THIRUTTUMTUMUTURUS009963747B2 (12 ) United States Patent ( 10 ) Patent No. : US 9 , 963, 747 B2 Bryant et al. (45 ) Date of Patent: *May 8 , 2018 (54 ) METHODS FOR THE IDENTIFICATION , ( 56 ) References Cited ASSESSMENT, AND TREATMENT OF PATIENTS WITH CANCER THERAPY U . S . PATENT DOCUMENTS 8 , 278, 038 B2 10 / 2012 Bryant et al. ( 71 ) Applicant: Millennium Pharmaceuticals , Inc. , 8 , 889 ,354 B2 11 /2014 Bryant et al. Cambridge , MA (US ) 2004 /0156854 A1 8 /2004 Mulligan et al. (72 ) Inventors : Barbara M . Bryant, Cambridge, MA FOREIGN PATENT DOCUMENTS (US ) ; Andrew I. Damokosh , West Hartford , CT (US ) ; George J . Wo WO 2003 / 053215 7 /2003 Mulligan , Lexington ,MA (US ) WO WO 04 /053066 A2 6 / 2004 WO WO 2006 / 133420 12 /2006 @( 73 ) Assignee : Millennium Pharmaceuticals , Inc ., Cambridge , MA (US ) OTHER PUBLICATIONS Adams, Julian , et al. , " Proteasome Inhibitors: A Novel Class of ( * ) Notice : Subject to any disclaimer, the term of this Potent and Effective Antitumor Agents , ” Cancer Research , vol. 59 patent is extended or adjusted under 35 ( Jun . 1 , 1999 ) pp . 2615 - 2622 . U . S .C . 154 (b ) by 0 days . days . Adams, Julian , “ Development of the Proteasome Inhibitor PS - 341, " The Oncologist , vol. 7 ( 2002 ) pp . 9 - 16 . This patent is subject to a terminal dis Lightcap , Eric S . , et al. , " Proteasome Inhibition Measurements: claimer . Clinical Application , ” Clinical Chemsitry , vol. 46 , No. 5 ( 2000 ) pp . 673 -683 . (21 ) Appl . No. : 14 /516 , 719 Hochwald , Steven N ., et al. , “ Antineoplastic Therapy in Colorectal Cancer through Proteasome Inhibition ,” The American Surgeon , ( 22 ) Filed : Oct . 17 , 2014 vol. 69 ( Jan . 2003) pp . 15 - 23 . Richardson , Paul G . , et al. , “ A Phase 2 Study of Bortezomib in Relapsed , Refractory Myeloma, ” The New England Journal of (65 ) Prior Publication Data Medicine , vol. 348 , No. 26 ( Jun . 26 , 2003 ) pp . 2609- 2617 . US 2015 /0252430 A1 Sep . 10 , 2015 Van de Vijver, Marc J. , et al. , “ A Gene - Expression Signature as a Predictor of Survival in Breast Cancer, " The New England Journal of Medicine , vol. 347 , No. 25 (Dec . 19 , 2002 ) pp . 1999 -2009 . Related U . S . Application Data Barden , Catherine B ., et al . , “ Classification of Follicular Thyroid Tumors by Molecular Signature : Results of Gene Profiling , ” Clini (63 ) Continuation of application No . 13 /600 ,260 , filed on cal Cancer Research , vol. 9 (May 2003 ) pp . 1792 - 1800 . Aug . 31 , 2012 , now Pat. No. 8 , 889, 354 , which is a Mulligan , George , et al. , “Gene expression profiling and correlation continuation of application No . 11 /449 , 195 , filed on with outcome in clinical trials of the proteasome inhibitor Jun . 8 , 2006 , now Pat. No. 8 ,278 ,038 . bortezomib ,” Blood , vol. 109 , No . 8 ( Apr. 15 , 2007 ) pp . 3177 - 3188 . Toczyski , David P ., et al. , “ The Epstein -Barr virus (EBV ) small RNA EBER1 binds and relocalizes ribosomal protein L22 in (60 ) Provisional application No. 60 /688 ,634 , filed on Jun . EBV -infected human B lymphocytes, ” Proceedings of the National 8 , 2005 . Academy of Science USA , vol . 91 ( Apr. 1994 ) pp . 3463 - 3467 . Maglott , Donna , et al. , “ Entrez Gene: gene -centered information at (51 ) Int . Cl. NCBI , ” Nucleic Acids Research , vol. 33 , Database issue ( 2005 ) pp . C120 1 /68 ( 2018 .01 ) D54 -D58 . GOIN 33 / 574 (2006 .01 ) (Continued ) G06F 19 / 20 ( 2011 .01 ) G06Q 50 / 22 ( 2018 .01 ) Primary Examiner — Sean E Aeder A61K 31 /69 ( 2006 .01 ) (74 ) Attorney , Agent, or Firm — Millennium G060 20 / 40 ( 2012 .01 ) Pharmaceuticals , Inc . G06F 19 /24 ( 2011. 01) U . S . CI. (57 ) ABSTRACT ??? ...... C12Q 1 /6886 (2013 .01 ); A61K 31/ 69 The present invention is directed to the identification of ( 2013 .01 ) ; GOIN 33 /57407 (2013 .01 ) ; GOIN predictive markers that can be used to determine whether 33 / 57415 (2013 .01 ) ; GOIN 33 / 57419 patients with cancer are clinically responsive or non - respon ( 2013 .01 ) ; GOIN 33 /57423 ( 2013 .01 ) ; GOIN sive to a therapeutic regimen prior to treatment. In particular , 33 /57426 (2013 .01 ) ; GOIN 33 / 57434 the present invention is directed to the use of certain ( 2013 . 01 ) ; GOIN 33 / 57438 ( 2013 . 01 ) ; GOIN individual and /or combinations of predictive markers, 33 /57449 (2013 . 01 ) ; G06F 19 /20 ( 2013 .01 ) ; wherein the expression of the predictive markers correlates G06Q 20 /40 (2013 .01 ) ; G06Q 50 / 22 with responsiveness or non - responsiveness to a therapeutic ( 2013 . 01 ) ; C120 2600 / 106 ( 2013 . 01 ); C120 regimen . Thus , by examining the expression levels of indi 2600 / 136 (2013 .01 ); C12Q 2600 /158 vidual predictive markers and /or predictive markers com ( 2013 .01 ) ; GOIN 2800 / 44 (2013 .01 ) ; GOOF prising a marker set, it is possible to determine whether a 19 /24 (2013 .01 ); YO2A 90 /22 (2018 . 01 ) ; YO2A therapeutic agent, or combination of agents , will be most 90 /26 (2018 .01 ) likely to reduce the growth rate of tumors in a clinical (58 ) Field of Classification Search setting. None See application file for complete search history . 19 Claims, No Drawings US 9 , 963, 747 B2 Page 2

References Cited Alberts , Bruce, et al. , Molecular Biology of the Cell, Third Edition , (56 ) published by Garland Publishing, Inc ., New York , New York ( 1994 ) p . 465 . OTHER PUBLICATIONS Greenbaum , Dov, et al. , “ Comparing protein abundance and mRNA Affymetrix , Inc ., “ Annotation Methodology, ” Affymetrix — Anno expression levels on a genomic scale , " Genome Biology , vol. 4 , No. 9 (2003 ) pp . 117 . 1 - 117. 8 . tation Methodology Technical Note [ online ] Affymetrix , Santa Lichtinghagen , Ralf, et al. , “ DifferentmRNA and protein expression Clara , CA , downloaded Jul. 23 , 2008 ( 3 pages) . of matrix metalloproteinases 2 and 9 and tissue inhibitor of metal Affymetrix , Inc ., “ GeneChip Human Genome U133 Set ,” [online ] loproteinases 1 in benign and malignant prostate tissue, ” European 701092 Rev . 3 ( Nov . 2004 ) downloaded Jul. 23 , 2008 ( 2 pages ) . Urology , vol. 42 ( 2002 ) pp . 398 - 406 . Affyrnetrix , Inc ., “ Human GenomeU133 Set - Support Materials , " Wessels , Lodewyk F . A . , et al. , “ A protocol for building and Affymetrix - Technical Support Documentation for Human evaluating predictors of disease state based on microarray data , " Genome U133 Set [ online ] downloaded Jul. 24 , 2008 ( 3 pages ) . Bioinformatics, vol . 21, No . 19 ( Apr. 7 , 2005 ) pp . 3755 - 3762 . International Preliminary Examination Report ( IPER ) and Written International Prelminary Examination Report (IPER ) and Written Opinion of the International Seaarching Authority dated Dec . 11 , Opinion of the International Searching Authority dated Dec . 11 , 2007 in corresponding PCT Application PCT/ US06 /022515 . 2007 in corresponding PCT Application PCT/ US06 / 022515 . Office Action dated Jun . 23. 2008 in co -pending U . S . Appl. No. Office Action dated Jun . 23 , 2008 in co - pending U . S . Appl. No. 10 / 728 ,055 . 10 / 728 ,055 . Maglott , Donna , et al. , “ Entrez Gene : gene - centered information at Supplementary Partial European Search Report dated Jun . 24 , 2009 NCBI, ” Nucleic Acids Research , vol. 33, Database issue (2005 ) pp . in European Patent Application No . 06784710 . 3 which corresponds D54 -D58 . to U . S . Appl. No. 11/ 449, 195 . Affymetrix , Inc ., “ Human Genome U133 Set - Support Materials ,” Glatt , Christine M ., et al ., “ Molecular characterization of thyroid Affymetrix — Technical Support Documentation for Human Genome U133 Set [online ] downloaded Jul. 24 , 2008 ( 3 pages ). toxicity : anchoring gene expression profiles to biochemical and Toczyski, David P. , et al. , “ The Epstein - Barr virus (EBV ) small pathologic end points ,” Environmental Health Perspectives , vol. RNA EBER1 binds and relocalizes ribosomal protein L22 in 113 , No . 10 ( Oct. 2005 ) pp . 1354 - 1361. EBV -infected human B lymphocytes, ” Proceedings of the National Wessels, Lodewyk F . A . , et al ., " A protocol for building and Academy of Science US, vol. 91 ( Apr. 1994 ) pp . 3463 - 3467 . evaluating predictors of disease state based on microarray data ,” Chauhan , Dharminder, et al. , “ Identification of genes regulated by Bioinfomatics, vol . 21, No . 19 ( Apr. 7 , 2005 ) pp . 3755 - 3762 . Dexamethasone in multiple myeloma cells using oligonucleotide Tockman , Melvyn S ., et al. , “ Considerations in bringing a cancer arrays, ” Oncogene , vol . 21 ( 2002 ) pp . 1346 - 1358 . biomarker to clinical application , ” Cancer Research (Supplement ) , International Search Report of the International Searching Authority vol. 52 (May 1 , 1992 ) pp . 27118 -27182 . dated Nov . 26 , 2007 issued in International Application No . PCT/ Ferrandina , Gabriella , et al. , “ Cyclooxygenase - 2 (Cox - 2 ) expres US06 /022515 , which corresponds to U . S . Appl. No . 11/ 449 , 195 . sion and resistance to platinum versus platinum / paclitaxel contain Search Report dated Jan . 2 , 2014 of co -pending European patent ing chemotherapy in advanced ovarian cancer, ” BMC Cancer , vol. application No . 13181515 . 1 6 No. 182 ( Jul. 11 , 2006 ) pp . 1 - 5 . Bohen et al. , “ Variation in Gene Expression Patterns in Follicular Chauhan , Dharminder, et al. , “ Identification of genes regulated by Lymphoma and the Response to Rituximab ” , PNAS, 2003 , vol. 100 , Dexamethasone in multiple myeloma cells using oligonudeotide No . 4 , pp . 1926 - 1930 . arrays ,” Oncogene , vol. 21 ( 2002 ) pp . 1346 - 1358 . Barlow , et al. , Predicting Event- Free and Overall Survival after Hideshima, Teru , et al. , “ The proteasome inhibitor PS -341 inhibits Treatment ofMyeloma with the Proteasome Inhibitor Bortezomib , growth , induces apoptosis , and overcomes drug resistance in human Blood , 2004 , vol. 104, Abstract 1481. multiple myeloma cells ,” Cancer Research , vol. 61 ( Apr. 1 , 2001) Barlogie , et al ., “ Treatment of Multiple Myeloma” , Blood , 2004 , pp . 3071 - 3076 . vol. 103 , pp . 20 -32 . US 9 , 963 , 747 B2 METHODS FOR THE IDENTIFICATION , regulation of genes involved in the immune and inflamma ASSESSMENT, AND TREATMENT OF tory responses. For example , Read et al. demonstrated that PATIENTS WITH CANCER THERAPY the ubiquitin -proteasome pathway is required for expression of cell adhesion molecules, such as E - selectin , ICAM - 1 , and CROSS REFERENCE TO RELATED 5 VCAM - 1 . See Immunity 2 :493 - 506 (1995 ) . Additional find APPLICATIONS ings further support the role for proteasome inhibition in cancer therapy, as Zetter found that cell adhesion molecules This application is a Continuation of U .S . patent appli - are involved in tumor metastasis and angiogenesis in vivo , cation Ser. No. 13 /600 , 260 , filed Aug . 31 , 2012 , now U . S . by directing the adhesion and extravastation of tumor cells Pat . No. 8 , 889, 354 , which is a Continuation of U . S . patent 10 to and from the vasculature to distant tissue sites within the application Ser. No. 11/ 449 , 195 , filed Jun . 8 , 2006 , now U . S . body . See , e . g ., Seminars in Cancer Biology 4 :219 - 229 Pat. No . 8 ,278 ,038 , which claims the benefit of U . S . Provi- ( 1993 ) . Moreover, Beg and Baltimore , found that NF -kB is sional Application No . 60 /688 ,634 , filed Jun . 8 , 2005 . The an anti -apoptotic factor, and inhibition of NF -kB activation entire contents of each of these applications are incorporated makes cells more sensitive to environmental stress and herein by reference . 15 cytotoxic agents . See Science 274 :782 (1996 ). The contents of the Sequence Listing were submitted on The first proteasome inhibitor described as having anti compact disc in the parent U . S . patent application Ser. No . tumor activity , bortezomib (N - pyrazinecarbonyl - L - phenyl 11 / 449, 195 and are being transferred to this application . The alanine - L - leucineboronic acid , PS -341 ) (VELCADE® for compact disc has a copy of the Sequence Listing file , created injection , Millennium Pharmaceuticals , Inc . , Cambridge , on Sep . 26 , 2008 and named “ sequence listing . txt, ” the 20 Mass .; Johnson & Johnson Pharmaceutical Research and contents of which are incorporated herein by this reference . Development L . L . C . ) has been approved for treatment of This file is 5 . 91 MB (6 ,203 , 392 bytes ) and was copied onto relapsed multiple myeloma. Presently clinical trials are compact disc on Sep . 30 , 2008 . underway in additional indications, including additional hematological cancers as well as solid tumors . This and BACKGROUND OF THE INVENTION 25 other peptide boronic ester and acid proteasome inhibitors have been described by Adams et al. See , e . g . , U . S . Pat. No . One of the continued problems with therapy in cancer 5 ,780 ,454 (1998 ), U .S . Pat . No . 6 , 066 , 730 ( 2000 ) , and U . S . patients is individual differences in response to therapies . Pat. No. 6 ,083 , 903 ( 2000 ) . They describe the use of the With the narrow therapeutic index and the toxic potential of disclosed boronic ester and boronic acid compounds to many available cancer therapies , such differential responses 30 reduce the rate of muscle protein degradation , to reduce the potentially contribute to patients undergoing unnecessary activity of NF -kB in a cell, to reduce the rate of degradation ineffective and even potentially harmful therapy regimens. If of p53 protein in a cell, to inhibit cyclin degradation in a cell , a designed therapy could be optimized to treat individual to inhibit the growth of a cancer cell, and to inhibit NF -kB patients, such situations could be reduced or even elimi- dependent cell adhesion . nated . Furthermore, targeted designed therapy may provide 35 Bortezomib specifically and selectively inhibits the pro more focused , successful patient therapy overall . Accord - teasome by binding tightly (Ki = 0 . 6 nM ) to one of the ingly , there is a need to identify particular cancer patients enzyme' s active sites . Bortezomib is selectively cytotoxic , which are particularly responsive to particular cancer thera - and has a novel pattern of cytotoxicity in National Cancer pies, either alone or in combination with other chemothera - Institute (NCI ) in vitro and in vivo assays . Adams J , et al . pies. It would therefore be beneficial to provide for the 40 Cancer Res 59 : 2615 - 22 . ( 1999) . In addition , bortezomib has diagnosis , staging , prognosis , and monitoring of cancer cytotoxic activity in a variety of xenograft tumor models . patients , including , e . g . , hematological cancer patients ( e . g ., Teicher B A , et al. Clin Cancer Res. 5 : 2638 - 45 ( 1999 ) . multiple myeloma, leukemias, lymphoma , etc ) as well as Bortezomib inhibits nuclear factor- kB (NF -KB ) activation , solid tumor cancer patients ( e . g ., lung , breast, prostate , attenuates interleukin - 6 ( IL - 6 ) mediated cell growth , and has ovary , colon , kidney , liver ) , who would benefit from par - 45 a direct apoptotic effect, and possibly an anti- angiogenic ticular cancer inhibition therapies ; or to indicate a predis - effect. Additionally , bortezomib is directly cytotoxic to position of such patients to non -responsiveness to therapy, myeloma cells in culture , independent of their p53 status. thus resulting in appropriate preventative measures . See , e .g . , Hideshima T , et al. Cancer Res. 61 :3071 -6 (2001 ) . Proteasome inhibition represents an important strategy in In addition to a direct cytotoxic effect of bortezomib on cancer treatment. The proteasome is a multi -enzyme com - 50 myeloma cells, bortezomib inhibits tumor necrosis factor plex present in all cells which play a role in degradation of alpha ( TNFa stimulated intercellular adhesion molecule - 1 proteins involved in regulation of the cell cycle . For ( ICAM - 1 ) expression by myeloma cells and ICAM - 1 and example, King et al. , demonstrated that the ubiquitin - pro - vascular cell adhesion molecule - 1 (VCAM - 1 ) expression on teasome pathway plays an essential role in regulating cell bone marrow stromal cells (BMSCs ) , resulting in decreased cycle , neoplastic growth and metastasis . A number of key 55 adherence of myeloma cells and , consequently , in decreased regulatory proteins, including p53, cyclins, and the cyclin cytokine secretion . Hideshima T , et al. Oncogene . 20 :4519 dependent kinases p21 and p27Kipi, are temporally degraded 27 (2001 ) . By inhibiting interactions ofmyeloma cells with during the cell cycle by the ubiquitin - proteasome pathway . the surrounding bone marrow , bortezomib can inhibit tumor The ordered degradation of these proteins is required for the growth and survival, as well as angiogenesis and tumor cell cell to progress through the cell cycle and to undergo 60 migration . The antineoplastic effect of bortezomib may mitosis . See , e . g . , Science 274 : 1652 - 1659 ( 1996 ) . Further involve several distinctmechanisms , including inhibition of more , the ubiquitin -proteasome pathway is required for cell growth signaling pathways, dysregulation of the cell transcriptional regulation . Palombella et al. , teach that the cycle , induction of apoptosis , and inhibition of cellular activation of the transcription factor NF - kB is regulated by adhesion molecule expression . Notably , bortezomib induces proteasome-mediated degradation of the inhibitor protein 65 apoptosis in cells that over express B - cell lymphoma 2 IkB . See International Patent Application Publication No. (Bcl - 2 ) , a genetic trait that confers unregulated growth and WO 95 /25533 . In turn , NF -kB plays a central role in the resistance to conventional chemotherapeutics. McConkey D US 9 ,963 , 747 B2 J , et al. The proteasome as a new drug target in metastatic tion : 1 ) methods and compositions for determining whether prostate cancer. 7th Annual Genitourinary Oncology Con - a proteasome inhibition therapy and /or a glucocorticoid ference , Houston , Tex . Abstract ( 1999 ) . therapy will or will not be effective in stopping or slowing Glucocorticoidal steroids are capable of causing apoptotic tumor growth and patient treatment ; 2) methods and com death of many varieties of cells , and a selection of gluco - 5 positions for monitoring the effectiveness of a proteasome corticoidal steroids have consequently be used in the treat - inhibition therapy ( a proteasome inhibitor agent or a com ment of various malignancies, including lymphoid malig bination of agents ) and / or a glucocorticoid therapy used for nancies, and combination therapies in solid tumors . For the treatment of tumors ; 3 ) methods and compositions for example, the optimal therapy for relapsed myeloma is not treatments of tumors comprising proteasome inhibition established , but high - dose dexamethasone is commonly 10 therapy and / or glucocorticoid therapy ; and 4 ) methods and used . See , e . g . , Kumar A , et al. Lancet Oncol; 4 : 293 -304 compositions for identifying specific therapeutic agents and ( 2003 ) ; Alexanian R , et al. Ann Intern Med . 105 : 8 - 11 ( 1986 ) ; combinations of therapeutic agents that are effective for the Friedenberg W R , et al . Am J Hematol. 36 : 171 -75 . ( 1991 ) . treatment of tumors in specific patients . Response rates with this treatment are similar to those with The markers of the present invention , whose expression vincristine, doxorubicin , and dexamethasone ( VAD ) , and the 15 correlates with the response to an agent, are identified in dexamethasone component is estimated to account for 85 Table 1A , Table 1B , Table 2A , Table 2B , and Table 3 . By percent of the effect of VAD . See , e . g ., Alexanian R , et al. examining the expression of one or more of the identified Blood . 80 :887 - 90 (1992 ) ; Sonneveld P , et al. Br J Haematol. markers ormarker sets in a tumor, it is possible to determine 115 :895 - 902 . (2001 ) . High - dose chemotherapy followed by which therapeutic agent or combination of agents will be autologous stem cell transplantation improves patient sur- 20 most likely to reduce the growth rate of the cancer cells . By vival , but in most cases the disease relapses. Attal M et al. examining the expression of one or more of the identified N Engl J Med . 335 : 91 - 97 ( 1996 ) ; Child JA , et al. N Engl J markers or marker sets in a cancer, it is also possible to Med . 348 : 1875 - 83 ( 2003) . determine which therapeutic agent or combination of agents In addition to use of dexamethasone , additional corticos - will be the least likely to reduce the growth rate of cancer teroids have demonstrated use in cancer treatments , includ - 25 cells . By examining the expression of one or more of the ing hydrocortisone in combination therapy for prostate can identified markers or marker sets , it is therefore possible to cer , predisolone in leukemia , prednisolone in lymphoma eliminate ineffective or inappropriate therapeutic agents treatment, and triamcinolone has recently demonstrated Importantly , these determinations can be made on a patient some anti -cancer activity . See , e . g . , Scholz M ., et al. , J Urol. by patient basis or on an agent by agent basis . Thus, one can 173: 1947 - 52 . ( 2005 ) ; Sano J. , et al , Res Vet Sci. (May 10 , 30 determine whether or not a particular therapeutic regimen is 005 ); Zinzani PL . et al ., Semin Oncol. 32 ( 1 Suppl 1 ) :S4 - 10 . likely to benefit a particular patient or type of patient , and / or ( 2005 ) ; and Abrams, M T et al. , J Cancer Res Clin Oncol. whether a particular regimen should be continued . 131: 347 -54 (2005 ). It is believed gene transcription result The present invention is directed to methods of identify ing from treatment with glucocorticoids results in apoptotic ing and /or selecting a cancer patient who is responsive to a death and therapeutic effect. Analysis of sensitive and resis - 35 therapeutic regimen . In particular, the methods are directed tant cell lines have demonstrated differential gene expres - to identifying or selecting a cancer patient who is responsive sion patterns , suggesting expression differences account for to a therapeutic regimen comprising proteasome inhibition varied response rates to glucocorticoid therapy . See , e . g ., therapy and /or glucocorticoid therapy . Additionally pro Thompson , E . B ., et al. , Lipids. 39 :821 - 5 ( 2004 ) , and refer vided are methods of identifying a patient who is non ences cited therein . 40 responsive to such a therapeutic regimen . These methods While advances in development of successful cancer typically include the determining the level of expression of therapies progress, individual patient responses continue to one or more predictive markers in a patient ' s tumor ( e . g ., a demonstrate subsets of patient response to any particular patient ' s cancer cells ) , comparing the level of expression to therapy. We have conducted gene expression analysis stud - reference expression level, and identifying whether ies to assess patient populations undergoing glucocorticoid 45 expression in the sample includes a pattern or profile of therapy or proteasome inhibition therapy . Analyses werewere expression of a selected predictive marker or marker set carried out to identify predictive markers associated with which corresponds to response or non -response to protea particular patients who respond well to treatment ( respond - some inhibition therapy and /or glucocorticoid therapy . ers ) with a glucocorticoid and /or proteasome inhibitor ver Additionally provided methods include therapeutic meth sus those patients who do not respond to treatment (non - 50 ods which further include the step of beginning , continuing , responders ) with a glucocorticoid and / or proteasome or commencing, or stopping, discontinuing or halting a inhibitor. therapy accordingly where a patient' s predictive marker profile indicates that the patient would respond or not DESCRIPTION OF THE INVENTION respond to the proteasome inhibition and /or glucocorticoid 55 therapeutic regimen . In another aspect, methods are pro The present invention is based , in part, on the identifica - vided for analysis of a patient not yet being treated with a tion of individual markers and marker sets that can be used proteasome inhibition therapy or glucocorticoid therapy and to determine whether a tumor may be effectively treated by identification and prediction that the patient would not be a treatment with a proteasome inhibition therapy and / or a responder to the therapeutic agent and such patient should glucocorticoid therapy. For example , the compositions and 60 not be treated with the proteasome inhibition therapy and /or methods provided herein can be used to determine whether glucocorticoid therapy when the patient ' s marker profile a patient will be responsive or non - responsive to a protea - indicates that the patient is a non - responder . Thus, the some inhibition therapeutic agent. Furthermore the compo - provided methods of the invention can eliminate ineffective sitions and methods provided herein can be used to deter - or inappropriate use of proteasome inhibition therapy and /or mine whether a patient will be responsive or non - responsive 65 glucocorticoid therapy regimens . to a glucocorticoid therapeutic agent . Based on these iden - Additionally provided are classifiers which can be used to tifications, the present invention provides, without limita - develop a diagnostic test or a readable array useful for US 9 , 963, 747 B2 identifying patients who will be responsive or non - respon - Clara , Calif. ) , also hereby are incorporated by reference . In sive to proteasome inhibition therapy and/ or glucocorticoid the case of conflict, the present specification , including therapy . Probes or peptides identified in a classifier of the definitions, will control . invention can be included in a diagnostic or prognostic test The articles “ a ” and “ an ” are used herein to refer to one to select a therapy , e . g . , proteasome inhibition therapy 5 or to more than one ( i. e . at least one of the grammatical and / or glucocorticoid therapy or a test which is used to object of the article . By way of example , “ an element" determine continuation of therapy, e . g ., proteasome inhibi means at least one element and can include more than one tion therapy and / or glucocorticoid therapy . Additional methods include methods to determine the element. activity of an agent, the efficacy of an agent, or identify new 10 A “marker " is a naturally - occurring polymer correspond therapeutic agents or combinations. Such methods include ing to at least one of the nucleic acids or proteins associated methods to identify an agent useful as a proteasome inhibitor with Affymetrix probe set identifiers listed in any one of and / or a glucocorticoid inhibitor, for treating a cancer, e . g . Table 1A , Table 1B , Table 2A , Table 2B , and Table 3 . For a hematological cancer ( e . g . , multiple myeloma, leukemias , example, markers include , without limitation , sequences lymphoma, etc ) or cancer from a solid tumor ( e. g ., in lung, 15 recognized by the Affymetric probes and probeset identifi breast, prostate , ovary, colon , kidney or liver ), based on its ers , sense and anti - sense strands of genomic DNA ( i. e . ability to affect the expression ofmarkers in a marker set of including any introns occurring therein ), RNA generated by the invention . For example , an inhibitor which decreases or transcription of genomic DNA (i .e . prior to splicing ), RNA increases the level of expression of a marker or markers generated by splicing of RNA transcribed from genomic provided as upregulated or downregulated , respectively , in a 20 DNA , and proteins generated by translation of spliced RNA set predictive for responsiveness to proteasome inhibition of ( i . e . including proteins both before and after cleavage of the cancer would be a candidate inhibitor for the cancer. In normally cleaved regions such as transmembrane signal another example , an inhibitor which decreases or increases sequences ) . As used herein , a " marker ” may also include a the level of expression of a marker or markers provided as cDNA made by reverse transcription of an RNA generated upregulated or downregulated , respectively, in a set predic - 25 by transcription of genomic DNA ( including spliced RNA ) . tive for responsiveness to glucocorticoid inhibition of the A “ marker set” is a group of markers , comprising two or cancer would be a candidate inhibitor for the cancer . more predictive markers of the invention . Markers of the The present invention is also directed to methods of treating a cancer patient, with a therapeutic regimen , in present invention include the predictive markers identified in particular a proteasome inhibitor therapy ( e .g ., a proteasome 30 Table 1A , Table 1B , Table 2A , Table 2B , and Table 3 ; as inhibitor agent, alone , or in combination with an additional identified by the particular probeset identifier, representative agent such as a chemotherapeutic agent ) and /or glucocorti public identifier , title , gene symbol, and / or Entrez gene coid therapy regimen ( a glucocorticoid agent, alone or in identifier, and include the representative nucleotide and /or combination with an additional agent) , which includes the protein sequence or fragment thereof which corresponds to step of selecting a patient whose predictive marker profile 35 me indicates that the patient will respond to the therapeutic A “ predictive marker ” as used herein , includes a marker regimen , and treating the patient with the proteasome inhi which has been identified as having differential expression bition therapy and /or glucocorticoid therapy . in tumor cells of a patient and furthermore that expression is Additional methods include selecting patients that are characteristic of a patient who is responsive in either a unlikely to experience response or increased time to pro - 40 positive or negative manner to treatment with a proteasome gression upon treatment with a cancer therapy ( e . g ., protea inhibitor regimen and/ or glucocorticoid regimen . For some inhibition therapy , glucocorticoid therapy ) . Further example , a predictive marker includes a marker which is more provided are methods for selection of a patient having demonstrates higher expression in a non - responsive patient ; aggressive disease and more rapid time to progression . alternatively a predictive marker includes a marker which Additional methods include a method to evaluate whether 45 demonstrates higher expression in a responsive patient. to treat or pay for the treatment of cancer, e . g . hematological Similarly , a predictive marker is intended to include those cancer ( e . g ., multiple myeloma , leukemias , lymphoma, etc ) markers which demonstrate lower expression in a non or cancer from a solid tumor ( e . g . , in lung , breast , prostate , responsive patient as well as those markers which demon ovary, colon , kidney or liver ), by reviewing a patient' s strate lower expression in a responsive patient. Thus , as used predictive marker profile for responsiveness or non - respon - 50 herein , predictive marker is intended to include each and siveness to proteasome inhibition and/ or glucococorticoid every one of these possibilities, and further can include each therapy. single marker individually as a predictive marker ; or alter Definitions natively can include one or more , or all of the characteristics Unless otherwise defined , all technical and scientific collectively when reference is made to " predictive markers " terms used herein have the same meaning as commonly 55 or " predictive marker sets. ” A predictive marker set also can understood by one of ordinary skill in the art to which this be known as a “ classifier. ” invention belongs. Although methods and materials similar As used herein , a “ naturally occurring” refers to a mol or equivalent to those described herein can be used in the ecule ( e . g . , RNA , DNA , protein , etc . ) that occurs in nature practice or testing of the present invention , preferred meth - ( e . g . encodes a natural protein , a naturally produced protein , ods and materials are described herein . The content of all 60 etc ) . database accession records ( e . g ., representative public iden - The term “ probe ” refers to any molecule which is capable tifier ID from Affymetrix HG133 annotation files, Entrez , of selectively binding to a specifically intended target mol GenBank , RefSeq ) cited throughout this application ( includ ecule , for example a marker of the invention . Probes can be ing the Tables ) are also hereby incorporated by reference . either synthesized by one skilled in the art, or derived from The contents of files disclosing the Affymetrix HG - 133A 65 appropriate biological preparations. For purposes of detec Probe Sequences and HG - 133B Probe Sequences , both tion of the target molecule , probes may be specifically FASTA files dated Jun . 9 , 2003 ( Affymetrix , Inc. , Santa designed to be labeled , as described herein . Examples of US 9 , 963, 747 B2 molecules that can be utilized as probes include , but are not a responsive patient, as described herein . Expression of that limited to , RNA , DNA , proteins, antibodies, and organic marker below a given threshold ( e . g . , below an informative monomers. level) may be indicative of a non - responsive patient The “ normal ” level of expression of a marker is the level A cancer or tumor is treated or diagnosed according to the of expression of the marker in cells in a similar environment 5 present methods. “ Cancer ” or “ tumor” is intended to include or response situation , in a patient not afflicted with cancer. any neoplastic growth in a patient, including an inititial A normal level of expression of a marker may also refer to tumor and any metastases. The cancer can be of the liquid or the level of expression of a “ reference sample ” , ( e . g . , sample solid tumor type. Liquid tumors include tumors of hemato from a healthy subjects not having the marker associated logical origin , including , e . g . , myelomas ( e . g . , multiple disease ) . A reference sample expression may be comprised 10 myeloma) , leukemias ( e . g ., Waldenstrom ' s syndrome, of an expression level of one or more markers from a chronic lymphocytic leukemia , other leukemias ) , and lym reference database . Alternatively , a " normal " level of phomas (eg , B - cell lymphomas , non -Hodgkins lymphoma ) . expression of a marker is the level of expression of the Solid tumors can originate in organs , and include cancers marker in non -tumor cells in a similar environment or such as lung , breast, prostate , ovary , colon , kidney , and liver . response situation from the same patient that the tumor is 15 As used herein , cancer cells , including tumor cells, refer to derived from . cells that divide at an abnormal ( increased ) rate . Cancer cells “ Differential expression ” of a marker refers to expression include , but are not limited to , carcinomas , such as of a marker that varies in level across patients . Furthermore , squamous cell carcinoma, basal cell carcinoma, sweat gland in this invention we refer to a marker as “ differentially carcinoma, sebaceous gland carcinoma, adenocarcinoma, expressed ” when its expression level is correlated with , or 20 papillary carcinoma, papillary adenocarcinoma, cystadeno otherwise indicative of, response or non - response to treat carcinoma, medullary carcinoma, undifferentiated carci ment. noma , bronchogenic carcinoma , melanoma , renal cell car " Complementary ” refers to the broad concept of sequence cinoma, hepatoma- liver cell carcinoma, bile duct carcinoma , complementarity between regions of two nucleic acid cholangiocarcinoma, papillary carcinoma, transitional cell strands or between two regions of the same nucleic acid 25 carcinoma , choriocarcinoma , semonoma, embryonal carci strand. It is known that an adenine residue of a first nucleic noma, mammary carcinomas, gastrointestinal carcinoma, acid region is capable of forming specific hydrogen bonds colonic carcinomas , bladder carcinoma, prostate carcinoma, ( “ base pairing” ) with a residue of a second nucleic acid and squamous cell carcinoma of the neck and head region ; region which is antiparallel to the first region if the residue sarcomas , such as fibrosarcoma, myxosarcoma, liposar is thymine or uracil. Similarly , it is known that a cytosine 30 coma, chondrosarcoma, osteogenic sarcoma, chordosar residue of a first nucleic acid strand is capable of base coma , angiosarcoma, endotheliosarcoma, lymphangiosar pairing with a residue of a second nucleic acid strand which coma, synoviosarcoma and mesotheliosarcoma ; is antiparallel to the first strand if the residue is guanine . A hematologic cancers , such as myelomas, leukemias ( e . g . , first region of a nucleic acid is complementary to a second acute myelogenous leukemia , chronic lymphocytic leuke region of the same or a different nucleic acid if , when the 35 mia , granulocytic leukemia , monocytic leukemia , lympho two regions are arranged in an antiparallel fashion , at least cytic leukemia ) , and lymphomas ( e . g . , follicular lymphoma, one nucleotide residue of the first region is capable of base mantle cell lymphoma, diffuse large Bcell lymphoma, pairing with a residue of the second region . Preferably , the malignant lymphoma, plasmocytoma, reticulum cell sar first region comprises a first portion and the second region coma, or Hodgkins disease ) ; and tumors of the nervous comprises a second portion , whereby, when the first and 40 system including glioma , meningoma, medulloblastoma , second portions are arranged in an antiparallel fashion , at schwannoma or epidymoma . least about 50 % , and preferably at least about 75 % , at least cancer is " responsive " to a therapeutic agent if its rate about 90 % , or at least about 95 % of the nucleotide residues of growth is inhibited as a result of contact with the of the first portion are capable of base pairing with nucleo - therapeutic agent, compared to its growth in the absence of tide residues in the second portion . More preferably , all 45 contact with the therapeutic agent. Growth of a cancer can nucleotide residues of the first portion are capable of base be measured in a variety of ways, for instance , the size of a pairing with nucleotide residues in the second portion . tumor or the expression of tumor markers appropriate for As used herein , “ informative ” expression is intended to that tumor type may bemeasured . For example , the response refer to the expression level of a differentially expressed definitions used to identify markers associated with predictive marker which corresponds to responsiveness or 50 myeloma and its response to proteasome inhibition therapy non - responsiveness . The expression level of a marker in a and /or glucocorticoid therapy , the Southwestern Oncology patient is “ informative ” if it is greater than a reference level Group (SWOG ) criteria as described in Blade et al. , Br J by an amount greater than the standard error of the assay Haematol. 1998 September; 102 ( 5 ) : 1115 - 23 were used employed to assess expression . Alternatively, a marker that (also see e . g . , Table C ) . These criteria define the type of is differentially expressed will have typical ranges of expres - 55 response measured in myeloma and also the characterization sion level that are predictive of responsiveness or non - of time to disease progression which is another important responsiveness. An informative expression level is a level measure of a tumor ' s sensitivity to a therapeutic agent. The that falls within the responsive or non - responsive range of quality of being responsive to a proteasome inhibition expressions. Still further , a set of markers may together be therapy and /or glucocorticoid therapy is a variable one , with “ informative ” if the combination of their expression levels 60 different cancers exhibiting different levels of " responsive either meets or is above or below a pre - determined score for ness ” to a given therapeutic agent, under different condi a predictive marker set as determined by methods provided tions . Still further, measures of responsiveness can be herein. assessed using additional criteria beyond growth size of a A given marker may be indicative of both responsive and tumor , including patient quality of life , degree of metastases , non - responsive patients ; for example , expression of a pre - 65 etc . In addition , clinical prognostic markers and variables dictive marker provided herein above a given threshold can be assessed ( e . g . , M protein in myeloma, PSA levels in ( e. g ., an informative expression level ) may be indicative of prostate cancer) in applicable situations . US 9 , 963 , 747 B2 10 A cancer is “ non - responsive ” to a therapeutic agent if its lymphomas , non - Hodgkins lymphoma) . Solid tumors can rate of growth is not inhibited , or inhibited to a very low originate in organs, and include cancers such as lung , breast, degree , as a result of contact with the therapeutic agent when prostate , ovary , colon , kidney , and liver . compared to its growth in the absence of contact with the The invention provides methods for determining or therapeutic agent . As stated above , growth of a cancer can be 5 assessing an appropriate cancer therapy regimen for treating measured in a variety of ways, for instance , the size of a a tumor in a patient. The cancer therapy regimens appro tumor or the expression of tumor markers appropriate for priate for use in or in conjunction with the provided methods that tumor type may be measured . For example , the response comprise proteasome inhibition therapy and / or glucocorti definitions used to identify markers associated with non - coid therapy. For example , proteasome inhibitor therapy response of multiple myeloma to therapeutic agents, the 10 comprises treatment of a patient with a proteasome inhibitor Southwestern Oncology Group ( SWOG ) criteria as (e . g ., bortezomib , or any other proteasome inhibitor described in Blade et . al . were used in the experiments described in further detail herein ) , alone or in combination described herein . The quality of being non -responsive to a with one or more additional agents . In another example , therapeutic agent is a highly variable one , with different glucocorticoid therapy comprises treatment of a patient with cancers exhibiting different levels of " non -responsiveness ” 15 a glucocorticoid ( e . g . , dexamethasone , or any other gluco to a given therapeutic agent, under different conditions. Still corticoid described in further detail herein ) , alone or in further , measures of non -responsiveness can be assessed combination with one or more additional agents . using additional criteria beyond growth size of a tumor , The provided methods comprise measuring the level of including patient quality of life , degree ofmetastases , etc . In expression of at least one predictive marker in the patient' s addition , clinical prognostic markers and variables can be 20 tumor and determining a cancer therapy regimen for treating assessed ( e . g . , M protein in myeloma, PSA levels in prostate the tumor based on the expression level of the predictive cancer ) in applicable situations. marker or markers , as relevant. An informative expression " Treatment” shall mean preventing or inhibiting further level of a predictive marker or markers in the patient sample tumor growth , as well as causing shrinkage of a tumor. is an indication that the patient is a responsive patient and Treatment is also intended to include prevention of metas - 25 would benefit from proteasome inhibition therapy and/ or tasis of tumor. A tumor is “ inhibited ” or “ treated ” if at least glucocorticoid therapy when the predictive marker or one symptom (as determined by responsiveness /non - respon marker set provided herein indicate such responsiveness. siveness , time to progression , or indicators known in the art Additionally , an informative expression level of a predictive and described herein ) of the cancer or tumor is alleviated , momarker or markers in a patient is an indication that the terminated , slowed , minimized , or prevented . Any amelio - 30 patient is a non -responsive patient and would not benefit ration of any symptom , physical or otherwise , of a tumor from proteasome inhibition therapy and / or glucocorticoid pursuant to treatment using a therapeutic regimen ( e . g ., therapy when the marker or markers provided herein indi proteasome inhibition regimen , glucocorticoid regimen ) as cate such non -responsiveness . further described herein , is within the scope of the invention . The invention further provides methods for determining As used herein , the term " agent” is defined broadly as 35 whether a patient will be responsive to a cancer therapy anything that cancer cells , including tumor cells , may be regimen for treating a tumor . Such methods comprise mea exposed to in a therapeutic protocol. In the context of the suring the level of expression of at least one predictive present invention , such agents include, but are not limited to , marker in the patient ' s tumor and determining a proteasome proteasome inhibition agents , glucocorticoidal steroid inhibition based regimen and /or glucocorticoid based regi agents , as well as chemotherapeutic agents as known in the 40 men for treating the tumor based on the expression level of art and described in further detail herein . the predictive marker or marker set. An informative expres A “ kit ” is any article of manufacture ( e . g . a package or s ion level of a predictive marker in the patient sample is an container ) comprising at least one reagent, e . g . a probe , for indication that the patient is a responsive patient and would specifically detecting a marker or marker set of the inven benefit from proteasome inhibition and / or glucocorticoid tion . The article of manufacture may be promoted , distrib - 45 therapy . An informative expression level of a predictive uted , or sold as a unit for performing the methods of the marker set in the patient is an indication that the patient is present invention . The reagents included in such a kit a responsive patient and would benefit from proteasome comprise probes /primers and / or antibodies for use in detect - inhibition therapy and / or glucocorticoid therapy when the ing responsive and non -predictive marker expression . In marker or markers provided herein indicate such respon addition , the kits of the present invention may preferably 50 siveness . Selected predictive markers for use in themethods contain instructions which describe a suitable detection comprise predictive markers which demonstrate increased assay . Such kits can be conveniently used , e . g ., in clinical expression in responsive patients and/ or longer time to settings , to diagnose and evaluate patients exhibiting symp - disease progression . toms of cancer, in particular patients exhibiting the possible The invention provides methods for determining whether presence of an a cancer capable of treatment with protea - 55 a patient has aggressive disease and will progress in disease some inhibition therapy and /or glucocorticoid therapy, faster than a patient not demonstrating aggressive disease. A including , e . g . , hematological cancers e . g . , myelomas ( e . g . , patient indicative of having aggressive disease also may be multiple myeloma) , lymphomas ( e . g . , non -hodgkins lym - non - responsive to a cancer therapy regimen for treating a phoma) , leukemias, and solid tumors ( e . g . , lung , breast, tumor. Such methods comprise measuring the level of ovarian , etc . ) . 60 expression of at least one predictive marker in the patient ' s The present methods and compositions are designed for tumor and identifying the patient as having aggressive use in diagnostics and therapeutics for a patient suffering disease based on the expression level of the predictive from cancer . The cancer can be of the liquid or solid tumor marker or marker set . An informative expression level of a type . Liquid tumors include tumors of hematological origin , predictive marker in the patient sample is an indication that including, e . g . , myelomas ( e . g . , multiple myeloma) , leuke - 65 the patient has aggressive disease patient and is likely to mias (e . g ., Waldenstrom 's syndrome, chronic lymphocytic progress and may not benefit from proteasome inhibition leukemia , other leukemias) , and lymphomas ( e . g . , B - cell based regimen and /or glucocorticoid based regimen therapy . US 9 , 963 , 747 B2 12 An informative expression level of a predictive marker set in ( e . g . , multiple myeloma , leukemias, lymphoma, etc ) or the patient is an indication that the patient is a patient having cancer from a solid tumor ( e . g ., in lung , breast , prostate , aggressive disease and would not benefit from proteasome ovary , colon , kidney or liver ) . In some embodiments , the inhibition based regimen and / or glucocorticoid based regi - predictive marker set includes a plurality of markers listed in men when the selected marker or marker set provided herein 5 Table 1A , Table 1B , Table 2A , Table 2B , or Table 3 . In some indicate such disease aggressiveness . Selected predictive embodiments the predictive marker set includes at least markers for use in the methods comprise predictive markers about 1 % , about 5 % , about 10 % , about 20 % , about 30 % , which demonstrate increased expression in non -responsive about 40 % , about 50 % , about 60 % , about 70 % , about 80 % , patients and /or shorter time to disease progression in about 90 % , about 95 % , about 96 % , about 97 % , about 98 % , patients and are not specific to treatment with proteasome 10 or about 99 % of the markers listed in Table 1B , Table 2A , inhibition therapy or glucocorticoid therapy . Table 2B , or Table 3 . Selected predictive marker sets can be Still further , the invention provides methods for deter - assembled from the predictive markers provided usingmeth mining whether a patient will be non - responsive to a cancer o ds provided herein and analogous methods known in the therapy regimen for treating a tumor. Such methods com art . An exemplary predictive marker sets is provided in prise measuring the level of expression of at least one 15 Table 4 . In certain aspects , the markers comprise those set predictive marker in the patient' s tumor and determining a forth in Table 4 . proteasome inhibition based regimen and /or glucocorticoid Methods of the invention further provide the ability to based regimen for treating the tumor based on the expression construct marker sets from the individual predictive markers level of the predictive marker or marker set . An informative set forth in Table 1A , Table 1B , Table 2A , Table 2B , and expression level of a predictive marker in the patient sample 20 Table 3 using the methods described in further detail herein . is an indication that the patient is a non -responsive patient In a further aspect , more than one marker set can be used in and would not benefit from proteasome inhibition based combination for the diagnostic , prognostic and treatment regimen and /or glucocorticoid based regimen therapy. An methods provided . informative expression level of a predictive marker set in the The methods of the invention can be performed such that patient is an indication that the patient is a non - responsive 25 determination of the level of expression of a predictive patient and would not benefit from proteasome inhibition marker is measured prior to tumor therapy in order to based regimen and /or glucocorticoid based regimen when identify whether the patient will be responsive to a protea the selected marker or marker set provided herein indicate some inhibition therapy and /or glucocorticoid therapy . such non - responsiveness. Selected predictive markers for In addition , the methods of the invention can be per use in the methods comprise predictive markers which 30 formed concurrently with ongoing tumor therapy to deter demonstrate increased expression in non -responsive patients mine if the patient is either responding to present protea and / or shorter time to disease progression . some inhibition therapy and / or glucocorticoid therapy or The invention further provides methods for treating a will respond to additional therapy comprising proteasome tumor in a patient with a proteasome inhibition based inhibition therapy and /or glucocorticoid therapy . regimen and / or glucocorticoid based regimen therapy . Such 35 Still further, the methods of the invention can be per therapeutic methods comprise measuring the level of expres - formed after tumor therapy has been carried out in order to sion of at least one predictive marker in a patient' s tumor ; assess whether the patient will be responsive to future course determining whether a proteasome inhibition based regimen of proteasome inhibition therapy and / or glucocorticoid and / or glucocorticoid based regimen for treating the tumor therapy . is appropriate based on the expression level of the predictive 40 Whether the methods are performed during ongoing marker or markers , and treating a patient with a proteasome tumor therapy or after a course of tumor therapy, the tumor inhibition based therapy and/ or glucocorticoid based therapy therapy can comprise proteasome inhibition therapy and/ or when the patient ' s expression level indicates a responsive glucocorticoid therapy , alone or alternative forms of cancer patient. An informative expression level of predictive therapy . The methods provided are designed to determine if marker in the patient sample is an indication that the patient 45 the patient will benefit from additional or future proteasome is a responsive patient and would benefit from proteasome inhibition and / or glucocorticoid therapy, and can include inhibition based regimen and / or glucocorticoid based regi - such proteasome inhibition and / or glucocorticoid therapy men therapy when the predictive marker or marker set alone or in combination with additional therapeutic agents . provided herein indicate the patient is a responsive patient. In certain aspects , the level of expression of predictive Methods of the invention use at least one of the predictive 50 marker in the patient' s tumor is measured by isolating a markers set forth in any one of Table 1A , Table 1B , Table sample of the tumor and performing analysis on the isolated 2A , Table 2B , and Table 3 . Additionally , the methods sample , or a portion thereof. In another aspect, the level of provided can use two , three , four, five , six , or more markers expression of predictive marker in the patient' s tumor is to form a predictive marker set. For example , marker sets measured using in vivo imaging techniques . selected from the markers in Table 1A , Table 1B , Table 2A , 55 In certain aspects , determining the level of expression of Table 2B , and / or Table 3 can be generated using the methods a predictive marker comprises detection of mRNA. Such provided herein and can comprise between two , and all of detection can be carried out by any relevant method , includ the markers set forth in Table 1A , Table 1B , Table 2A , Table ing e . g . , PCR , northern , nucleotide array detection , in vivo 2B , or Table 3 and each and every combination in between imaging using probes capable of detection of the appropriate ( e . g . , four selected markers , 16 selected markers , 74 selected 60 nucleic acid . In other aspects , determining the level of markers , etc . ) . In some embodiments , the predictive marker expression of the predictive marker comprises detection of set comprises at least 5 , 10 , 20 , 40 , 60 , 100 , 150 , 200 , or 300 protein . Such detection can be carried out using any relevant or more markers. In other embodiments , the predictive method for protein detection , including e . g . , ELISA , western marker set comprises no more than 5 , 10 , 20 , 40 , 60 , 100 , blot , immunoassay , protein array detection , in vivo imaging 150 , 200 , 300 , 400 , 500 , 600 or 700 markers . In some 65 using probes capable of detection of the appropriate peptide . embodiments , the predictive marker set includes a plurality Determining the level of expression of a predictive of genes associated with cancer , e .g . a hematological cancer marker is compared to a reference expression level . For US 9 , 963, 747 B2 13 14 example , a reference expression level can be a predeter- therapy. In addition , the markers of the present invention can mined standard reference level of expression in order to be used to identify a patient that has become or is at risk of evaluate if expression of a marker or marker set is informa - becoming refractory to treatment with proteasome inhibition tive and make an assessment for determining whether the therapy and /or glucocorticoid therapy . The invention also patient is responsive or non - responsive . Additionally , deter - 5 features combinations of markers , referred to herein as mining the level of expression of a predictive marker can be " marker sets , ” that can predict whether a patient is likely to compared to an internal reference marker level of expression respond or not to respond to a proteasome inhibition therapy which is measured at the same time as the predictive marker and / or glucocorticoid therapy regimen . in order to make an assessment for determining whether the Table 1 sets forth predictive markers identified using patient is responsive or non - responsive . For example , 10 statistical analysis applied to samples from 224 patients , expression of a distinct marker or markers which is /are not which are specific identifiers of response or non - response to predictive markers of the invention , but which is known to proteasome inhibition therapy ( e . g ., bortezomib ) . The mark demonstrate a constant expression level can be assessed as ers in Table 1 are differentially expressed in samples from an internal reference marker level , and the level of the patients that are either responsive or non - responsive to predictive marker expression is determined as compared to 15 treatment with the proteasome inhibitor bortezomib . Thus , the reference . In an alternative example , expression of the one would appreciate that the markers identified can func selected predictive marker or markers in a tissue sample tion in a predictive model to prospectively identify patients ' which is a non - tumor sample can be assessed as an internal response to proteasome inhibition therapy , including reference marker level. The level of expression of a marker response to bortezomib or other proteasome inhibition thera or markers may be determined as having increased expres - 20 pies known in the art as well as those described in further sion in certain aspects . The level of expression of a marker detail herein . In particular, the markers in Table 1 are or markers may be determined as having decreased expres - correlated with a positive response to therapy (referred to sion in other aspects . The level of expression may be herein as " responsive , ( R ) ” ) ; or a long time until disease determined as no informative change in expression as com - progression (TTP ) as determined by a Cox proportional pared to a reference level. In still other aspects , the level of 25 hazard analysis , as described in further detail herein . A expression is determined against a pre - determined standard patient with a positive response ( either complete , partial or expression level as determined by the methods provided minimal ; see Table C ) to therapy is hereinafter referred to as herein . a " responder ” . Predictors of long time to progression are The invention also relates to various reagents and kits for useful as additional indicators of patients who are likely to diagnosing , staging, prognosing , monitoring and treating a 30 progress in disease at a slower rate and may be more likely cancer patient ( e . g . , a patient with a liquid tumor or a solid to be responsive to therapy than other patients . Additionally , tumor) , with proteasome inhibition therapy and / or glucocor - the predictive markers in Table 1 are correlated with a ticoid therapy . Provided are reagents for detection of mark - negative or poor response to an agent ( referred to herein as ers and marker sets and for use in the methods of the “ non - responsive, (NR ) ” ), or a short time to disease progres invention comprising at least two isolated predictive mark - 35 sion ( TTP ) . A patient with a poor response ( called a pro ers set forth in Table 1A , Table 1B , Table 2A , Table 2B , and gressive or refractory disease ; see Table C ) to treatment is Table 3 . Such reagents include nucleic acid probes, primers , hereinafter referred to as a " non -responder ” . These identi antibodies, antibody derivatives, antibody fragments , and fied predictive markers are useful as additional indicators of peptide probes for detection of the relevant predictive mark - patients who are likely to progress in disease at a faster rate , ers set forth in Table 1A , Table 1B , Table 2A , Table 2B , and 40 and less likely to be responsive to therapy than other Table 3 . patients. A patient with no response to treatment is herein Further provided are kits for use in the provided methods. after referred to as “ stable ” . The kits of the invention include reagents for assessing Table 1A provides predictive markers which are upregu predictive markers ( e . g ., at least one predictive marker ) and lated indicators of non - response and / or correlate with predictive marker sets ( e . g . , at least two , three , four or more 45 shorter time to progression . Marker nos. 1 - 547 in Table 1A markers selected from Table 1A , Table 1B , Table 2A , Table are newly identified predictive markers , and predictive 2B , and Table 3 ) , as well as instructions for use in accor - markers no . 548 -657 have been previously identified as dance with the methods provided herein . In certain aspects , associated markers predictive of non - response and/ or cor the kits provided contain nucleic acid probes for assessment relation with shorter time to progression . See , International of predictive markers . In still other aspects , the kits provided 50 Patent Publication No . WO04053066 , published Jun . 24 , contain antibody , antibody derivative antibody fragment, or 2004 . Table 1B provides predictive markers which are peptide reagents for assessment of predictive markers . upregulated indicators of response and /or correlate with Identification of Responsive and Non - Responsive Markers longer time to progression . Marker nos . 658 - 876 in Table 1B The present invention provides markers that are expressed are newly associated predictive markers , and predictive in a tumor that is responsive to proteasome inhibition 55 markers no . 877 - 911 have been previously identified as therapy and /or glucocorticoid therapy and whose expression associated markers predictive of response and / or correlation correlates with responsiveness to that therapeutic agent. The with longer time to progression . See , International Patent present invention also provides markers that are expressed in Publication No . WO04053066 , published Jun . 24 , 2004 . a tumor that is non - responsive to proteasome inhibition Table 2 sets forth predictive markers identified using therapy and / or glucocorticoid therapy and whose expression 60 statistical analysis applied to samples from 224 patients , correlates with non -responsiveness to such therapy. Accord - which are specific identifiers of response or non - response to ingly , one or more of the markers can be used to identify glucocorticoid therapy ( e . g . , dexamethasone ) . The markers cancers that can be successfully treated by proteasome in Table 2 are differentially expressed in samples from inhibition therapy and / or glucocorticoid therapy . One or patients that are either responsive or non - responsive to more of the markers of the present invention can be used to 65 treatment with the glucocorticoidal steroid agent dexam identify patients that can be successfully treated using ethasone . Thus , one would appreciate that the markers proteasome inhibition therapy and /or glucocorticoid identified can function in a predictive model to prospectively US 9 , 963 , 747 B2 15 16 identify patients ' response to glucocorticoid therapy , includ level in one sample , e . g . , a tumor sample , to another sample , ing response to dexamethasone or other glucocorticoid e . g . , a non - tumor sample , or between samples from different therapies known in the art as well as those described in sources . further detail herein . As in Table 1 , Table 2 sets forth Further, the expression level can be provided as a relative predictive markers identified which are specific identifiers of 5 expression level. To determine a relative expression level of response or long time to progression ; or non - response or a marker or marker set, the level of expression of the short time to progression upon therapy with glucocorticoid predictive marker or marker set is determined for 10 or more individual samples , preferably 50 or more individual treatment ( e. g ., dexamethasone ). samples in order to establish a baseline , prior to the deter Table 2A provides predictive markers which are upregu 10 mination of the expression level for the sample in question . lated indicators of non - response and /or correlate with To establish a baseline measurement, mean expression level shorter time to progression . Table 2B provides predictive of each of the predictive markers or marker sets assayed in markers which are upregulated indicators of response and /or the larger number of samples is determined and this is used correlate with longer time to progression . as a baseline expression level for the predictive markers or Table 3 sets forth predictive markers identified which do 15 marker sets in question . The expression level of the marker not distinguish between response to proteasome inhibition or marker set determined for the test sample ( absolute level therapy and response to glucocorticoid therapy , rather are of expression ) is then divided by the mean expression value indicator predictive markers of response / longer time to obtained for that marker or marker set. This provides a progression or non -response / shorter time to progression relative expression level and aids in identifying extreme with regard to either therapy , and are indicators of general 20 cases of responsive or non - responsive -ness . disease aggressiveness . Marker nos . 1203 -1423 in Table 3 Determining Responsiveness or Non - Responsiveness to an are newly associated predictive markers , and predictive Agent markers no . 1424 - 1474 have been previously identified as The expression level (including protein level) of the associated markers predictive of non - response / correlation identified predictive markers of responsive / non - responsive with shorter time to progression and /or response /correlation 25 patients and may be used to : 1) determine if a patient can be with longer time to progression related to advanced stage treated by an agent or combination of agents ; 2 ) determine patient' s response to bortezomib treatment. See , Interna if a patient is responding to treatment with an agent or tional Patent Publication No . W004053066 , published Jun . combination of agents ; 3 ) select an appropriate agent or 24 , 2004 . combination of agents for treating a patient; 4 ) monitor the level of 30 effectiveness of an ongoing treatment; 5 ) identify new In the methods of the present invention , the level of 30 eucancer therapy treatments ( either single agent proteasome expression of one or more predictive markers selected from inhibitor and /or glucocorticoid agents or complementary the group consisting of the markers identified in Table 1A , agents which can be used alternatively or in combination Table 1B , Table 2A , Table 2B , and Table 3 , is determined . with proteasome inhibition and /or glucocorticoid agents ) ; 6 ) As used herein , the level or amount of expression refers LOto 35 identify aggressiveness of a cancer ; and 7 ) select an appro the absolute level of expression of an mRNA encoded by the priate agent or combination of agents in treating early and marker or the absolute level of expression of the protein late recurrence of a cancer. In particular, the identified encoded by themarker (i . e ., whether or not expression is or predictive markers may be utilized to determine appropriate is not occurring in the cancer cells ) . therapy , to monitor clinical therapy and human trials of a Generally , it is preferable to determine the expression of 40 drug being tested for efficacy, and to develop new agents and two or more of the identified responsive or non - predictive therapeutic combinations . markers , or three or more of the identified responsive or cancer may be predisposed to respond to an agent if one non - predictive markers , or still further a larger a set of the or more of the corresponding predictive markers identified identified responsive and / or non -predictive markers, in Table 1B , Table 2B , and Table 3 (as indicated by ( + ) in selected from the predictive markers identified in Table 1A , 45 Table 3 ) demonstrate increased expression . In certain Table 1B , Table 2A , Table 2B , and Table 3 . For example , aspects of the invention , the predisposition of a cancer to be Table 4 sets forth marker sets identified using the methods responsive to an agent is determined by the methods of the described herein and can be used in the methods of the present invention , wherein informative expression of the present invention . Still further, additional and / or alternative individual predictive markers of the marker sets identified in marker sets comprising the predictive markers identified 50 Table 4 is evaluated . Likewise , the predisposition of a herein can be generated using the methods and predictive patient to be responsive to an agent is determined by the markers provided . Thus , it is possible to assess the expres - methods of the present invention , wherein a marker set sion of a panel of responsive and non - predictive markers generated using to the methods described herein wherein the using the methods and compositions provided herein . markers comprising the marker set include predictive mark As an alternative to making determinations based on the 55 ers set forth in Table 1B , Table 2B , and Table 3 , and the absolute expression level of selected markers, determina expression of the marker set is evaluated . tionsmay be based on normalized expression levels . Expres - A cancer may be predisposed to non -responsiveness to an sion levels are normalized by correcting the absolute expres - agent if one or more of the corresponding non -predictive sion level of a predictive marker by comparing its markers demonstrates informative expression levels . A can expression to the expression of a reference marker that is not 60 cer may be predisposed to non -responsiveness to an agent if a predictive marker, e . g . , a housekeeping gene that is con - one or more of the corresponding predictive markers iden stitutively expressed . Suitable markers for normalization tified in Table 1A , Table 2A , and Table 3 ( as indicated by ( - ) include housekeeping genes , such as the actin gene . Con in Table 3 ) demonstrate informative increased expression . In stitutively expressed genes are known in the art and can be certain aspects of the invention , the predisposition of a identified and selected according to the relevant tissue 65 cancer to be non - responsive to an agent is determined by the and / or situation of the patient and the analysis methods. methods of the present invention , wherein informative Such normalization allows one to compare the expression expression of the individual predictive markers of the US 9 , 963, 747 B2 17 18 marker sets identified in Table 4 is evaluated . Likewise, the glucocorticoid therapy ; and identifying that an agent is or is predisposition of a patient to be non - responsive to an agent not appropriate to treat the tumor based on the evaluation . is determined by the methods of the present invention , In such methods , a proteasome inhibition therapy and /or wherein a marker set is generated using the methods glucocorticoid therapy regimen is determined appropriate to described herein wherein the markers comprising the marker 5 treat the tumor when the expression profile of the predictive set include predictive markers set forth in Table 1A , Table marker or predictive marker set demonstrates increased 2A , and/ or Table 3 , and the expression of the marker set is responsiveness or decreased non - responsiveness according evaluated . In one aspect, the predictive marker set for evaluation of to the expression profile of the predictive markers in the a cancer predisposed to respond or predisposed to not 10 presence" of the agent. respond to a therapy comprises markers selected from those The invention also provides a method for determining set forth in any of Table 1A Table 1B , Table 2A Table 2B and whether treatment with an proteasome inhibitor therapy Table 3 . Still a further aspect contemplates markers set forth should be initiated in in a patient selected from a multiple in either Table 1A and Table 1B alone or in combination with myeloma patient, a lymphoma patient, a leukemia patient, a markers set for the in Table 2A and Table 2B and /or Table 15 lung cancer patient, a breast cancer patient, and an ovarian 3 , or alternatively , those markers set forth in Table 2A and cancer patient, a prostate cancer patient, a colon cancer Table 2B alone or in combination with Table 1A and Table patient, a kidney cancer patient, and a liver cancer patient ; 1B and /or Table 3 . In still another aspect the predictive comprising obtaining one or more samples , followed by marker or markers evaluated are selected from those set determining the level of expression of one or more markers forth in Table 3 . In certain aspects , the marker set is selected 20 which correspond to markers identified in any of Table 1A , from those set forth in Table 4 . According to the methods, Table 1B , Table 2A , Table 2B , and Table 3 in the sample ; and proteasome inhibition therapy and / or glucocorticoid therapy initiating proteasome inhibitor therapy when the expression would be continued where the expression profile indicates profile of the predictive markers identified in any one of continued responsiveness, or decreased non - responsiveness Table 1A , Table 1B , Table 2A , Table 2B , and Table 3 is using the evaluation methods described herein . 25 indicative of a responsive patient to such treatment. Alter The present invention provides methods for determining natively , the treatment is not initiated when the expression whether a cancer therapy e . g . , a proteasome inhibitor and /or profile of the predictive markers identified in any one of glucocorticoid agent, can be used to reduce the growth rate Table 1A , Table 1B , Table 2A , Table 2B , and Table 3 is of a tumor comprising evaluating expression of at least one indicative of a non - responsive patient to treatment. predictive marker or a predictive marker set in a tumor 30 The invention also provides a method for determining sample ; and identifying that proteasome inhibition therapy whether treatment with proteasome inhibition therapy and/ or glucocorticoid therapy is or is not appropriate to should be initiated in in a patient selected from a multiple reduce the growth rate of the tumor based on the evaluation . myeloma patient, a lymphoma patient, a leukemia patient, a The invention provides a method for determining whether lung cancer patient, a breast cancer patient , and an ovarian a proteasome inhibition therapeutic regimen ( e . g ., a protea - 35 cancer patient, a prostate cancer patient, a colon cancer some inhibitor agent ( e . g . , bortezomib ) alone or in combi- patient, a kidney cancer patient, and a liver cancer patient; nation with another chemotherapeutic agent ) can be used to comprising obtaining one or more samples of tumor cells reduce the growth rate of a tumor comprising determining from a patient, followed by determining the expression the expression profile of a predictive marker or predictive profile in the sample of a predictive marker set comprising marker set; and identifying that a proteasome inhibition 40 markers identified in Table 1A , Table 1B , Table 2A , Table therapeutic agent is or is not appropriate to reduce the 2B , and Table 3 ; and initiating the proteasome inhibitor growth rate of the myeloma cells based on the expression treatment when the expression profile of the predictive profile . markers identified in Table 1A , Table 1 B , Table 2A , Table Additionally provided are methods for determining 2B , and Table 3 is indicative of a responsive patient . whether a proteasome inhibitor therapy can be used to 45 Alternatively , the treatment is not initiated when the expres reduce the growth of a tumor, comprising obtaining a sample sion profile of the predictive markers identified in Table 1A , of tumor cells , evaluating the expression of one or more Table 1B , Table 2A , Table 2B , and /or Table 3 is indicative individual markers or a marker set, both in tumor cells of a non -responsive patient . exposed to the agent and in tumor cells that have not been The invention also provides a method for determining exposed to the proteasome inhibition therapy ; and identify - 50 whether treatment with an glucocorticoid therapy should be ing that an agent is or is not appropriate to treat the tumor initiated in in a patient selected from a multiple myeloma based on the evaluation . patient, a lymphoma patient, a leukemia patient, a lung The invention provides a method for determining whether cancer patient , a breast cancer patient, and an ovarian cancer a glucocorticoid regimen (e . g. , glucocorticoidal steroid patient, a prostate cancer patient, a colon cancer patient, a agent ( e . g . , dexamethasone ) alone or in combination with 55 kidney cancer patient, and a liver cancer patient; comprising another chemotherapeutic agent) can be used to reduce the obtaining one or more samples , followed by determining the growth rate of a tumor comprising determining the expres - level of expression of one or more markers which corre sion profile of a predictive marker or predictive marker set; spond to markers identified in any of Table 1A , Table 1B , and identifying that a glucocorticoid therapeutic agent is or Table 2A , Table 2B , and Table 3 in the sample ; and initiating is not appropriate to reduce the growth rate of the tumor 60 glucocorticoid therapy when the expression profile of the based on the expression profile . predictive markers identified in any one of Table 1A , Table Additionally provided are methods for determining 1B , Table 2A , Table 2B , and Table 3 is indicative of a whether a glucocorticoid therapy can be used to reduce the responsive patient to such treatment. Alternatively , the treat growth of a tumor, comprising obtaining a sample of tumor ment is not initiated when the expression profile of the cells , evaluating the expression of one or more individual 65 predictive markers identified in any one of Table 1A , Table markers or a marker set, both in tumor cells exposed to the 1B , Table 2A , Table 2B , and Table 3 is indicative of a agent and in tumor cells that have not been exposed to the non -responsive patient to treatment. US 9 ,963 , 747 B2 20 The invention also provides a method for determining cancer treatment including proteasome inhibition therapy whether treatment with glucocorticoid therapy should be and / or glucocorticoid therapy . In general, it is conceivable to initiated in in a patient selected from a multiple myeloma obtain a first sample from the patient prior to beginning patient, a lymphoma patient, a leukemia patient , a lung therapy and one or more samples during treatment. In such cancer patient, a breast cancer patient , and an ovarian cancer 5 a use , a baseline of expression prior to therapy is determined , patient, a prostate cancer patient, a colon cancer patient, a then changes in the baseline state of expression is monitored kidney cancer patient, and a liver cancer patient; comprising during the course of therapy. Alternatively , two or more obtaining one or more samples of tumor cells from a patient , successive samples obtained during treatment can be used followed by determining the expression profile in the sample without the need of a pre - treatment baseline sample . In such of a predictive marker set comprising markers identified in 10 a use , the first sample obtained from the subject is used as Table 1A , Table 1B , Table 2A , Table 2B , and Table 3 ; and a baseline for determining whether the expression of a initiating the glucocorticoid treatment when the expression particular marker or marker set is increasing or decreasing . profile of the predictive markers identified in Table 1A , In general, when monitoring the effectiveness of a thera Table 1B , Table 2A , Table 2B , and Table 3 is indicative of peutic treatment , two or more samples from a patient are a responsive patient. Alternatively , the treatment is not 15 examined . In another aspect, three or more successively initiated when the expression profile of the predictive mark - obtained samples are used , including at least one pretreat ers identified in Table 1A , Table 1B , Table 2A , Table 2B , ment sample . and /or Table 3 is indicative of a non -responsive patient. The invention provides methods for determining whether Monitoring the Effectiveness of an Anti - Cancer Agent treatment with a proteasome inhibitor therapy regimen As discussed above , the identified responsive and non - 20 should be continued in a patient selected from a multiple predictive markers can be used as pharmacodynamic mark - myeloma patient, a lymphoma patient, a leukemia patient, a ers to assess whether the tumor has become refractory to an lung cancer patient, a breast cancer patient, and an ovarian ongoing treatment ( e . g ., a proteasome inhibition therapy cancer patient, a prostate cancer patient, a colon cancer and / or glucocorticoid therapy ) . When the cancer is not patient, a kidney cancer patient, and a liver cancer patient; responding to a treatment the expression profile of the tumor 25 comprising obtaining two or more samples of tumor cells cells will change : the level or relative expression of one or from a patient at different times during the course of a more of the predictive markers ( e . g . , those predictive mark - proteasome inhibition therapy regimen , followed by evalu ers identified in Table 1A , Table 1B , Table 2A , Table 2B , ating the expression of one or more markers which corre Table 3 ) such that the expression profile represents a non spond to markers identified in any of Table 1A , Table 1B , responsive patient. 30 Table 2A , Table 2B , and Table 3 in the two ormore samples ; In one such use , the invention provides methods for and continuing the treatment when the expression profile of determining whether a cancer therapy comprising protea - the predictive markers identified in any one of Table 1A , some inhibition therapy and/ or glucocorticoid therapy Table 1B , Table 2A , Table 2B , and Table 3 is indicative of should be continued in a cancer patient, comprising deter a responsive patient during the course of the treatment. In mining the expression of at least one predictive marker or a 35 such methods, a proteasome inhibition therapy and regimen marker set, wherein the markers are selected from those set is determined appropriate to treat the patient when the forth in any of Table 1A , Table 1B , Table 2A , Table 2B , or expression profile of the predictive marker or predictive Table 3 , in a tumor sample of a patient exposed to a marker set demonstrates increased responsiveness or proteasome inhibition therapy and /or glucocorticoid decreased non -responsiveness according to the expression therapy ; and continuing treatment when the expression 40 profile of the predictive markers in the presence of the agent. profile of the marker or marker set demonstrates respon - Additionally provided are methods for determining siveness to the agent being used . whether treatment with a proteasome inhibitor therapy regi In another such use , the invention provides methods for men should be continued in in a patient selected from a determining whether a proteasome inhibition therapy and / or multiple myeloma patient, a lymphoma patient, a leukemia glucocorticoid therapy should be discontinued in a cancer 45 patient, a lung cancer patient, a breast cancer patient, and an patient, comprising determining the expression of at least ovarian cancer patient, a prostate cancer patient, a colon one predictive marker or a predictive marker set , wherein the cancer patient, a kidney cancer patient, and a liver cancer markers are selected from those set forth in any of Table 1A , patient; comprising obtaining two or more samples of tumor Table 1B , Table 2A , Table 2B , or Table 3 in a tumor sample cells from a patient at different times during the course of of a patient exposed to a proteasome inhibition therapy 50 anti- cancer therapy treatment, followed by evaluating the and / or glucocorticoid therapy ; and discontinuing or altering expression of of a predictive marker set comprising markers treatment when the expression profile of the markers iden identified in any of Table 1A , Table 1B , Table 2A , Table 2B , tified in any one of Table 1A , Table 1B , Table 2A , Table 2B , and Table 3 in the two or more samples ; and continuing the or Table 3 demonstrates non -responsiveness to the agent treatment when the expression profile of the predictive being used . 55 marker set indicates increased responsiveness or decreased As used herein , a patient refers to any subject undergoing non -responsiveness according to the expression during the proteasome inhibition therapy and / or glucocorticoid therapy course of treatment. Alternatively , the treatment is discon for cancer treatment . The subject may be a human patient tinued when the expression profile of the marker set dem undergoing proteasome inhibition (e .g ., bortezomib or other onstrates decreased responsiveness and /or increased non related agent ) and / or glucocorticoid ( e . g . , dexamethasone ) 60 responsiveness during the course of treatment. therapy using a sole therapeutic agent. The subject may be The invention provides methods for determining whether a human patient undergoing proteasome inhibition ( e . g ., treatment with a glucocorticoid therapy regimen should be bortezomib or other related agent) and / or glucocorticoid continued in a patient selected from a multiple myeloma ( e . g . , dexamethasone ) therapy using a therapeutic agent in patient, a lymphoma patient, a leukemia patient, a lung conjunction with another agent ( e . g . , a chemotherapy treat- 65 cancer patient, a breast cancer patient, and an ovarian cancer ment) . The present invention also includes comparing two or patient , a prostate cancer patient, a colon cancer patient, a more samples obtained from a patient undergoing anti - kidney cancer patient, and a liver cancer patient ; comprising US 9 , 963, 747 B2 obtaining two or more samples of tumor cells from a patient A skilled artisan can readily select and obtain the appro at different times during the course of a glucocorticoid priate cancer cells that are used in the present method . For therapy regimen , followed by evaluating the expression of cancer cell lines , sources such as The National Cancer one or more markers which correspond to markers identified Institute , for the NCI -60 cells , are preferred . For cancer cells in any of Table 1A , Table 1B , Table 2A , Table 2B , and Table 5 obtained from a patient, standard biopsy methods, such as a 3 in the two or more samples ; and continuing the treatment needle biopsy , can be employed when the expression profile of the predictive markers iden - Myeloma samples were used to identify the markers of tified in any one of Table 1A , Table 1B , Table 2A , Table 2B , the present invention . Further, the expression level of mark and Table 3 is indicative of a responsive patient during the ers can be evaluated in other tissue types including disorders course of treatment . In such methods, a glucocorticoid 10 of related hematological cell types , including, e . g ., Walden therapy regimen is determined appropriate to treat the stroms macrogobulinemia , Myelodysplastic syndrome and patient when the expression profile of the predictive marker other hematological cancers including lymphomas, leuke or predictive marker set demonstrates increased responsive - mias , as well as tumors of various solid tissues . It will thus ness or decreased non - responsiveness according to the be appreciated that cells from other hematologic malignan expression profile of the predictive markers in the presence 15 cies including , e .g ., B - cell Lymphomas, Non - Hodgkins of the agent. Lymphoma, Waldenstrom ' s syndrome, or other leukemias Additionally provided are methods for determining will be useful in the methods of the present invention . Still whether treatment with a glucocorticoid therapy regimen further, the predictive markers predicting disease aggres should be continued in in a patient selected from a multiple siveness as well as responsiveness and non - responsiveness myeloma patient, a lymphoma patient, a leukemia patient, a 20 to proteasome inhibition therapeutic agents in solid tumors lung cancer patient, a breast cancer patient, and an ovarian (e .g ., lung , breast , prostate , ovary, colon , kidney, and liver) , cancer patient, a prostate cancer patient, a colon cancer can also be useful in the methods of the present invention . patient, a kidney cancer patient, and a liver cancer patient; Preferably , the samples used will be from similar tumors comprising obtaining two or more samples of tumor cells or from non -cancerous cells of the same tissue origin as the from a patient at different times during the course of a 25 tumor in question . The choice of the cell source is dependent glucocorticoid therapy regimen , followed by evaluating the on the use of the relative expression level data . For example , expression of of a predictive marker set comprising markers using tumors of similar types for obtaining a mean expres identified in any of Table 1A , Table 1B , Table 2A , Table 2B , sion score allows for the identification of extreme cases of and Table 3 in the two or more samples ; and continuing the responsive or non - responsive -ness . Using expression found treatment when the expression profile of the predictive 30 in normal tissues as a mean expression score aids in vali marker set indicates increased responsiveness or decreased dating whether the responsive/ non - predictive marker or non - responsiveness according to the expression during the marker set assayed is tumor specific (versus normal cells ) . course of treatment . Alternatively , the treatment is discon - Such a later use is particularly important in identifying tinued when the expression profile of the marker set dem whether a responsive or non -predictive marker or marker set onstrates decreased responsiveness and / or increased non - 35 can serve as a target marker or marker set . In addition , as responsiveness during the course of treatment. more data is accumulated , the mean expression value can be The invention certain aspects of the invention relate to revised , providing improved relative expression values methods of treatment and / or diagnosis of a patient with based on accumulated data . cancer utilizing samples . The source of the cancer cells used Detection Assays in the present methods will be based on how the method of 40 Various methods are available to examine the expression the present invention is being used . For example , if the of the markers , including gene array / chip technology , RT method is being used to determine whether a patient' s PCR , in -situ hybridization , immunohistochemistry , immu cancer can be treated with an agent, or a combination of noblotting , FISH ( fluorescence in - situ hybridization ) , FACS agents , then the preferred source of sample will be cancer analyses, northern blot, southern blot or cytogenetic analy cells obtained from a tumor from the patient, e . g . , a tumor 45 ses . A skilled artisan can select from these or other appro biopsy ( including a solid or a liquid tumor ), a blood sample , priate and available methods based on the nature of the a plasma sample , a urine sample , a saliva sample , a lymph marker (s ) , tissue sample and disease in question . Different sample or other sample can be used . A sample obtained from methods or combinations of methods could be appropriate in a tumor can be enriched for tumor cells to increase the different cases or, for instance in different solid or hemato specificity of the analysis . A variety ofmethods known in the 50 logical tumor types . art can be used to enrich for tumor cells , including differ - In certain aspects of the invention , the expression of ential centrifugation , fluorescence cell sorting analysis predictive marker or markers identified in Table 1A , Table ( FACS) , isolating cells based on growth independent of 1B , Table 2A , Table 2B , and Table 3 is detected by mea substrate attachment, binding to a selection agent, e . g . to an suring mRNA which corresponds to the predictive marker or antibody to a tumor marker and furthermore attaching the 55 marker set . In yet another aspects of the invention , the antibody and thus the bound tumor cell to a solid support, expression of markers which correspond to markers or etc . Alternatively , a cancer cell line similar to the type of marker sets identified in Table 1A , Table 1B , Table 2A , Table cancer being treated can be assayed . For example , if mul 2B , and Table 3 , is detected by measuring protein which tiple myeloma is being treated , then a myeloma cell line can corresponds to the marker or marker set . be used . If the method is being used to predict or monitor the 60 An exemplary method for detecting the presence or effectiveness of a therapeutic protocol, then a tissue or blood absence of a nucleic acid or polypeptide corresponding to a sample from a patient being treated is a preferred source . If marker of the invention in a biological sample involves the method is being used to determine the activity of an obtaining a biological sample ( e . g . a tumor sample ) from a agent, the efficacy of an agent, or identify new therapeutic test subject and contacting the biological sample with a agents or combinations , any cancer cells , e . g ., cells of a 65 compound or an agent capable of detecting the polypeptide cancer cell line , cells isolated from a tumor of an animal or nucleic acid ( e . g . , mRNA , genomic DNA , or cDNA ) . The model, can be used . detection methods of the invention can thus be used to detect US 9 , 963, 747 B2 23 24 mRNA , protein , cDNA , or genomic DNA , for example , in or indirectly , with detectable labels discussed herein and a biological sample in vitro as well as in vivo . For example , which are well - known to one skilled in the art . in vitro techniques for detection ofmRNA include Northern It is also possible to directly detect marker/ probe complex hybridizations. in situ hybridizations, and TaqMan assays formation without further manipulation or labeling of either (Applied Biosystems) under GLP approved laboratory con - 5 component (marker or probe ), for example by utilizing the ditions . In vitro techniques for detection of a polypeptide technique of fluorescence energy transfer (see , for example , corresponding to a marker of the invention include enzyme Lakowicz et al. , U . S . Pat. No . 5 ,631 , 169 ; Stavrianopoulos, et al ., U .S . Pat. No . 4 ,868 , 103 ). A fluorophore label on the linked immunosorbent assays (ELISAs ) , Western blots , first, ' donor ' molecule is selected such that, upon excitation immunoprecipitations and immunofluorescence . In vitro 10 with incident light of appropriate wavelength , its emitted techniques for detection of genomic DNA include Southern fluorescent energy will be absorbed by a fluorescent label on hybridizations . Furthermore, in vivo techniques for detec a second “ acceptormolecule , which in turn is able to tion of a polypeptide corresponding to a marker of the fluoresce due to the absorbed energy . Alternately , the invention include introducing into a subject a labeled anti ‘donor ' protein molecule may simply utilize the natural body directed against the polypeptide . For examplemple ;, methe 15 fluofluorescent energy of tryptophan residues . Labels are chosen antibody can be labeled with a radioactive marker whose that emit different wavelengths of light, such that the ' accep presence and location in a subject can be detected by tor ' molecule label may be differentiated from that of the standard imaging techniques. ' donor ' . Since the efficiency of energy transfer between the A general principle of such diagnostic and prognostic labels is related to the distance separating the molecules , assays involves preparing a sample or reaction mixture that 20 spatial relationships between the molecules can be assessed . may contain a marker , and a probe , under appropriate In a situation in which binding occurs between the mol conditions and for a time sufficient to allow the marker and ecules, the fluorescent emission of the “ acceptor ' molecule probe to interact and bind , thus forming a complex that can label in the assay should be maximal. An FET binding event be removed and /or detected in the reaction mixture . These can be conveniently measured through standard fluorometric assays can be conducted in a variety of ways . 25 detection means well known in the art ( e . g . , using a fluo For example , one method to conduct such an assay would rimeter ) . involve anchoring the marker or probe onto a solid phase In another example , determination of the ability of a probe support , also referred to as a substrate , and detecting target to recognize a marker can be accomplished without labeling marker /probe complexes anchored on the solid phase at the either assay component ( probe or marker ) by utilizing a end of the reaction . In one example of such a method , a 30 technology such as real - time Biomolecular Interaction sample from a subject, which is to be assayed for presence Analysis ( BIA ) ( see , e . g ., Sjolander , S . and Urbaniczky , C . , and / or concentration of marker , can be anchored onto a 1991, Anal. Chem . 63 : 2338 - 2345 and Szabo et al. , 1995 , carrier or solid phase support . In another example , the Curr . Opin . Struct. Biol. 5 :699 - 705 ) . As used herein , “ BIA " reverse situation is possible , in which the probe can be or “ surface plasmon resonance ” is a technology for studying anchored to a solid phase and a sample from a subject can 35 biospecific interactions in real time, without labeling any of be allowed to react as an unanchored component of the the interactants (e . g ., BIAcore ). Changes in the mass at the assay . One example of such an example includes use of an binding surface ( indicative of a binding event) result in array or chip which contains a predictive marker or marker alterations of the refractive index of light near the surface set anchored for expression analysis of the sample . ( the optical phenomenon of surface plasmon resonance There are many established methods for anchoring assay 40 (SPR ) ) , resulting in a detectable signal which can be used as components to a solid phase . These include , without limi- an indication of real- time reactions between biologicalmol tation , marker or probe molecules which are immobilized ecules . through conjugation of biotin and streptavidin . Such bioti - Alternatively , in another example , analogous diagnostic nylated assay components can be prepared from biotin -NHS and prognostic assays can be conducted with marker and ( N -hydroxy - succinimide ) using techniques known in the art 45 probe as solutes in a liquid phase . In such an assay , the ( e . g . , biotinylation kit , Pierce Chemicals, Rockford , 111 . ) , and complexed marker and probe are separated from uncom immobilized in the wells of streptavidin - coated 96 well plexed components by any of a number of standard tech plates (Pierce Chemical) . In certain aspects , the surfaces niques, including but not limited to : differential centrifuga with immobilized assay components can be prepared in tion , chromatography , electrophoresis and advance and stored . Other suitable carriers or solid phase 50 immunoprecipitation . In differential centrifugation ,marker / supports for such assays include any material capable of probe complexes may be separated from uncomplexed assay binding the class of molecule to which the marker or probe components through a series of centrifugal steps, due to the belongs. Well- known supports or carriers include, but are different sedimentation equilibria of complexes based on not limited to , glass, polystyrene, nylon , polypropylene , their different sizes and densities ( see, for example , Rivas, nylon , polyethylene , dextran , amylases , natural and modi - 55 G . , and Minton , A . P ., 1993 , Trends Biochem Sci . 18 ( 8 ) : 284 fied celluloses, polyacrylamides, gabbros , and magnetite . 7 ) . Standard chromatographic techniques may also be uti In order to conduct assays with the above mentioned lized to separate complexed molecules from uncomplexed approaches , the non - immobilized component is added to the ones . For example , gel filtration chromatography separates solid phase upon which the second component is anchored molecules based on size , and through the utilization of an After the reaction is complete , uncomplexed components 60 appropriate gel filtration resin in a column format, for may be removed ( e . g . , by washing ) under conditions such example , the relatively larger complex may be separated that any complexes formed will remain immobilized upon from the relatively smaller uncomplexed components . Simi the solid phase . The detection of marker /probe complexes larly , the relatively different charge properties of the marker/ anchored to the solid phase can be accomplished in a number probe complex as compared to the uncomplexed compo ofmethods outlined herein . In one example , when the probe 65 nents may be exploited to differentiate the complex from is the unanchored assay component, can be labeled for the uncomplexed components , for example through the utiliza purpose of detection and readout of the assay , either directly tion of ion -exchange chromatography resins. Such resins US 9 , 963 , 747 B2 25 26 and chromatographic techniques are well known to one amplification method , followed by the detection of the skilled in the art ( see, e . g . , Heegaard , N . H . , 1998 , J Mol. amplified molecules using techniques well known to those Recognit. Winter 11 ( 1 - 6 ) : 141- 8 ; Hage , D . S ., and Tweed , S . of skill in the art . These detection schemes are especially A . J Chromatogr B Biomed Sci Appl 1997 Oct . 10 ; 699 ( 1 - useful for the detection of nucleic acid molecules if such 2 ): 499 - 525 ) . Gel electrophoresis may also be employed to 5 molecules are present in very low numbers . As used herein , separate complexed assay components from unbound com - amplification primers are defined as being a pair of nucleic ponents (see , e . g . , Ausubel et al ., ed ., Current Protocols in acid molecules that can anneal to 5 ' or 3 ' regions of a gene Molecular Biology , John Wiley & Sons, New York , 1987 - (plus and minus strands, respectively , or vice - versa ) and 1999 ) . In this technique , protein or nucleic acid complexes contain a short region in between . In general, amplification are separated based on size or charge , for example . In order 10 primers are from about 10 to 30 nucleotides in length and to maintain the binding interaction during the electropho - flank a region from about 50 to 200 nucleotides in length . retic process, non - denaturing gel matrix materials and con - Under appropriate conditions and with appropriate reagents , ditions in the absence of reducing agent are typically pre - such primers permit the amplification of a nucleic acid ferred . Appropriate conditions to the particular assay and molecule comprising the nucleotide sequence flanked by the components thereof will be well known to one skilled in the 15 primers . art. For in situ methods, mRNA does not need to be isolated The level ofmRNA corresponding to the marker can be from the cancer cells prior to detection . In such methods , a determined both by in situ and by in vitro formats in a cell or tissue sample is prepared /processed using known biological sample using methods known in the art . The term histological methods . The sample is then immobilized on a " biological sample ” is intended to include tissues , cells , 20 support , typically a glass slide , and then contacted with a biological fluids and isolates thereof, isolated from a subject, probe that can hybridize to mRNA that encodes the marker. as well as tissues, cells and fluids present within a subject. As an alternative to making determinations based on the Many expression detection methods use isolated RNA . For absolute expression level of the marker, determinations may in vitro methods, any RNA isolation technique that does not be based on the normalized expression level of the marker. select against the isolation of mRNA can be utilized for the 25 Expression levels are normalized by correcting the absolute purification of RNA from tumor cells ( see , e . g ., Ausubel et expression level of a marker by comparing its expression to al. , ed ., Current Protocols in Molecular Biology, John Wiley the expression of a reference gene that is not a marker, e . g . , & Sons, New York 1987 - 1999 ) . Additionally , large numbers a housekeeping gene that is constitutively expressed . Suit of tissue samples can readily be processed using techniques able genes for normalization include housekeeping genes well known to those of skill in the art , such as , for example , 30 such as the actin gene , or epithelial cell -specific genes . This the single -step RNA isolation process of Chomczynski normalization allows the comparison of the expression level ( 1989, U . S . Pat. No . 4 ,843 , 155 ) . in one sample , e . g ., a patient sample , to another sample , e . g . , One diagnostic method for the detection of mRNA levels a non - cancer sample , or between samples from different involves contacting the isolated mRNA with a nucleic acid sources . molecule (probe ) that can hybridize to the mRNA encoded 35 Alternatively , the expression level can be provided as a by the gene being detected . The nucleic acid probe can be , relative expression level. To determine a relative expression for example , a full- length cDNA , or a portion thereof, such level of a marker, the level of expression of the marker is as an oligonucleotide of at least 7 , 15 , 30 , 50 , 100 , 250 or determined for 10 or more samples of normal versus cancer 500 nucleotides in length and sufficient to specifically cell isolates, preferably 50 or more samples , prior to the hybridize under stringent conditions to a mRNA or genomic 40 determination of the expression level for the sample in DNA encoding a marker of the present invention . Other question . The mean expression level of each of the markers suitable probes for use in the diagnostic assays of the and marker sets assayed in the larger number of samples is invention are described herein . Hybridization of an mRNA determined and this is used as a baseline expression level for with the probe indicates that the marker in question is being the marker. The expression level of the marker determined expressed . 45 for the test sample ( absolute level of expression ) is then In one format, the mRNA is immobilized on a solid divided by the mean expression value obtained for that surface and contacted with a probe , for example by running marker . This provides a relative expression level . the isolated mRNA on an agarose gel and transferring the In another aspect of the present invention , a polypeptide mRNA from the gel to a membrane , such as nitrocellulose . corresponding to a marker is detected . A preferred agent for In an alternative format, the probe( s ) are immobilized on a 50 detecting a polypeptide of the invention is an antibody solid surface and the mRNA is contacted with the probe (s ), capable of binding to a polypeptide corresponding to a for example , in an Affymetrix gene chip array. A skilled marker of the invention , preferably an antibody with a artisan can readily adapt known mRNA detection methods detectable label. Antibodies can be polyclonal, or more for use in detecting the level of mRNA encoded by the preferably , monoclonal. An intact antibody, or a fragment markers of the present invention . 55 thereof (e . g ., Fab or F (ab ') 2 ) can be used . The term An alternative method for determining the level of mRNA " labeled ” , with regard to the probe or antibody, is intended corresponding to a marker of the present invention in a to encompass direct labeling of the probe or antibody by sample involves the process of nucleic acid amplification , coupling ( i. e ., physically linking ) a detectable substance to e . g . , by rtPCR ( the experimental description set forth in the probe or antibody , as well as indirect labeling of the Mullis , 1987 , U . S . Pat . No. 4 ,683 , 202 ) , ligase chain reaction 60 probe or antibody by reactivity with another reagent that is (Barany , 1991 , Proc. Natl. Acad . Sci . USA , 88 : 189 - 193 ) , self directly labeled . Examples of indirect labeling include sustained sequence replication (Guatelli et al ., 1990 , Proc . detection of a primary antibody using a fluorescently labeled Natl . Acad . Sci . USA 87: 1874 - 1878 ), transcriptional ampli secondary antibody and end - labeling of a DNA probe with fication system (Kwoh et al. , 1989 , Proc. Natl. Acad . Sci. biotin such that it can be detected with fluorescently labeled USA 86 : 1173 - 1177 ) , Q - Beta Replicase (Lizardi et al , 1988 , 65 streptavidin . Bio / Technology 6 : 1197 ) , rolling circle replication (Lizardi et A variety of formats can be employed to determine al. , U .S . Pat. No. 5, 854 , 033 ) or any other nucleic acid whether a sample contains a protein that binds to a given US 9 , 963 , 747 B2 27 28 antibody. Examples of such formats include, but are not Electronic apparatus , including readable arrays compris limited to , enzyme immunoassay (ETA ) , radioimmunoassay ing at least one predictive marker of the present invention is (RIA ), Western blot analysis and enzyme linked immuno also contemplated for use in conjunction with the methods absorbant assay (ELISA ) . A skilled artisan can readily adapt of the invention . As used herein , " electronic apparatus known protein / antibody detection methods for use in deter- 5 readable media ” refers to any suitable medium for storing , mining whether a sample comprising cancer cells express a holding or containing data or information that can be read marker of the present invention . and accessed directly by an electronic apparatus . As used herein , the term “ electronic apparatus ” is intended to include In one format, antibodies , or antibody fragments , can be any suitable computing or processing apparatus or other used in methods such as Western blots or immunofluores 10 device configured or adapted for storing data or information . cence techniques to detect the expressed proteins. In such Examples of electronic apparatus suitable for use with the uses , it is generally preferable to immobilize either the present invention and monitoring of the recorded informa antibody or proteins on a solid support. Suitable solid phase tion include stand - alone computing apparatus ; networks , supports or carriers include any support capable of binding including a local area network (LAN ) , a wide area network an antigen or an antibody. Well - known supports or carriers 15 (WAN ) Internet. Intranet . and Extranet: electronic appli include glass, polystyrene, polypropylene, polyethylene , ances such as a personal digital assistants (PDAs ) , cellular dextran , nylon , amylases , natural and modified celluloses, phone, pager and the like ; and local and distributed process polyacrylamides, gabbros , and magnetite . ing systems. As used herein , " recorded ” refers to a process One skilled in the art will know many other suitable for storing or encoding information on the electronic appa carriers for binding antibody or antigen , and will be able to 20 ratus readable medium . Those skilled in the art can readily adapt such support for use with the present invention . For adopt any of the presently known methods for recording example , protein isolated from tumor cells can be run on a information on known media to generate manufactures polyacrylamide gel electrophoresis and immobilized onto a comprising the markers of the present invention . solid phase support such as nitrocellulose . The support can For example , microarray systems are well known and then be washed with suitable buffers followed by treatment 25 used in the art for assessment of samples, whether by with the detectably labeled antibody . The solid phase sup - assessment gene expression ( e . g . , RNA detection , protein port can then be washed with the buffer a second time to detection ) , or metabolite production , for example . Microar remove unbound antibody. The amount of bound label on rays for use according to the invention include one or more the solid support can then be detected by conventional probes of predictive marker ( s ) of the invention characteristic means . 30 of response and /or non -response to a therapeutic regimen as Another method for determining the level of a polypep - described herein . In one embodiment, the microarray com tide corresponding to a marker is mass spectrometry . For prises one or more probes corresponding to one or more of example , intact proteins or peptides , e . g . , tryptic peptides markers selected from the group consisting of markers can be analyzed from a sample , e . g . , a tumor sample , blood , which demonstrate increased expression in responsive plasma, urine , etc , containing one or more polypeptide 35 patients , and genes which demonstrate non -response in markers . The method can further include treating the sample patients . A number of different microarray configurations to lower the amounts of abundant proteins, e . g . serum and methods for their production are known to those of skill albumin , to increase the sensitivity of the method . For in the art and are disclosed , for example , in U . S . Pat. Nos . example , liquid chromatography can be used to fractionate 5 ,242 , 974 ; 5 , 384 ,261 ; 5 , 405 ,783 ; 5 ,412 , 087 ; 5 ,424 ,186 ; the sample so portions of the sample can be analyzed 40 5 ,429 , 807 ; 5 , 436 , 327 ; 5 ,445 , 934 ; 5 ,556 ,752 ; 5 ,405 ,783 ; separately by mass spectrometry . The steps can be per 5 , 412 ,087 ; 5 ,424 ,186 ; 5 ,429 ,807 ; 5 ,436 , 327 ; 5 ,472 ,672 ; formed in separate systems or in a combined liquid chro 5 ,527 ,681 ; 5 ,529 , 756 ; 5 ,545 ,531 ; 5 ,554 ,501 ; 5 , 561, 071 ; matography /mass spectrometry system (LC /MS , see for 5 ,571 ,639 ; 5 , 593 , 839 ; 5 ,624 ,711 ; 5 ,700 ,637 ; 5 ,744 , 305 ; example , Liao , et al. Arthritis Rheum . 50 : 3792 -3803 5 ,770 ,456 ; 5 , 770 ,722 ; 5 , 837 , 832 , 5 ,856 , 101 ; 5 , 874 , 219 ; ( 2004 )) . The mass spectrometry system also can be in 45 5 , 885 , 837 ; 5 ,919 , 523 ; 5 , 981 , 185 ; 6 ,022 , 963 ; 6 ,077 ,674 ; tandem (MS /MS ) mode . The charge state distribution of the 6 , 156 , 501 ; 6 , 261 ,776 ; 6 , 346 ,413 ; 6 ,440 ,677 ; 6 ,451 ,536 ; protein or peptide mixture can be acquired over one or 6 ,576 ,424 ; 6 ,610 ,482 ; 5 , 143 , 854 ; 5 ,288 ,644 ; 5 , 324 ,633 ; multiple scans and analyzed by statistical methods , e . g . 5 ,432 , 049 ; 5 , 470 ,710 ; 5 ,492 ,806 ; 5 ,503 , 980 ; 5 ,510 , 270 : using the retention time and mass - to - charge ratio ( m / z ) in 5 ,525 ,464 ; 5 ,547 ,839 ; 5 , 580 ,732 ; 5 , 661, 028 ; 5 , 848 ,659 ; and the LC /MS system , to identify proteins expressed at statis - 50 5 , 874 , 219 ; Shena , et al ., Tibtech 16 : 301, 1998 ; Duggan , et tically significant levels differentially in samples from al. , Nat. Genet . 21: 10, 1999 ; Bowtell, et al. , Nat. Genet . patients responsive or non - responsive to proteasome inhi- 21: 25 , 1999 ; Lipshutz , et al. , 21 Nature Genet . 20 - 24 , 1999 ; bition and /or glucocorticoid therapy . Examples of mass Blanchard , et al. , 11 Biosensors and Bioelectronics , 687 - 90 , spectrometers which can be used are an ion trap system 1996 ; Maskos, et al. , 21 Nucleic Acids Res. 4663 -69 , 1993 ; ( ThermoFinnigan , San Jose , Calif . ) or a quadrupole time- 55 Hughes , et al. , Nat . Biotechol. 19 : 342 , 2001 ; each of which of- flight mass spectrometer (Applied Biosystems, Foster are herein incorporated by reference . A tissue microarray can City, Calif .) . The method can further include the step of be used for protein identification (see Hans et al Blood peptide mass fingerprinting, e . g . in a matrix - assisted laser 103 :275 - 282 ( 2004 ) ) . A phage - epitope microarray can be desorption ionization with time- of - flight (MALDI - TOF ) used to identify one or more proteins in a sample based on mass spectrometry method . The method can further include 60 whether the protein or proteins induce auto -antibodies in the the step of sequencing one or more of the tryptic peptides. patient (Bradford et al . Urol. Oncol. 24 : 237 - 242 (2006 ) ) . Results of this method can be used to identify proteins from A microarray thus comprises one or more probes corre primary sequence databases, e . g. maintained by the National sponding to one or more predictive markers identified in Center for Biotechnology Information , Bethesda , Md. , or the Table 1A , Table 1B , Table 2A , Table 2B , and Table 3 . The Swiss Institute for Bioinformatics, Geneva , Switzerland , and 65 microarray may comprise probes corresponding to , for based on mass spectrometry tryptic peptide m / z base peaks . example , at least 10 , at least 20 , at least 50 , at least 100 , or Electronic Apparatus Readable Arrays at least 1000 predictive markers of the invention character US 9 , 963 , 747 B2 29 30 istic of patient response to proteasome inhibition therapy different antibody which binds to either the polypeptide or and / or glucocorticoid therapy . The microarray may com the first antibody and is conjugated to a detectable label. prise probes corresponding to one or more predictive mark - For oligonucleotide - based kits , the kit can comprise , for ers as set forth herein . Still further, the microarray may example : ( 1 ) an oligonucleotide , e . g . , a detectably labeled comprise complete marker sets as set forth herein and which 5 oligonucleotide, which hybridizes to a nucleic acid sequence may be selected and compiled according to the methods set encoding a polypeptide corresponding to a marker of the forth herein . The microarray can be used to assay expression invention ; ( 2 ) a pair of primers useful for amplifying a of one or more predictive markers or predictive marker sets nucleic acid molecule corresponding to a marker of the in the array . In one example , the array can be used to assay invention ; or ( 3 ) a marker set comprising oligonucleotides more than one predictive marker or marker set expression in 10 which hybridize to at least two nucleic acid sequences a sample to ascertain an expression profile of markers in the encoding polypeptide predictive markers of the invention . array . In this manner , up to about 44 , 000 markers can be The kit can also comprise , e . g . , a buffering agent , a preser simultaneously assayed for expression . This allows a profile vative , or a protein stabilizing agent. The kit can further to be developed showing a battery of markers specifically comprise components necessary for detecting the detectable expressed in one or more samples. Still further, this allows 15 label ( e . g . , an enzyme or a substrate ) . For marker sets , the kit a profile to be developed to assess responsiveness to one or can comprise a marker set array or chip for use in detecting more therapies ( e . g . , glucocorticoid therapy or proteasome the predictive markers . The kit can also contain a reference inhibition therapy ) . sample or a series of reference samples which can be The array is also useful for ascertaining differential assayed and compared to the test sample. Each component expression patterns of one or more markers in normal and 20 of the kit can be enclosed within an individual container and abnormal ( e . g . , sample , e . g ., tumor ) cells . This provides a all of the various containers can be within a single package , battery of predictive markers that could serve as a tool for along with instructions for interpreting the results of the ease of identification of responsive and non - responsive assays performed using the kit . patients . Further , the array is useful for ascertaining expres - Therapeutic Agents sion of reference markers for reference expression levels . In 25 The markers and marker sets of the present invention are another example , the array can be used to monitor the time predictive of patients who are responsive or non - responsive course of expression of one or more predictive markers in (sensitive or resistant) proteasome inhibition therapy and/ or the array . glucocorticoid therapy regimens , generally . In addition to such qualitative determination , the inven - Therapeutic agents for use in the methods of the invention tion allows the quantitation of marker expression . Thus , 30 include a class of therapeutic agents known as proteosome predictive markers can be grouped on the basis of marker inhibitors . “ Proteasome inhibitor ” shallmean any substance sets or responsive and non - responsive indications by the which directly or indirectly inhibits the 20S or 26S protea level of expression in the sample . This is useful, for some or the activity thereof. Preferably, such inhibition is example , in ascertaining the responsive or non -responsive specific , i. e ., the proteasome inhibitor inhibits proteasome indication of the sample by virtue of scoring the expression 35 activity at a concentration that is lower than the concentra levels according to the methods provided herein . tion of the inhibitor concentration required to produce The array is also useful for ascertaining the effect of the another, unrelated biological effect. Preferably , the concen expression of a marker on the expression of other predictive tration of the proteasome inhibitor required for proteasome markers in the same cell or in different cells . This provides , inhibition is at least 2 - fold lower, more preferably at least for example , a selection of alternate molecular targets for 40 5 - fold lower , even more preferably at least 10 - fold lower, therapeutic intervention if the proteasome inhibition regi - and most preferably at least 20 - fold lower than the concen men and / or glucocorticoid therapy regimen is non - respon - tration required to produce an unrelated biological effect. sive . Proteasome inhibitor compounds of this invention are those Reagents and Kits compounds which are useful for inhibiting tumor growth , The invention also encompasses kits for detecting the 45 ( e . g ., multiple myeloma tumor growth , other hematological presence of a polypeptide or nucleic acid corresponding to or solid tumors as described in further detail herein ) in a marker of the invention in a sample ( e . g . a tumor sample ). patients . Proteasome inhibitor also is intended to include Such kits can be used to determine if a subject is predisposed pharmaceutically acceptable salts of the compounds . to response or non - response to an anti -cancer therapy regi- Proteasome inhibition therapy , generally comprises at men . In another aspect, the invention provides a test kit for 50 least an agent which inhibits proteasome activity in a cell, monitoring the efficacy of a compound or therapeutic in a and can comprise additional therapeutic agents . In certain sample . For example , the kit may comprise a labeled probe applications of the invention , the agent used in methods of capable of detecting a polypeptide or an mRNA encoding a the invention is a proteasome inhibitor . One example of a polypeptide corresponding to a marker of the invention in a proteosome inhibitor has been approved for treatment of biological sample and means for determining the amount of 55 multiple myeloma patients who have received at least two the polypeptide or mRNA in the sample ( e . g ., an antibody prior therapies and have demonstrated disease progression which binds the polypeptide or an oligonucleotide probe on the last therapy and is presently being tested in clinical which binds to DNA or mRNA encoding the polypeptide ). trials for additional indications is bortezomib . Proteasome Kits may further include instructions for use of the provided inhibition therapy regimens can also include additional kits and interpreting the results obtained using the kit ; 60 therapeutic agents such as chemotherapeutic agents . Some additional reagents for preparation of probes for use in the examples of traditional chemotherapeutic agents are set methods provided ; and detectable label, alone or conjugated forth in Table A . Alternatively or in combination with these to the provided probe( s ). chemotherapeutic agents, newer classes of chemotherapeu For antibody- based kits , the kit can comprise , for tic agents can also be used in proteasome inhibition therapy . example : ( 1 ) a first antibody ( e . g ., attached to a solid 65 The examples described herein entail use of the protea support ) which binds to a polypeptide corresponding to a some inhibitor N - pyrazinecarbonyl- L -phenylalanine - L -leu marker of the invention ; and , optionally , (2 ) a second , cineboronic acid , bortezomib (( VELCADETM ); formerly US 9 ,963 , 747 B2 32 known as MLN341 or PS -341 ) . The language “ proteasome coids utilized in treatmentof hematological and combination inhibitor” is intended to include bortezomib , compounds therapy in solid tumors include hydrocortisone, predisolone , which are structurally similar to bortezomib and / or analogs prednisone, and triamcinolone . Glucocorticoid therapy regi of bortezomib . “ Proteasome inhibitor ” can also include mens can be used alone , or can be used in conjunction with “ mimics” . “ Mimics” is intended to include compounds 5 additional chemotherapeutic agents . Chemotherapeutic which may not be structurally similar to bortezomib but agents are known in the art and described in further detail mimic the therapeutic activity of bortezomib or structurally herein . Examples of chemotherapeutic agents are set forth in similar compounds in vivo . Table A . As with proteasome inhibition therapy , new classes Proteasome inhibitors for use in the practice of the of cancer therapies may be combined with glucocorticoid invention include additional peptide boronic acids such as 10 therapy regimens as they are developed . Finally , the meth those disclosed in Adams et al ., U . S . Pat. No. 5 , 780 ,454 ods of the invention include combination of proteasome ( 1998 ), U . S . Pat . No . 6 ,066 ,730 (2000 ), U . S . Pat . No . inhibition therapy with glucocorticoid therapy, either alone, 6 , 083, 903 ( 2000 ) , U . S . Pat. No. 6 , 548, 668 ( 2003 ) , and or in conjunction with further agents . Siman et al. WO 91/ 13904 , each of which is hereby incor - In one aspect, one or more of the markers listed in any one porated by reference in its entirety , including all compounds 15 of Table 1A , Table 1B , Table 2A , Table 2B , and /or Table 3 , and formulae disclosed therein . Preferably , a boronic acid can be used to identify candidate agents for use in a compound for use in the present invention is selected from treatment regimen which will produce a response in a the group consisting of: N - (4 -morpholine ) carbonyl. -beta .- patient. For example , the method can identify an agent or a ( 1 -naphthyl ) - L -alanine - L - leucine boronic acid ; N - ( 8 - quino - combination of agents useful as a proteasome inhibitor. In line ) sulfonyl- . beta .- (1 -naphthyl ) - L -alanine - L -alanine - L - 20 another example , the method can identify an agent or leucine boronic acid ; N - ( 2 -pyrazine ) carbonyl- L - combination of glucocorticoids . In another example , the phenylalanine - L -leucine boronic acid , and N - ( 4 - method can identify a set of patients likely to be non morpholine ) carbonyl- [ O - ( 2 -pyridylmethyl ) ] - L - tyrosine - L responsive to current therapies , and therefore good candi leucine boronic acid . dates for inclusion in a clinical trial of a drug aimed at Additionally , proteasome inhibitors include peptide alde - 25 meeting the unmet need of non -responsive patients . For hyde proteasome inhibitors such as those disclosed in Stein example , a marker or marker set associated with non et al. U . S . Pat. No . 5 ,693 ,617 (1997 ) , and International response to bortezomib can identify a patient or a test system patent publications WO 95 /24914 published Sep . 21 , 1995 comprising the capacity to express the marker or marker set . and Siman et al. WO 91/ 13904 published Sep . 19 , 1991 ; The method can identify a candidate agent which achieves Iqbal et al . J . Med . Chem . 38 : 2276 -2277 ( 1995 ) , as well as 30 a response in such a patient or test system . In the method , an Bouget et al. Bioorg Med Chem 17 : 4881 - 4889 (2003 ) each assay composition comprising a cell, e . g . a tumor cell , of which is hereby incorporated by reference in its entirety, capable of expressing a marker or a plurality of markers including all compounds and formulae disclosed therein . listed in any one of Table 1A , Table 1B , Table 2A , Table 2B , Further, proteasome inhibitors include lactacystin and and/ or Table 3 is contacted with the test agent, e . g . for an lactacycstin analogs which have been disclosed in Fentany 35 amount of time for the test agent to affect the level of marker, et al, U . S . Pat. No. 5 , 756 ,764 ( 1998 ) , and U . S . Pat . No. detecting the level of the marker and comparing the level to 6 , 147, 223 (2000 ) , Schreiber et al U . S . Pat. No. 6 ,645 , 999 the level in a reference cell , e . g ., a cell contacted with a ( 2003 ) , and Fenteany et al . Proc . Natl . Acad . Sci . USA known proteasome inhibitor ( e . g ., bortezomib ) or glucocor ( 1994 ) 91 : 3358, each of which is hereby incorporated by ticoid ( e .g ., dexamethasone ) or a normal cell , and identify reference in its entirety , including all compounds and for - 40 ing the agent as a candidate proteasome inhibitor or gluco mulae disclosed therein . corticoid if the test agent produces an informative Additionally , synthetic peptide vinyl sulfone proteasome expression level of the marker or markers typical of a inhibitors and epoxyketone proteasome inhibitors have been responsive patient. Conversely, the test agent may not be disclosed and are useful in the methods of the invention . See , identified as a candidate agent if it is used in the method and e . g ., Bogyo et al. , Proc . Natl. Acad . Sci. 94 :6629 ( 1997 ) ; 45 produces an informative expression level typical of a non Spaltenstein et al. Tetrahedron Lett . 37: 1343 ( 1996 ) ; Meng responsive patient. The assay composition can comprise a L , Proc . Natl . Acad Sci 96 : 10403 ( 1999 ) ; and Meng L H , tumor cell isolated from a patient with cancer, e . g . a hema Cancer Res 59 : 2798 ( 1999 ), each of which is hereby tological cancer ( e . g . , multiple myeloma, leukemias, lym incorporated by reference in its entirety . phoma , etc ) or cancer from a solid tumor ( e . g . , in lung , Still further, naturally occuring compounds have been 50 breast, prostate , ovary , colon , kidney or liver ) . Alternatively , recently shown to have proteasome inhibition activity can be the assay composition can comprise a tumor cell line . The used in the present methods. For example , TMC - 95A , a composition comprising the cell can be an in vivo tumor cyclic peptide , or Gliotoxin , both fungal metabolites or model, e . g . an immunocompromised mouse or a rat with an polyphenols compounds found in green tea have been iden ectopic , e . g . subcutaneous or ascites, tumor, e . g . a human tified as proteasome inhibitors . See , e . g . , Koguchi Y , Anti - 55 tumor. The assay composition can be a human subject. biot ( Tokyo ) 53: 105 . ( 2000 ); Kroll M , Chem Biol 6 :689 Further to the above, the language, proteasome inhibition ( 1999 ) ; and Nam S , J . Biol Chem 276 : 13322 ( 2001 ) , each of therapy regimen and /or glucocorticoid therapy regimen can which is hereby incorporated by reference in its entirety . include additional agents in addition to proteasome inhibi Additional therapeutic agents for use in the methods of tion agents , including chemotherapeutic agents . A “ chemo the invention comprise a known class of therapeutic agents 60 therapeutic agent” is intended to include chemical reagents comprising glucocorticoid steroids. Glucocorticoid therapy, which inhibit the growth of proliferating cells or tissues generally comprises at least one glucocorticoid agent ( e . g ., wherein the growth of such cells or tissues is undesirable. dexamethasone ). In certain applications of the invention , the Chemotherapeutic agents such as anti -metabolic agents , agent used in methods of the invention is a glucocorticoid e . g ., Ara AC , 5 - FU and methotrexate , antimitotic agents , agent. One example of a glucocorticoid utilized in the 65 e . g . , taxane , vinblastine and vincristine , alkylating agents , treatment of multiple myeloma patients as well as other e . g . , melphanlan , BCNU and nitrogen mustard , Topoi cancer therapies is dexamethasone . Additional glucocorti somerase II inhibitors , e . g . , VW - 26 , topotecan and Bleomy US 9 , 963, 747 B2 33 34 cin , strand -breaking agents , e . g ., doxorubicin and DHAD , different mechanisms of action , e . g . , the use of an anti cross -linking agents , e . g . , cisplatin and CBDCA , radiation mitotic agent in combination with an alkylating agent and a and ultraviolet light. In a preferred embodiment, the agent is proteasome inhibitor . a proteasome inhibitor ( e . g . , bortezomib or other related The agents disclosed herein may be administered by any compounds ) . are well known in the art ( see e . g . , Gilman A . 5 route , including intradermally , subcutaneously , orally , G ., et al. , The Pharmacological Basis of Therapeutics, 8th intraarterially or intravenously . Preferably , administration Ed ., Sec 12: 1202- 1263 ( 1990 )) , and are typically used to will be by the intravenous route . Preferably parenteral treat neoplastic diseases . The chemotherapeutic agents gen - administration may be provided in a bolus or by infusion . erally employed in chemotherapy treatments are listed The concentration of a disclosed compound in a pharma below in Table A . 10 ceutically acceptable mixture will vary depending on several The agents tested in the present methods can be a single factors , including the dosage of the compound to be admin agent or a combination of agents . For example , the present istered , the pharmacokinetic characteristics of the methods can be used to determine whether a single chemo - compound ( s ) employed , and the route of administration . The therapeutic agent, such as methotrexate , can be used to treat agent may be administered in a single dose or in repeat a cancer or whether a combination of two or more agents can 15 doses . Treatments may be administered daily or more fre be used in combination with a proteasome inhibitor (e . g ., quently depending upon a number of factors, including the bortezomib ) and / or a glucocorticoid agent ( e . g ., dexametha overall health of a patient, and the formulation and route of sone ). Preferred combinations will include agents that have administration of the selected compound (s ) . TABLE A Chemotherapeutic Agents TYPE OF NONPROPRIETARY NAMES CLASS AGENT (OTHER NAMES ) Alkylating Nitrogen Mechlorethamine (HN2 ) Mustards Cyclophosphamide Ifosfamide Melphalan ( L -sarcolysin ) Chlorambucil Ethylenimines Hexamethylmelamine And Thiotepa Methylmelamines Alkyl Sulfonates Busulfan Alkylating Nitrosoureas Carmustine ( BCNU ) Lomustine (CCNU ) Semustine (methyl - CCNU ) Streptozocin ( streptozotocin ) Alkylating Triazenes Decarbazine (DTIC ; dimethyltriazenoimi dazolecarboxamide ) Alkylator cis -diamminedichloroplatinum II (CDDP ) Antimetab Folic Acid Methotrexate (amethopterin ) olites Analogs Pyrimidine Fluorouracil (' 5 - fluorouracil ; 5 -FU ) Analogs Floxuridine ( fluorode -oxyuridine ; FUR ) Cytarabine ( cytosine arabinoside ) Purine Analogs Mercaptopuine (6 -mercaptopurine ; 6 -MP ) and Related Thioguanine (6 - thioguanine ; TG ) Inhibitors Pentostatin ( 2' - deoxycoformycin ) Vinca Alkaloids Vinblastin (VLB ) Vincristine Topoisomerase Etoposide Inhibitors Teniposide Camptothecin Topotecan 9 -amino - campotothecin CPT- 11 Natural Antibiotics Dactinomycin (actinomycin D ) Products Adriamycin Daunorubicin (daunomycin ; rubindomycin ) Doxorubicin Bleomycin Plicamycin (mithramycin ) Mitomycin (mitomycin C ) TAXOL Taxotere Enzymes L - Asparaginase Natural Biological Interfon alfa Products Response Interleukin 2 Modifiers Platinum cis - diamminedichloroplatinum II ( CDDP) Coordination Carboplatin Complexes Anthracendione Mitoxantrone Substituted Urea Hydroxyurea US 9, 963, 747 B2 35 36 TABLE A - continued Chemotherapeutic Agents TYPE OF NONPROPRIETARY NAMES CLASS AGENT (OTHER NAMES) Miscellaneous Methyl Hydraxzine Procarbazine Agents Derivative ( N -methylhydrazine , (MIH ) Adrenocortical Mitotane ( 0 , p '- DDD ) Suppressant Aminoglutethimide Hormones and Progestins Hydroxyprogesterone caproate Antagonists Medroxyprogesterone acetate Megestrol acetate Estrogens Diethylstilbestrol Ethinyl estradiol Antiestrogen Tamoxifen Androgens Testosterone propionate Fluoxymesterone Antiandrogen Flutamide Gonadotropin Leuprolide releasing Hormone analog

Isolated Nucleic Acid Molecules, Vectors and Host Cells thereto , e . g . , a radioisotope , a fluorescent compound , an One aspect of the invention pertains to isolated nucleic enzyme, or an enzyme co - factor. Such probes can be used as acid molecules that correspond to a predictive marker of the part of a diagnostic test kit for identifying cells or tissues invention , including nucleic acids which encode a polypep - 25 which express the protein , such as by measuring levels of a tide corresponding to a predictive marker of the invention or nucleic acid molecule encoding the protein in a sample of a portion of such a polypeptide . Isolated nucleic acids of the cells from a subject, e. g . , detecting mRNA levels or deter invention also include nucleic acid molecules sufficient for mining whether a gene encoding the protein has been use as hybridization probes to identify nucleic acid mol- mutated or deleted . ecules that correspond to a predictive marker of the inven - 30 In addition to the nucleotide sequences described in the tion , including nucleic acids which encode a polypeptide database records described herein , it will be appreciated by corresponding to a predictive marker of the invention , and those skilled in the art that DNA sequence polymorphisms fragments of such nucleic acid molecules , e . g . , those suit - that lead to changes in the amino acid sequence can exist able for use as PCR primers for the amplification or mutation within a population ( e . g . , the human population ) . Such of nucleic acid molecules . As used herein , the term “ nucleic 35 genetic polymorphisms can exist among individuals within acid molecule ” is intended to include DNA molecules ( e . g . , a population due to naturally occurring allelic variation . An cDNA or genomic DNA ) and RNA molecules ( e . g ., mRNA ) allele is one of a group of genes which occur alternatively at and analogs of the DNA or RNA generated using nucleotide a given genetic locus . In addition , it will be appreciated that analogs . The nucleic acid molecule can be single- stranded or DNA polymorphisms that affect RNA expression levels can double - stranded , but preferably is double -stranded DNA . 40 also exist that may affect the overall expression level of that A nucleic acid molecule of the present invention , e . g . , a gene ( e . g . , by affecting regulation or degradation ) . nucleic acid encoding a protein corresponding to a marker As used herein , the terms " gene ” and “ recombinant gene ” listed in any one of Table 1A , Table 1B , Table 2A , Table 2B , refer to nucleic acid molecules comprising an open reading and / or Table 3 , can be isolated and manipulated ( e . g ., frame encoding a polypeptide corresponding to a marker of amplified , cloned , synthesized , etc . ) using standard molecu - 45 the invention , including, e . g . , sequences which differ , due to lar biology techniques and the sequence information in the degeneracy of the genetic code , from the nucleotide database records described herein . ( e . g . , described in Sam - sequence of nucleic acids encoding a protein which corre brook et al. , ed . , Molecular Cloning : A Laboratory Manual, sponds to a marker of the invention , and thus encode the 2nd ed ., Cold Spring Harbor Laboratory Press , Cold Spring same protein . Harbor, N . Y ., 1989 ) . 50 As used herein , the phrase “ allelic variant ” refers to a Moreover , a nucleic acid molecule of the invention can nucleotide sequence which occurs at a given locus or to a comprise only a portion of a nucleic acid sequence , wherein polypeptide encoded by the nucleotide sequence . Such natu the full length nucleic acid sequence comprises a predictive rally occurring allelic variations can typically result in 1 - 5 % marker of the invention or which encodes a polypeptide variance in the nucleotide sequence of a given gene . Alter corresponding to a marker of the invention . Such nucleic 55 native alleles can be identified by sequencing the gene of acids can be used , for example , as a probe or primer. The interest in a number of different individuals . This can be probe/ primer typically is used as one or more substantially readily carried out by using hybridization probes to identify purified oligonucleotides . The oligonucleotide typically the same genetic locus in a variety of individuals . Any and comprises a region of nucleotide sequence that hybridizes all such nucleotide variations and resulting amino acid under stringent conditions to at least about 7 , preferably 60 polymorphisms or variations that are the result of naturally about 15 , more preferably about 25 , 50 , 75 , 100 , 125 , 150 , occurring allelic variation and that do not alter the functional 175 , 200 , 250 , 300 , 350 , or 400 or more consecutive activity are intended to be within the scope of the invention . nucleotides of a nucleic acid of the invention . The present invention encompasses antisense nucleic acid Probes based on the sequence of a nucleic acid molecule molecules , i . e . , molecules which are complementary to a of the invention can be used to detect transcripts or genomic 65 sense nucleic acid of the invention , e .g ., complementary to sequences corresponding to one or more predictive markers the coding strand of a double -stranded DNA molecule of the invention . The probe comprises a label group attached corresponding to a marker of the invention or complemen US 9 , 963, 747 B2 37 38 tary to an mRNA sequence corresponding to a marker of the protocols as described in Hyrup et al. ( 1996 ), supra ; Perry invention . Accordingly, an antisense nucleic acid of the O 'Keefe et al. (1996 ) Proc. Natl . Acad . Sci. USA 93 :14670 invention can hydrogen bond to ( i. e . anneal with ) a sense 675 . nucleic acid of the invention . The antisense nucleic acid can PNAs can be used in therapeutic and diagnostic applica be complementary to an entire coding strand , or to only a 5 tions. For example , PNAs can be used , e . g . , in the analysis portion thereof , e . g ., all or part of the protein coding region of single base pair mutations in a gene by, e . g . , PNA directed PCR clamping ; as artificial restriction enzymes when used in ( or open reading frame ). An antisense nucleic acid molecule combination with other enzymes , e . g . , S1 nucleases (Hyrup can also be antisense to all or part of a non -coding region of (1996 ) , supra ; or as probes or primers for DNA sequence and the coding strand of a nucleotide sequence encoding *a * 10 hybridization (Hyrup , 1996 , supra ; Perry - O 'Keefe et al ., polypeptide of the invention . The non - coding regions (“ 5 ! 1996 , Proc . Natl. Acad . Sci. USA 93 : 14670 -675 ) . and 3 ' untranslated regions" ) are the 5 ' and 3 ' sequences In another aspect, PNAs can be modified , e . g ., to enhance which flank the coding region and are not translated into their stability or cellular uptake , by attaching lipophilic or amino acids . other helper groups to PNA , by the formation of PNA -DNA An antisense oligonucleotide can be, Torfor example , about 15 chimeras, or by the use of liposomes or other techniques of 5 , 10 , 15 , 20 , 25 , 30 , 35 , 40 , 45 , or 50 or more nucleotides drug delivery known in the art . For example , PNA -DNA in length . An antisense nucleic acid of the invention can be chimeras can be generated which can combine the advan constructed using chemical synthesis and enzymatic ligation tageous properties of PNA and DNA . Such chimeras allow reactions using procedures known in the art . For example , an DNA recognition enzymes, e . g . , RNASE H and DNA poly antisense nucleic acid ( e . g . , an antisense oligonucleotide ) 20 merases, to interact with the DNA portion while the PNA can be chemically synthesized using naturally occurring portion would provide high binding affinity and specificity . nucleotides or variously modified nucleotides designed to PNA -DNA chimeras can be linked using linkers of appro increase the biological stability of the molecules or to priate lengths selected in terms of base stacking, number of increase the physical stability of the duplex formed between bonds between the nucleobases , and orientation (Hyrup , the antisense and sense nucleic acids, e . g . , phosphorothioate 25 1996 , supra ) . The synthesis of PNA -DNA chimeras can be derivatives and acridine substituted nucleotides can be used . performed as described in Hyrup ( 1996 ) , supra , and Finn et Examples of modified nucleotides which can be used to al. ( 1996 ) Nucleic Acids Res. 24 ( 17 ) : 3357 -63 . For example , parila DNA chain can be synthesized on a solid support using generate the antisense nucleic acid include 5 - fluorouracil , standard phosphoramidite coupling chemistry and modified 5 - bromouracil, 5 - chlorouracil , 5 - iodouracil, hypoxanthinema , 30 nucleoside analogs. Compounds such as 5 '- ( 4 -methoxytri xanthine, 4 - acetylcytosine , 5 - ( carboxyhydroxylmethyl) ura tyl) amino - 5 ' -deoxy - thymidine phosphoramidite can be used cil , 5 -carboxymethylaminomethyl - 2 - thiouridine, 5 -car as a link between the PNA and the 5 ' end of DNA (Mag et boxymethylaminomethyluracil , dihydrouracil , beta - D - ga al. , 1989, Nucleic Acids Res . 17 :5973 -88 ). PNA monomers lactosylqueosine , inosine , N6 -isopentenyladenine , are then coupled in a step -wise manner to produce a chi 1 -methylguanine , 1 -methylinosine , 2 , 2 - dimethylguaninenyiguan .ne , a35 meric molecule with a 5 ' PNA segment and a 3 ' DNA 2 -methyladenine , 2 -methylguanine , 3 -methylcytosine , segment (Finn et al. , 1996 , Nucleic Acids Res . 24 ( 17 ) :3357 5 - methylcytosine , N6 - adenine, 7 -methylguanine , 5 -methyl 63 ) . Alternatively , chimeric molecules can be synthesized aminomethyluracil , 5 -methoxyaminomethyl - 2 - thiouracil, with a 5 ' DNA segment and a 3 ' PNA segment (Peterser et beta - D -mannosylqueosine , 5 ' -methoxycarboxymethyluracil , al. , 1975 . Bioorganic Med . Chem . Lett . 5 : 1119 - 11124 ) . 5 -methoxyuracil , 2 -methylthio - N6 - isopentenyladenine , ura - 40 The oligonucleotide can include other appended groups cil - 5 -oxyacetic acid ( v ) , wybutoxosine , pseudouracil , queo - such as peptides ( e . g . , for targeting host cell receptors in sine, 2 - thiocytosine, 5 -methyl - 2 - thiouracil, 2 - thiouracil, vivo ) , or agents facilitating transport across the cell mem 4 - thiouracil , 5 -methyluracil , uracil -5 - oxyacetic acid methy - brane ( see , e . g. , Letsinger et al ., 1989 , Proc. Natl. Acad . Sci . lester , uracil - 5 - oxyacetic acid ( v ) , 5 -methyl - 2 - thiouracil, USA 86 :6553 -6556 ; Lemaitre et al. , 1987 , Proc . Natl. Acad . 3 - ( 3 - amino - 3 - N - 2 - carboxypropyl) uracil , ( acp3 ) w , and 2 , 6 - 45 Sci . USA 84 : 648 -652 ; PCT Publication No . WO 88 /09810 ) diaminopurine. Alternatively , the antisense nucleic acid can or the blood -brain barrier ( see , e . g ., PCT Publication No . be produced biologically using an expression vector into WO 89 / 10134 ) . In addition , oligonucleotides can be modi which a nucleic acid has been sub - cloned in an antisense fied with hybridization - triggered cleavage agents ( see , e . g . , orientation ( i . e . , RNA transcribed from the inserted nucleic Krol et al. , 1988 , Bio / Techniques 6 : 958 - 976 ) or intercalating acid will be of an antisense orientation to a target nucleic 50 agents ( see , e . g ., Zon , 1988 , Pharm . Res. 5 :539 -549 ) . To this acid of interest, described further in the following subsec - end , the oligonucleotide can be conjugated to another mol tion ) . ecule , e . g. , a peptide , hybridization triggered cross -linking The nucleic acid molecules of the invention can be agent, transport agent , hybridization - triggered cleavage modified at the base moiety , sugar moiety or phosphate agent , etc . backbone to improve, e . g . , the stability , hybridization , or 55 The invention also includes molecular beacon nucleic solubility of the molecule . For example , the deoxyribose acids having at least one region which is complementary to phosphate backbone of the nucleic acids can be modified to a marker of the invention , such that the molecular beacon is generate peptide nucleic acids (see Hyrup et al ., 1996 , useful for quantitating the presence of the predictive marker Bioorganic & Medicinal Chemistry 4 ( 1 ) : 5 - 23 ) . As used of the invention in a sample . A “ molecular beacon ” nucleic herein , the terms “ peptide nucleic acids” or “ PNAs" refer to 60 acid is a nucleic acid comprising a pair of complementary nucleic acid mimics , e . g . , DNA mimics , in which the deoxy - regions and having a fluorophore and a fluorescent quencher ribose phosphate backbone is replaced by a pseudopeptide associated therewith . The fluorophore and quencher are backbone and only the four natural nucleobases are retained . associated with different portions of the nucleic acid in such The neutral backbone of PNAs has been shown to allow for an orientation that when the complementary regions are specific hybridization to DNA and RNA under conditions of 65 annealed with one another, fluorescence of the fluorophore low ionic strength . The synthesis of PNA oligomers can be is quenched by the quencher. When the complementary performed using standard solid phase peptide synthesis regions of the nucleic acid are not annealed with one US 9 ,963 , 747 B2 39 40 another, fluorescence of the fluorophore is quenched to a sequences homologous to a nucleic acid molecules of the lesser degree . Molecular beacon nucleic acids are described , invention . BLAST protein searches can be performed with for example , in U .S . Pat. No . 5 ,876 ,930 . the XBLAST program , score = 50 , wordlength = 3 to obtain Vectors , including expression vectors , containing a amino acid sequences homologous to a protein molecules of nucleic acid encoding a polypeptide corresponding to a 5 the invention . To obtain gapped alignments for comparison predictive marker of the invention can be used for produc - purposes , Gapped BLAST can be utilized as described in tion of nucleic acid and proteins corresponding to predictive Altschul et al . ( 1997 ) Nucleic Acids Res . 25 : 3389 - 3402 . markers of the invention ; as well as for production of Alternatively , PSI- Blast can be used to perform an iterated compositions relating to the predictive markers . Useful search which detects distant relationships between mol vectors further comprise promoter and /or regulatory 10 ecules. When utilizing BLAST, Gapped BLAST, and PSI sequences for effective expression of the nucleic acid and / or Blast programs, the default parameters of the respective protein corresponding to the predictive marker of interest. In programs ( e . g . , XBLAST and NBLAST ) can be used (acces certain instances, promoters can include constitutive pro - sible at the website maintained by National Center for moter/ regulatory sequences , inducible promoter/ regulatory Biotechnology Information , Bethesda , Md. , USA . Another sequences, tissue specific promoter/ regulatory sequences , or 15 example of a mathematical algorithm utilized for the com the naturally occurring endogenous promoter/ regulatory parison of sequences is the algorithm of Myers and Miller, sequences corresponding to the predictive marker of inter ( 1988 ) CABIOS 4 : 11 - 17 . Such an algorithm is incorporated est , as required . Various expression vectors are well known into the ALIGN program (version 2 . 0 ) which is part of the in the art and can be adapted to suit the particular system for GCG sequence alignment software package . When utilizing expression . For example , recombinant expression vectors of 20 the ALIGN program for comparing amino acid sequences, a the invention can be designed for expression of a polypep - PAM120 weight residue table , a gap length penalty of 12 , tide corresponding to a marker of the invention in prokary - and a gap penalty of 4 can be used . Yet another useful otic (e .g . , E . coli ) or eukaryotic cells ( e. g ., insect cells ( using algorithm for identifying regions of local sequence similar baculovirus expression vectors }, yeast cells or mammalian ity and alignment is the FASTA algorithm as described in cells ) . Suitable host cells are known in the art and include 25 Pearson and Lipman ( 1988 ) Proc . Natl. Acad . Sci. USA those discussed in Goeddel , supra . Alternatively , the recom 85 : 2444 -2448 . When using the FASTA algorithm for com binant expression vector can be transcribed and translated in paring nucleotide or amino acid sequences, a PAM120 vitro , for example using T7 promoter regulatory sequences weight residue table can , for example , be used with a k - tuple and T7 polymerase . Vectors and host cells can be produced value of 2 . The percent identity between two sequences can using routine methodology known in the art. Furthermore , 30 be determined using techniques similar to those described use of vectors and host cells can be utilized for production above , with or without allowing gaps . In calculating percent of nucleic acids, polypeptides and fragments thereof corre - identity , only exact matches are counted . sponding to markers of the invention . The invention also provides chimeric or fusion proteins Isolated Proteins and Antibodies corresponding to a marker of the invention . As used herein , One aspect of the invention pertains to isolated proteins 35 a " chimeric protein ” or “ fusion protein ” comprises all or part which correspond to predictive markers of the invention , (preferably a biologically active part ) of a polypeptide and biologically active portions thereof, as well as polypep corresponding to a marker of the invention operably linked tide fragments suitable for use as immunogens to raise to a heterologous polypeptide ( i . e . , a polypeptide other than antibodies directed against a polypeptide corresponding to a the polypeptide corresponding to the marker ) . Within the predictive marker of the invention . Polypeptides for use in 40 fusion protein , the term " operably linked ” is intended to the invention can be isolated , purified , or produced using the indicate that the polypeptide of the invention and the het gene identification information provided herein in combi- erologous polypeptide are fused in - frame to each other. The nation with routine molecular biology, protein purification heterologous polypeptide can be fused to the amino - termi and recombinant DNA techniques well known in the art . nus or the carboxyl- terminus of the polypeptide of the Preferred polypeptides have the amino acid sequence 45 invention . Useful fusion proteins can include GST , c -myc , listed in the one of the GenBank and Entrez database records FLAG , HA , and any other well known heterologous tag for described herein . Other useful proteins are substantially use in fusion protein production . Such fusion proteins can identical ( e . g ., at least about 70 % , preferably 80 % , 90 % , facilitate the purification of a recombinant polypeptide of the 95 % , or 99 % to one of these sequences and retain the invention . functional activity of the protein of the corresponding natu - 50 In addition , fusion proteins can include a signal sequence rally - occurring protein yet differ in amino acid sequence due from another protein such as gp67, melittin , human placental to natural allelic variation or mutagenesis . alkaline phosphatase , and phoA . In yet another aspect, the The determination of percent identity between two fusion protein is an immunoglobulin fusion protein in which sequences can be accomplished using a mathematical algo - all or part of a polypeptide corresponding to a predictive rithm determining the number of identical positions shared 55 marker of the invention is fused to sequences derived from between two sequences . Determination can be carried out a member of the immunoglobulin protein family . The immu using any known method in the art for comparison of noglobulin fusion proteins of the invention can be used as identity and similarity . Examples of methods used can immunogens to produce antibodies directed against a poly include for example , a mathematical algorithm utilized for peptide of the invention in a subject , to purify ligands and in the comparison of two sequences is the algorithm of Karlin 60 screening assays to identify molecules which inhibit the and Altschul ( 1990 ) Proc . Natl . Acad . Sci. USA 87 :2264 - interaction of receptors with ligands. 2268, modified as in Karlin and Altschul ( 1993 ) Proc. Natl . An isolated polypeptide corresponding to a predictive Acad . Sci. USA 90 : 5873 -5877 . Such an algorithm is incor - marker of the invention , or a fragment thereof , can be used porated into the NBLAST and ) (BLAST programs of Alts - as an immunogen to generate antibodies using standard chul, et al. (1990 ) J . Mol. Biol. 215 :403 - 410 . BLAST 65 techniques for polyclonal and monoclonal antibody prepa nucleotide searches can be performed with the NBLAST ration . For example , an immunogen typically is used to program , score = 100 , wordlength = 12 to obtain nucleotide prepare antibodies by immunizing a suitable ( i e immuno US 9 , 963 , 747 B2 42 competent ) subject such as a rabbit, goat , mouse , or other standard recombinant DNA techniques, are within the scope mammal or vertebrate . An appropriate immunogenic prepa of the invention . A chimeric antibody is a molecule in which ration can contain , for example , recombinantly - expressed or different portions are derived from different animal species , chemically - synthesized polypeptide . The preparation can such as those having a variable region derived from a murine further include an adjuvant , such as Freund ' s complete or 5 mAb and a human immunoglobulin constant region . (See , incomplete adjuvant, or a similar immunostimulatory agent. e . g . , Cabilly et al. , U . S . Pat. No . 4 , 816 , 567 ; and Boss et al. , Accordingly , another aspect of the invention pertains to U . S . Pat. No . 4 ,816 , 397 , which are incorporated herein by antibodies directed against a polypeptide of the invention . reference in their entirety . ) Humanized antibodies are anti The terms “ antibody ” and “ antibody substance ” as used body molecules from non -human species having one or interchangeably herein refer to immunoglobulin molecules 10 more complementarily determining regions (CDRs ) from and immunologically active portions of immunoglobulin the non -human species and a framework region from a molecules , i. e. , molecules that contain an antigen binding human immunoglobulin molecule . (See , e . g ., Queen , U . S . site which specifically binds an antigen , such as a polypep - Pat. No . 5 ,585 ,089 , which is incorporated herein by refer tide of the invention , e . g . , an epitope of a polypeptide of the ence in its entirety .) Such chimeric and humanized mono invention . A molecule which specifically binds to a given 15 clonal antibodies can be produced by recombinant DNA polypeptide of the invention is a molecule which binds the techniques known in the art, for example using methods polypeptide , but does not substantially bind other molecules described in PCT Publication No. WO 87 /02671 ; European in a sample , e . g. , a biological sample , which naturally Patent Application 184 ,187 ; European Patent Application contains the polypeptide. Examples of immunologically 171, 496 ; European Patent Application 173 ,494 ; PCT Pub active portions of immunoglobulin molecules include F ( ab ) 20 lication No. WO 86 /01533 ; U . S . Pat . No . 4 ,816 , 567 ; Euro and F (ab ' ) , fragments which can be generated by treating the pean Patent Application 125 , 023 ; Better et al. ( 1988 ) Sci antibody with an enzyme such as pepsin . The invention ence 240 : 1041 - 1043 ; Liu et al. ( 1987 ) Proc . Natl . Acad . Sci. provides polyclonal and monoclonal antibodies . Synthetic USA 84 : 3439 - 3443 ; Liu et al . (1987 ) J . Immunol. 139 : 3521 and genetically engineered variants ( See U . S . Pat. No. 3526 ; Sun et al. ( 1987 ) Proc . Natl. Acad . Sci. USA 84 : 214 6 ,331 , 415 ) of any of the foregoing are also contemplated by 25 218 ; Nishimura et al . ( 1987 ) Cancer Res . 47: 999 - 1005 ; the present invention . Polyclonal and monoclonal antibodies Wood et al. ( 1985 ) Nature 314 : 446 -449 ; and Shaw et al . can be produced by a variety of techniques , including ( 1988 ) J . Natl. Cancer Inst. 80 : 1553 - 1559 ) ; Morrison conventional murine monoclonal antibody methodology ( 1985 ) Science 229 : 1202 - 1207; Oi et al. ( 1986 ) Bio / Tech e . g ., the standard somatic cell hybridization technique of niques 4 : 214 ; U . S . Pat. No . 5 , 225 , 539 ; Jones et al . ( 1986 ) Kohler and Milstein , Nature 256 : 495 ( 1975 ) the human B 30 Nature 321 : 552 -525 ; Verhoeyan et al. ( 1988 ) Science 239 : cell hybridoma technique (see Kozbor et al. , 1983, Immunol. 1534 ; and Beidler et al. ( 1988 ) J . Immunol. 141: 4053 - 4060 . Today 4 : 72 ), the EBV -hybridoma technique (see Cole et al. , Methods for making human antibodies are known in the pp . 77 - 96 In Monoclonal Antibodies and Cancer Therapy , art . One method for making human antibodies employs the Alan R . Liss , Inc . , 1985 ) or trioma techniques. See generally , use of transgenic animals , such as a transgenic mouse. These Harlow , E . and Lane , D . ( 1988 ) Antibodies : A Laboratory 35 transgenic animals contain a substantial portion of the Manual, Cold Spring Harbor Laboratory Press , Cold Spring human antibody producing genome inserted into their own Harbor, N . Y .; and Current Protocols in Immunology , Coli - genome and the animal' s own endogenous antibody produc gan et al . ed ., John Wiley & Sons, New York , 1994 . tion is rendered deficient in the production of antibodies. Preferably , for diagnostic applications , the antibodies are Methods for making such transgenic animals are known in monoclonal antibodies . Additionally , for use in in vivo 40 the art. Such transgenic animals can be made using XENO applications the antibodies of the present invention are MOUSETM technology or by using a " minilocus ” approach . preferably human or humanized antibodies . Hybridoma cells Methods for making XENOMICETM are described in U . S . producing a monoclonal antibody of the invention are Pat . Nos . 6 , 162 , 963 , 6 , 150 , 584 , 6 , 114 ,598 and 6 ,075 , 181 , detected by screening the hybridoma culture supernatants which are incorporated herein by reference . Methods for for antibodies that bind the polypeptide of interest, e . g . , 45 making transgenic animals using the “ minilocus ” approach using a standard ELISA assay. are described in U . S . Pat. Nos . 5 , 545 , 807 , 5 ,545 ,806 and If desired , the antibody molecules can be harvested or 5 ,625 , 825 ; also see International Publication No. W093 / isolated from the subject (e .g ., from the blood or serum of 12227 , which are each incorporated herein by reference . the subject) and further purified by well - known techniques, Antibody fragments may be derived from any of the such as protein A chromatography to obtain the IgG fraction . 50 antibodies described above . For example , antigen -binding Alternatively , antibodies specific for a protein or polypep - fragments , as well as full - length monomeric , dimeric or tide of the invention can be selected or ( e . g. , partially trimeric polypeptides derived from the above - described purified ) or purified by , e . g . , affinity chromatography to antibodies are themselves useful . Useful antibody homologs obtain substantially purified and purified antibody. By a of this type include (i ) a Fab fragment, a monovalent substantially purified antibody composition is meant, in this 55 fragment consisting of the VL , VH , CL and CH1 domains ; context, that the antibody sample contains at most only 30 % ( ii ) a F (ab ' ) , fragment, a bivalent fragment comprising two (by dry weight ) of contaminating antibodies directed against Fab fragments linked by a disulfide bridge at the hinge epitopes other than those of the desired protein or polypep - region ; ( iii ) a Fd fragment consisting of the VH and CH1 tide of the invention , and preferably at most 20 % , yet more domains ; (iv ) a Fv fragment consisting of the VL and VH preferably at most 10 % , and most preferably at most 5 % (by 60 domains of a single arm of an antibody, (v ) a dAb fragment dry weight) of the sample is contaminating antibodies. A (Ward et al. , Nature 341 : 544 -546 ( 1989 ) ) , which consists of purified antibody composition means that at least 99 % of the a VH domain ; ( vii ) a single domain functional heavy chain antibodies in the composition are directed against the antibody, which consists of a VHH domain (known as a desired protein or polypeptide of the invention . nanobody ) see e . g ., Cortez -Retamozo , et al . , Cancer Res. Additionally , recombinant antibodies , such as chimeric 65 64 : 2853 - 2857 (2004 ) , and references cited therein ; and ( vii ) and humanized monoclonal antibodies, comprising both an isolated complementarity determining region (CDR ) , human and non - human portions, which can be made using e . g ., one or more isolated CDRs together with sufficient US 9 , 963, 747 B2 43 44 framework to provide an antigen binding fragment. Further amino acid sequence of the present invention , an amino acid more , although the two domains of the Fv fragment, VL and sequence which is at least 95 % identical to an amino acid VH , are coded for by separate genes , they can be joined , sequence of the present invention wherein the percent using recombinant methods , by a synthetic linker that identity is determined using the ALIGN program of the enables them to be made as a single protein chain in which 5 GCG software package with a PAM120 weight residue the VL and VH regions pair to form monovalent molecules table , a gap length penalty of 12 , and a gap penalty of 4 ) and (known as single chain Fv (scFv ) ; see e . g ., Bird et al. an amino acid sequence which is encoded by a nucleic acid Science 242 : 423 -426 ( 1988 ) ; and Huston et al . Proc . Natl. molecule which hybridizes to a nucleic acid molecule con Acad . Sci. USA 85 :5879 - 5883 ( 1988 ) . Such single chain sisting of the nucleic acid molecules of the present inven antibodies are also intended to be encompassed within the 10 tion , or a complement thereof, under conditions of hybrid term “ antigen - binding fragment " of an antibody . These ization of 6xSSC at 45° C . and washing in 0 . 2xSSC , 0 . 1 % antibody fragments are obtained using conventional tech - SDS at 65° C . The monoclonal antibodies can be human , niques known to those with skill in the art, and the fragments humanized , chimeric and / or non -human antibodies . are screened for utility in the same manner as are intact The substantially purified antibodies or fragments thereof antibodies . Antibody fragments , such as Fv, F ( ab ' ) , and Fab 15 may specifically bind to a signal peptide, a secreted may be prepared by cleavage of the intact protein , e . g. by sequence , an extracellular domain , a transmembrane or a protease or chemical cleavage . cytoplasmic domain or cytoplasmic membrane of a poly An antibody directed against a polypeptide corresponding peptide of the invention . The substantially purified antibod to a predictive marker of the invention ( e . g . , a monoclonal ies or fragments thereof, the non -human antibodies or frag antibody ) can be used to detect the predictive marker ( e . g ., 20 ments thereof, and / or the monoclonal antibodies or in a cellular sample ) in order to evaluate the level and pattern fragments thereof, of the invention specifically bind to a of expression of the predictive marker. The antibodies can secreted sequence or an extracellular domain of the amino also be used diagnostically to monitor protein levels in acid sequences of the present invention . tissues or body fluids ( e . g . in an tumor sample ) as part of a The invention also provides a kit containing an antibody clinical testing procedure , e . g ., to , for example , determine 25 of the invention conjugated to a detectable substance , and the efficacy of a given treatment regimen . Detection can be instructions for use . Still another aspect of the invention is facilitated by coupling the antibody to a detectable sub - a diagnostic composition comprising an antibody of the stance . Examples of detectable substances include various invention and a pharmaceutically acceptable carrier. In cer enzymes , prosthetic groups , fluorescent materials , lumines - tain aspects , the diagnostic composition contains an anti cent materials , bioluminescent materials , and radioactive 30 body of the invention , a detectable moiety , and a pharma materials . Examples of suitable enzymes include horserad ceutically acceptable carrier . ish peroxidase , alkaline phosphatase, ß -galactosidase , or Sensitivity Assays acetylcholinesterase ; examples of suitable prosthetic group A sample of cancerous cells is obtained from a patient . An complexes include streptavidin /biotin and avidin / biotin ; expression level is measured in the sample for a marker examples of suitable fluorescent materials include umbellif- 35 corresponding to at least one of the predictive markers set erone , fluorescein , fluorescein isothiocyanate, rhodamine , forth in Table 1A , Table 1B , Table 2A , Table 2B , and /or dichlorotriazinylamine fluorescein , dansyl chloride or phy - Table 3. Preferably a marker set is utilized comprising coerythrin ; an example of a luminescent material includes markers identified in Table 1A , Table 1B , Table 2A , Table luminol; examples of bioluminescent materials include 2B , and /or Table 3 , and put together in a marker set using the luciferase , luciferin , and aequorin , and examples of suitable 40 methods described herein . For example , marker sets can radioactive material include 125 I, 131 1, 35S or ' H . comprise the marker sets identified in Table 4 , or any marker Accordingly , in one aspect , the invention provides sub - set prepared by similar methods. Such analysis is used to stantially purified antibodies or fragments thereof, and non - obtain an expression profile of the tumor in the patient. human antibodies or fragments thereof, which antibodies or Evaluation of the expression profile is then used to deter fragments specifically bind to a polypeptide comprising an 45 mine whether the patient is a responsive patient and would amino acid sequence encoded by a predictive marker iden - benefit from proteasome inhibition therapy ( e . g ., treatment tified herein . The substantially purified antibodies of the with a proteasome inhibitor (e . g. , bortezomib ) alone, or in invention , or fragments thereof, can be human , non - human , combination with additional agents ) and /or glucocorticoid chimeric and/ or humanized antibodies . therapy ( e .g ., treatment with a glucocorticoid ( e . g . , dexam In another aspect, the invention provides non -human 50 ethasone ) alone , or in combination with additional agents ) . antibodies or fragments thereof, which antibodies or frag - Evaluation can include use of one marker set prepared using ments specifically bind to a polypeptide comprising an any of the methods provided or other similar scoring meth amino acid sequence which is encoded by a nucleic acid ods known in the art ( e . g . , weighted voting , CTF ) . Still molecule of a predictive marker of the invention . Such further , evaluation can comprise use of more than one non - human antibodies can be goat, mouse , sheep , horse , 55 prepared marker set. A proteasome inhibition therapy and/ or chicken , rabbit , or rat antibodies . Alternatively , the non - glucocorticoid therapy will be identified as appropriate to human antibodies of the invention can be chimeric and /or treat the cancer when the outcome of the evaluation dem humanized antibodies . In addition , the non -human antibod - onstrates decreased non - responsiveness or increased respon ies of the invention can be polyclonal antibodies or mono - siveness in the presence of the agent. clonal antibodies . 60 In one aspect, the invention features a method of evalu In still a further aspect, the invention provides monoclo - ating a patient, e . g ., a patient with cancer, e . g . a hemato nal antibodies or fragments thereof, which antibodies or logical cancer ( e . g . , multiple myeloma , leukemias , lym fragments specifically bind to a polypeptide comprising an phoma, etc ) or cancer from a solid tumor ( e . g ., in lung , amino acid sequence selected from the group consisting of breast , prostate , ovary , colon , kidney , or liver ) for respon the amino acid sequences of the present invention , an amino 65 siveness or non -responsiveness to treatment with a protea acid sequence encoded by the cDNA of the present inven - some inhibition and /or a glucocorticoid therapy regimen The tion , a fragment of at least 15 amino acid residues of an method includes providing an evaluation of the expression US 9 , 963 , 747 B2 45 46 of the markers in a predictive marker set ofmarkers in the or more marker expression levels , e . g. , a predictive marker patient, wherein the predictive marker set has the following or predictive marker set , e . g . , a level of expression associ properties : it includes a plurality of genes, each of which is ated with responsiveness or non - responsiveness to a protea differentially expressed as between patients responsive or some inhibition therapy and / or glucocorticoid therapy ( e . g ., non - responsive to treatment with a proteasome inhibition 5 the informative expression level ) . For example , premiums and / or a glucocorticoid therapy regimen and non -afflicted can be increased ( e .g ., by a certain percentage ) if the subjects and it contains a sufficient number of differentially markers of a patient or a patient ' s predictive marker set expressed markers such that differential expression ( e . g . , as described herein are differentially expressed between an compared to a level in a non - afflicted reference sample ) of insured candidate (or a candidate seeking insurance cover each of the markers in the predictive marker set in a subject 10 age ) and a reference value ( e . g . , a non -afflicted person ) . As is predictive of responsiveness or nonresponsiveness with no another example , premiums can be decreased if levels of a more than about 15 % , about 10 % , about 5 % , about 2 . 5 % , or predictive marker or predictive marker set are sustained (as about 1 % false positives (wherein false positive means described herein ) after treatment with a proteasome inhibitor predicting that a patient as responsive or non - responsive or a glucocorticoid . Premiums can also be scaled depending when the subject is not ) ; and providing a comparison of the 15 on marker expression levels , e . g . , the result of evaluating a expression of each of the markers in the set from the patient predictive marker or predictive marker set described herein . with a reference value , thereby evaluating the patient. For example , premiums can be assessed to distribute risk , Examining the expression of one or more of the identified e . g . , as a function of marker expression levels , e . g . , the result markers or marker sets in a tumor sample taken from a of evaluating a predictive marker or predictive marker set patient during the course of proteasome inhibition therapy 20 described herein . In another example , premiums are and / or glucocorticoid treatment, it is also possible to deter - assessed as a function of actuarial data that is obtained from mine whether the therapeutic agent is continuing to work or patients that are enhanced or non - enhanced responders . whether the cancer has become non -responsive (refractory ) Information about marker expression levels , e . g. , the to the treatment protocol. For example , a patient receiving a result of evaluating a predictive marker or predictive marker treatment of bortezomib would have tumor cells removed 25 set described herein ( e . g . , the informative expression level) , and monitored for the expression of a marker or marker set . can be used , e . g . , in an underwriting process for life insur If the expression profile of one or more marker sets identi - ance . The information can be incorporated into a profile fied in Table 1A , Table 1B , Table 2A , Table 2B , and / or Table about a subject . Other information in the profile can include , 3 demonstrates increased responsiveness in the presence of for example , date of birth , gender, marital status, banking the agent, the treatment with proteasome inhibitor would 30 information , credit information , children , and so forth . An continue . However, if the expression profile of one or more insurance policy can be recommended as a function of the marker sets identified in Table 1A , Table 1B , Table 2A , Table information on marker expression levels, e . g ., the result of 2B , and / or Table 3 demonstrates increased non - responsive evaluating a predictive marker or predictive marker set ness in the presence of the agent, then the cancer may have described herein , along with one or more other items of become resistant to proteasome inhibition therapy and/ or 35 information in the profile . An insurance premium or risk glucocorticoid therapy , and another treatment protocol assessment can also be evaluated as function of the predic should be initiated to treat the patient. tive marker or predictive marker set information . In one Importantly , these determinations can be made on a implementation , points are assigned on the basis of being patient by patient basis or on an agent by agent (or combi responsive or non - responsive to a proteasome inhibition nations of agents ) . Thus , one can determine whether or not 40 therapy and / or glucocorticoid therapy. a particular proteasome inhibition therapy and / or glucocor - In one embodiment, information aboutmarker expression ticoid therapy is likely to benefit a particular patient or levels , e . g . , the result of evaluating a predictive marker or group / class of patients , or whether a particular treatment predictive marker set described herein , is analyzed by a should be continued . function that determines whether to authorize the transfer of Use of Information 45 funds to pay for a service or treatment provided to a subject In one method , information , e . g . , about the patient' s (or make another decision referred to herein ) . For example , marker expression levels ( e . g ., the result of evaluating a the results of analyzing a expression of a predictive marker predictive marker or predictive marker set described herein ), or predictive marker set described herein may indicate that or about whether a patient will be responsive or non - a subject is responsive or non - responsive to a proteasome responsive to a proteasome inhibition therapy and /or gluco - 50 inhibition therapy and/ or glucocorticoid therapy, suggesting corticoid therapy , is provided ( e . g ., communicated , e . g ., that a treatment course is needed , thereby triggering an electronically communicated ) to a third party , e . g . , a hospi- outcome that indicates or causes authorization to pay for a tal, clinic , a government entity , reimbursing party or insur - service or treatment provided to a subject. In one example , ance company ( e . g ., a life insurance company ) . For example , informative expression level of a predictive marker or a choice ofmedical procedure , payment for a medical proce - 55 predictive marker set selected from or derived from Table dure , payment by a reimbursing party , or cost for a service 1A , Table 1B , Table 2A , Table 2B , and Table 3 is determined or insurance can be function of the information . E . g ., the and payment is authorized if the informative expression third party receives the information , makes a determination level identifies a responsive patient. For example , an entity , based at least in part on the information , and optionally e. g. , a hospital, care giver, government entity , or an insur communicates the information or makes a choice of proce - 60 ance company or other entity which pays for , or reimburses dure , payment, level of payment, coverage , etc . based on the medical expenses, can use the outcome of a method information . In the method , informative expression level of described herein to determine whether a party , e. g. , a party a predictive marker or a predictive marker set selected from other than the subject patient, will pay for services ( e . g . , a or derived from Table 1A , Table 1B , Table 2A , Table 2B , and particular therapy ) or treatment provided to the patient. For Table 3 is determined . 65 example , a first entity , e . g ., an insurance company, can use In one embodiment, a premium for insurance ( e . g ., life or the outcome of a method described herein to determine medical) is evaluated as a function of information about one whether to provide financial payment to , or on behalf of, a US 9 , 963 , 747 B2 48 patient, e . g ., whether to reimburse a third party , e . g . , a refractory multiple myeloma ( Protocol M34101 - 039 ). See vendor of goods or services, a hospital, physician , or other Richardson et . al ., N . Engl . J Med . , 352 : 2487 - 2498 ( 2005 ) . care - giver , for a service or treatment provided to a patient. Patients were treated with either bortezomib ( 315 patients ) For example , a first entity , e . g . , an insurance company, can or high - dose dexamethasone (312 patients ) . use the outcome of a method described herein to determine 5 Treatment Dosage and Administration whether to continue , discontinue , enroll an individual in an Drug Supply and Storage insurance plan or program , e. g. , a health insurance or life Bortezomib for injection ( VELCADETM Millennium insurance plan or program . Pharmaceuticals , Inc ., Cambridge , Mass . ), a sterile In one aspect, the disclosure features a method of pro - lyophilized powder for reconstitution , was supplied in vials viding data . The method includes providing data described 10 containing 2 . 5 mg bortezomib and 25 mg mannitol USP . herein , e . g ., generated by a method described herein , to Each vial was reconstituted with 2 . 5 mL of normal ( 0 . 9 % ) provide a record , e. g. , a record described herein , for deter saline, Sodium Chloride Injection USP , such that the recon mining if a payment will be provided . In some embodi- stituted solution contained bortezomib at a concentration of ments , the data is provided by computer, compact disc , 1 mg/mL . The reconstituted solution was clear and colorless telephone, facsimile , email , or letter. In some embodiments , 15 with a final pH between 5 and 6 . the data is provided by a first party to a second party . In some Dexamethasome tablets (DECADRON® Merck & Co . , embodiments, the first party is selected from the subject , a Inc. ) . healthcare provider , a treating physician , a health mainte nance organization (HMO ) , a hospital , a governmental TABLE B entity , or an entity which sells or supplies the drug . In some 20 — embodiments , the second party is a third party payor, an Drug Information insurance company , employer, employer sponsored health ChemicalChemical Name N - Pyrazinecarbonyl - L plan , HMO , or governmental entity . In some embodiments , phenylalanine the first party is selected from the subject, a healthcare L -leucineboronic acid Research Name MLN341 or PS - 341 provider , a treating physician , an HMO , a hospital, an 25 Generic Name bortezomib dexamethasone insurance company , or an entity which sells or supplies the Proprietary Name VELCADE TM Decadron ® drug and the second party is a governmental entity . In some CAS Registry No. 179324 - 69 - 7 312 - 93 - 6 embodiments , the first party is selected from the subject, a U . S . Pat . No . 5 ,780 , 454 Classification Proteasome Inhibitor Steroid healthcare provider, a treating physician , an HMO , a hos Molecular Formula C19H253N404 C22H2. FO pital, an insurance company, or an entity which sells or 30 Molecular Weight 384 . 25 392 .47 supplies the drug and the second party is an insurance Structure Boronic acid derivative Synthetic company. of a leucine adrenocorticosteroid In another aspect, the disclosure features a record ( e . g . , phenylalanine dipeptide computer readable record ) which includes a list and value of expression for the predictive marker or predictive marker set 35 Patients were assigned to receive bortezomib or high - dose for a patient. In some embodiments , the record includes dexamethasone by random allocation at a 1 : 1 ratio . Ran more than one value for each marker. domization was to be stratified , based on the number of lines Exemplification of prior therapy (one prior line versus more than one prior Based on positive findings in multiple myeloma in Phase line of therapy ), time of progression relative to treatment 1 clinical trials (Orlowski , J Clin Oncol. 2002 Nov . 15 ; 40 ( progression while on their most recent therapy or within 6 20 (22 ) : 4420 - 7 . , Aghajanian , Clin Cancer Res . 2002 August ; months of stopping their most recent therapy, or relapse > 6 8 ( 8 ): 2505 - 11, ) Phase 2 myeloma studies were conducted in months after receiving their most recent therapy ) , and order to better to allow a more precise estimate of antitumor screening B2 -microglobulin levels ( > 2 .5 mg/ L versus s2 . 5 activity ofbortezomib in a more homogeneous population of mg/ L ) . patients . The safety and efficacy of bortezomib in subjects 45 Patients assigned to the bortezomib group received treat with multiple myeloma was investigated in two phase 2 ment for a maximum of 273 days . Patients in this treatment clinical studies , M34100 -024 (subjects with first relapse ) group received up to eight 3 -week treatment cycles followed and M34100 - 025 ( subjects with second or greater relapse by up to three 5 -week treatment cycles of bortezomib . and refractory to their last prior therapy ) . In Study M34100 . Within each 3 -week treatment cycle , the patient received 025 , the CR + PR rate to bortezomib alone was 27 % (53 of 50 bortezomib 1 . 3 mg/ m2 /dose alone as a bolus intravenous 193 patients ) , and the overall response rate (CR + PR +MR ) to (IV ) injection twice weekly for two weeks ( on Days 1 , 4 , 8 , bortezomib alone was 35 % (67 of 193 patients ) . See Rich - and 11) of a 21 - day cycle . Within each 5 -week treatment ardson P G , et al. N Engl J Med ., 348 :2609 - 17 (2003 ). In cycle , the patient received bortezomib 1 . 3 mg/ m² / dose alone Study M34100 -024 CR + PR rates of were 30 % and 38 % as a bolus IV injection once weekly ( on Days 1 , 8 , 15 , and were seen among patients with relapsed multiple myeloma 55 22 ) of a 35 - day cycle . treated with bortezomib 1 .0 mg/ m² and 1 .3 mg/m² , respec Patients assigned to the high -dose dexamethasone group tively . See Jagannath , Br J Haematol. 127 : 165 -72 ( 2004 ) . received treatment for a maximum of 280 days. Patients in Patient samples and response criteria from patients partici- this treatment group received up to four 5 -week treatment pating in these studies , as well as the following additional cycles, followed by up to five 4 -week treatment cycles . studies described below were sought for use in pharmacog - 60 Within each 5 -week treatment cycle , the patient received enomic analyses to identify markers associated with patient dexamethasone 40 mg/ day PO , once daily on Days 1 to 4 , 9 response to treatments to 12 , and 17 to 20 of a 35 - day cycle . Within each 4 -week An Open - Label Study Comparison of Bortezomib Versus treatment cycle , the patient received dexamethasone 40 High Dose Dexamethasone in Patients with Relapsed and mg /day PO once daily on Days 1 to 4 of a 28 day cycle . The Refractory Myeloma 65 protocol provided for patients in the dexamethasone group A multicenter, open - label, randomized study was con - who experienced confirmed progressive disease (PD ) to ducted , comprising 627 enrolled patients with relapsed or receive bortezomib on a companion study ( An International, US 9 ,963 , 747 B2 49 50 Non - Comparative, Open -Label Study of PS -341 Adminis and according to the protocol would have received at least tered to Patients with Multiple Myeloma Who Received four prior therapies . Pharmacogenomic samples were also High -dose Dexamethasone or Experienced Progressive Dis sought for these 240 patients . ease after Receiving at Least Four Previous Therapies , During the study, disease response was assessed accord (Protocol M34101 - 040 ) . An additional 240 patients who did 5 ing to the European Group for Blood and Marrow Transplant not participate in this study, enrolled in the companion study ( EBMT) criteria as presented in Table C . TABLE C Disease Response Criteria Table C Disease Response Criteria Response Criteria for response Complete Requires all of the following: response (CR ) Disappearance of the original monoclonal protein from the blood and urine on at least two determinations for a minimum of six we KS DV immunofixation studies. < 5 % plasma cells in the bone marrow3. No increase in the size or number of lytic bone lesions ( development of a compression fracture does not exclude response ). Disappearance of soft tissue plasmacytomas for at least six weeks. Partial PR includes patients in whom some, but not all , criteria for CR are response fulfilled providing the remaining criteria satisfy the requirements for ( PR ) PR . Requires all of the following : 250 % reduction in the level of serum monoclonal protein for at least two determinations six weeks apart . Iloille If present, reduction in 24 - hour urinary light chain excretion by either > 90 % or to < 200 mg for at least two determinations six weeks apart. 250 % reduction in the size of soft tissue plasmacytomas (by clinical or radiographic examination ) for at least six weeks . No increase in size or number of lytic bone lesions ( development of compression fracture does not exclude response ) . Minimal MR includes patients in whom some, but not all, criteria for PR are response fulfilled providing the remaining criteria satisfy the requirements for (MR ) MR. Requires all of the following : 225 % to 550 % reduction in the level of serum monoclonal protein for at least two determinations six weeks apart . If present, a 50 to 89 % reduction in 24 - hour light chain excretion , which still exceeds 200 mg/ 24 h , for at least two determinations six weeks apart . 25 -49 % reduction in the size of plasmacytomas (by clinical or radiographic examination ( e . g ., 2D MRI, CT scan ) . No increase in size or number of lytic bone lesions (development of compression fracture does not exclude response ) . No change (NC ) Not meeting the criteria for MR or PD . Progressive Requires one or more of the following: disease (PD ) > 25 % increase in the level of serum monoclonal paraprotein , which ( for patients must also be an absolute increase of at least 5 g / L and confirmed on a not in CR ) repeat investigation one to three weeks later4, > 25 % increase in 24 -hour urinary light chain excretion , which must also be an absolute increase of at least 200 mg/ 24 h and confirmed on a repeat investigation one to three weeks latert, s. > 25 % increase in plasma cells in a bone marrow aspirate or on trephine biopsy , which must also be an absolute increase of at least 10 % . Definite increase in the size of existing lytic bone lesions or soft tissue plasmacytomas . Development of new bone lesions or soft tissue plasmacytomas (not including compression fracture) . Development of hypercalcemia (corrected serum calcium > 11 . 5 mg/ dL or 2 . 8 mmol/ L not attributable to any other cause ) 4. Relapse Requires at least one of the following : from CR Reappearance of serum or urine monoclonal paraprotein on immunofixation or routine electrophoresis to an absolute value of > 5 g / L for serum and > 200 mg/ 24 hours for urine, and excluding oligoclonal immune reconstitution . Reappearance of monoclonal paraprotein must be confirmed by at least one follow -up . 25 % plasma cells in the bone marrow aspirate or biopsy . Development of new lytic bone lesions or soft tissue plasmacytomas or definite increase in the size of residual bone lesions (not including compression fracture ) . US 9, 963, 747 B2 51 52 TABLE C - continued Disease Response Criteria Table C Disease Response Criterial Response Criteria for response Development of hypercalcemia ( corrected serum calcium > 11. 5 mg/ dL or 2 . 8 mmol / L not attributable to any other cause ) . Based on the EBMT criteria . See , Blade J , et al. Br J Haematol; 102 (5 ) : 1115- 23 ( 1998 ). For proper evaluation of CR , bone marrow should be 20 % cellular and serum calcium should be within normal limits . A bone marrow collection and evaluation is required to document CR . Repeat collection and evaluation of bone marrow is not required to confirm CR for patients with secretory myeloma who have a sustained absence of monoclonal protein on immunofixation for a minimum of 6 weeks ; however, repeat collection and evaluation of bonemarrow is required at the Response Confirmation visit for patients with non - secretory myeloma. 4 The need for urgent therapy may require repeating these tests earlier or eliminating a repeat examination . For determination of PD , increase in paraprotein is relative to the nadir .

Patients were evaluable for response if they had received from log -rank test adjusted by actual randomization factors . at least one dose of study drug and had measurable disease See , Richardson et al. , New Engl J Med ., submitted . at baseline (627 total patients : 315 in the bortezomib group Median time to response was 43 days for patients in both and 312 in the dexamethasone group ) . The evaluation of 20 groups . Median duration of response was 8 months in the confirmed response to treatment with bortezomib or dexam - bortezomib group and 5 .6 months in the dexamethasone ethasone according to the European Group for Blood and group . Marrow Transplant ( EBMT) criteria is provided in Table D . Patients given bortezomib had a superior overall survival. One -year survival was 80 % on bortezomib and 66 % on Response and date of disease progression was determined by125 dexamethasone ( P < 0 . 0030 ) . This represents a 41 % decrease computer algorithm that integrated data from a central 2 in risk of death in the bortezomib group during the first year laboratory and case report forms from each clinical site , after enrollment. The hazard ratio for overall survival was according to the Blade criteria ( Table C ) . The response rate 0 . 57 ( P < 0 .0013 ) , favoring bortezomib . The analysis of over ( complete plus partial response (CR + PR ) ) in the bortezomib all survival includes data from 147 patients ( 44 percent ) in group was 38 percent; and in the dexamethasone group was 30 the dexamethasone group who had disease progression and 18 percent ( P < 0 .0001 ) . Complete response was achieved in subsequently crossed over to receive bortezomib in a com 20 patients (6 percent ) who received bortezomib , and in 2 panion study . patients ( < 1 percent) who received dexamethasone Quality of Life assessment can be analyzed to determine (P < 0 .001 ) , with complete response plus near -complete if response to therapy was accompanied by measurable response in 13 and 2 percent ( P < 0 .0001 ) in patients receiv - improvement in quality of life . Analysis is performed on ing bortezomib and dexamethasone, respectively . These data » summary scores as well as individual items, with specific have been submitted for publication . See Richardson PG , et analytical methods outlined in a formal statistical analysis al. [ submitted NEJM ). plan developed prior to database lock . TABLE D Summary of Best Confirmed Response to Treatment1 , 2 ( Population , N = 627 ) bortezomib dexamethasone Best Confirmed | n ( % ) n ( % ) Difference Response ( n = 315 ) ( n = 312) (95 % CI) p - value Overall Response Rate 121 (38 ) 56 ( 18 ) 0 .20 (0 .14 , 0 . 27 ) < 0 .0001 (CR + PR ) Complete Response 20 ( 6 ) 2 ( 1 ) 0 . 06 ( 0 .03 , 0 .09 ) 0 . 0001 Partial Response 101 ( 32 ) 54 ( 17 ) 0 . 15 ( 0 .08 , 0 .21 ) < 0 .0001 Near CR : IF + 21 ( 7 ) 3 ( 1 ) 0 .06 ( 0 .03 , 0 . 09 ) SWOG Remission 46 ( 15 ) 17 ( 5 ) 0 . 09 ( 0 . 05 , 0 . 14 ) Minor Response 25 ( 8 ) 52 (17 ) - 0 .09 ( - 0 . 14 , - 0 .04 ) CR + PR + MR 146 (46 ) 108 ( 35 ) 0 . 12 ( 0 . 04 , 0 . 19 ) No Change 137 (43 ) 149 (48 ) - 0 .04 ( - 0 . 12 , 0 .04 ) Progressive Disease 22 ( 7 ) 41 ( 13 ) - 0 .06 ( - 0 . 11 , - 0 .01 ) Not Evaluable 10 (3 ) 14 ( 4 ) - 0 .01 ( - 0 .04 , 0 .02 ) Response based on computer algorithm using the protocol- specified EBMT criteria . Percents calculated for the statistical output in section 14 are 'rounded to the nearest integer including percents 20 . 5 % but < 1 % rounding to 1 % ; these are reported in the in -text tables as < 1 % . " Asymptotic confidence interval for the difference in response rates. " P - value from the Cochran -Mantel - Haenszel chi- square test adjusted for the actual randomization stratifi cation factors. Disease progression was determined by Blade criteria as 60 Pharmacogenomic Samples Collected described in Table C and above. The median time to disease Pharmacogenomic tumor samples (bone marrow aspirate ) were collected from patients for evaluation of the expression progression in the bortezomib group was 6 .2 month (189 of globalmRNA levels . days ) ; and the in the dexamethasone group was 3 . 5 months 65 Statistical Procedures ( 106 days ) ( hazard ratio 0 .55 , P < 0 .0001 ) . The date of Summary tabulations were presented that displayed the progression was determined by computer algorithm . P -value number of observations, mean , standard deviation ,median , US 9 , 963 , 747 B2 53 54 minimum , and maximum for continuous variables , and the non - responder analysis utilizes 79 vs. 84 samples . The number and percent per category for categorical data . The responder vs. progression analysis utilizes 79 vs. 41 and the categories for summarization were the two assigned treat- progression vs. other analysis utilizes 41 vs. 122 samples . ment groups. For the dexamethasone analysis NR 25, Ns = 16 , and Np= 21. A formal statistical analysis plan was developed and 5 Accordingly , the responder vs . non - responder analysis uti finalized prior to database lock . The primary efficacy analy - lizes 25 vs . 37 samples. The responder vs . progression ses were performed on the intent- to - treat ( ITT) population analysis utilizes 25 vs . 21 and the progression vs. other The primary efficacy analysis was performed on the rates of analysis utilizes 21 vs. 41 samples . responders , where a responder was defined as a CR , PR , or 44 ,928 gene transcripts (Affymetrix probe sets ) were MR using the criteria prospectively established in Table C . 10 profiled for each sample on the two Affymetrix U133 Two - sided 90 % confidence limits on proportions of respond - microarrays ( A and B ) according to manufacturer ' s direc ers in each dose group were established , corresponding to a tions. Total RNA was isolated from homogenized patient 95 % one - sided lower limit . tumor tissue by TriazolTM ( Life Technologies, Inc .) and For those patients who participated in the pharmacog - stored at 80° C ., following the manufacturer ' s recommen enomic portion of the study, correlation between RNA 15 dations. Detailed methods for labeling the samples and expression levels and response to therapy were evaluated subsequent hybridization to the arrays are available from descriptively . In addition , duration of response, time to Affymetrix (Santa Clara, Calif .) . Briefly , 1. 5 ug of total RNA disease progression , quality of life , and overall patient was converted to double -stranded cDNA (Superscript ; Life survival may be analyzed using RNA expression as a factor. Technologies, Inc .) priming the first - strand synthesis with a 20 T7 - (dT ) 24 primer containing a T7 polymerase promoter TABLE E (Affymetrix Inc . ) . All of the double - stranded DNA was subsequently used as a template to generate biotinylated Summary of Pharmacogenomic Patient Response CRNA using the incorporated T7 promoter sequence in an in vitro transcription system (Megascript kit ; Ambion and Study CR PR MR NCPD IE TOTALresponse with evaluable 25 Bio -11 -CTP and Bio -16 -UTP ; Enzo ). Reference oligonucle otides and spikes were added to 6 - 10 ug of cRNA , which all 10 69 25 59 61 22 224 was then hybridized to U133 A and B oligonucleotide arrays 024 1 1 0 1 4 0 025 2 10 3 10 14 5 for 16 h at 45° C . with constant rotation . The arrays were 040 1 20 6 13 8 ON2 then washed and stained on an Affymetrix fluidics station 039 341 5 25 5 19 13 9 8g 30 using the EUKGE -WS 1 protocol and scanned on an 039 Dex 1 13 11uawoü 16 22 6 Affymetrix GeneArray scanner. Normalization and Logarithmic Transformation . A total of 224 patient samples were assessed for pharma - Expression values for all markers on each microarray cogenomic analyses. These patient samples were collected were normalized to a trimmed mean of 150 . Expression from the clinical trials of bortezomib for the treatment of 35 values were determined using MASS gene expression analy multiple myeloma See Table E . The overall response rate to sis data processing software ( Affymetrix , Santa Clara , bortezomib in this set of patients was 46 . 4 % ( CR + PR rate of Calif . ) . These values will be referred to as the “ normalized 35 % ) . The overall response rate to dexamethasone was expression ” in the remainder of this section . In a further 39 .7 % (CR + PR rate of 22 . 2 % ) . All pharmacogenomic processing step , the number 1 was added to each normalized analyses relied on the European Group for Blood and 40 expression value . The logarithm base 2 was taken of the Marrow Transplant (EBMT ) criteria of response category . resulting number, and this value will be referred to as the Identification of Responsive and Non -Predictive Markers “ log expression ” in the remainder of this section . Biopsies from 224 multiple myeloma patients resulted in Variance Components Analysis . generation of high quality gene expression data which was There were up to six replicate hybridizations for each used to identify predictive markers . Candidate markers that 45 patient : three replicate hybridizations for each of two T7 are correlated with the outcome of multiple myeloma RNA labelings . To summarize replicates into a single esti patients to proteasome inhibition ( e . g . , bortezomib ) therapy mate of intensity for each patient, a mixed effects linear or glucocorticoid ( e . g . , dexamethasone ) therapy were model was used . For each probe set , a model was fit which selected by using a combination of marker ranking algo included terms the patient sample specific random effect rithms . Supervised learning and feature selection algorithms 50 representing the deviation from the overall mean intensity , were then used to identify the markers of the present and the replicate hybridization random effect. These random invention . effects are referred to as the variance components of the A data set comprising 224 discovery samples, time to model . Model fitting includes assessing the variance due to progression data and short- term response categorization was these two random effects , resulting in estimates of patient used to identify genes associated with patient outcome to 55 sample variance and replicate variance . one of two treatments ( bortezomib or dexamethasone ) . The S ummarizing Expression Across Replicates . data set consisted of discovery samples matched with the The final summary expression value, for each sample on patient' s outcome as measured by best response and time to each probe set , was obtained by estimating the best linear disease progression . For best response , each patient was unbiased predictor (BLUP ) . The BLUP can be viewed as a classified as responder (N2 ) , stable disease (NS ), or progres - 60 weighted average of each subject ' s replicates with weights sion (Np ) . For marker identification , the three response inversely proportional to the linear combination of the classes were further grouped into responders vs . non - re - variance components. The weights influence how much each sponders ( stable and progression ) (NP - s ) , responders vs. subject 's estimate of intensity deviates away from the over progression , or progression vs. others ( stable and respond all mean . Details on mixed effects models and calculating ers ) (NR - s ) . The analyses further separated the patients 65 BLUP estimates can be found in most texts which discuss based on the treatment they received . For bortezomib analy - linear mixed effects models and variance components . See , ses Nr = 79, N = 43 , and Np = 41. Thus, the responder vs . for example , " Variance Components ” by Searle , Casella , US 9 , 963, 747 B2 55 56 and McCulloch . Wiley Series in Probability and Mathemati arbitrarily designated “ control” and “ tester. ” PFC finds cal Statistics , 1992 John Wiley & Sons . genes with higher expression in the tester than in the control Removal of Genes with Low Inter- Patient Variance . samples . For the two -class comparisons described in this The probe sets were reduced in number to include only invention , each class was used in turn as the tester. To those having more than 75 % of their variance due to patient 5 qualify as having higher expression , tester samples must be sample variance . Of 44 ,928 probe sets , 7 , 017 passed this above the kth percentile of the control sample . The fold filter and were carried on to further analysis . change values of tester samples are subjected to a nonlinear Optional Reverse Log Transformation . transformation that rises to a user- specified asymptote , in The BLUP expression value was used for differential order to distinguish moderate levels of fold - change , but not expression analysis with the t- test . For computing the digital 10 make distinctions between very large fold -changes . The differential expression scores, the final summary value, x , squashed fold - change values of the over - expressed tester was transformed back to the original scale by exponentiat samples are averaged to get the POOF score . In particular , ing , thus reversing the log transformation : PFC for a given tester sample , s, and gene , g , is computed y = 2x - 1 15 as the average across tester samples of the compressed Single Marker Selection . tester :control ratio R ( s , g ) : R ( s , g )= C ( x , / ( k + x . 9 ) ) , where Single gene transcripts that appear associated with patient C (x ) is the compression function C ( z ) = A ( 1 - e - 2/ 4 ) for zzT , response categories or with patient time to progression can and C ( z ) = 0 for z < T , where T is a threshold value no less than be identified using the feature ranking and filtering meth - 1 . 0 . A is an upper asymptote on the fold -change value, k is odology described below . Single marker identification of 20 a constant reflecting the additive noise in the data , i .e ., the predictive markers using the methodology described herein fixed component of the variance in repeated measurements . are set forth in Table 1 ( Table 1A and Table 1B ), Table 2 Xps is the expression value of gene g in sample s, x , 2 is the ( Table 2A and Table 2B ) , and Table 3 . Qth percentile of the control samples' expression value . Model Selection . A minimum fraction f of the tester samples must have A set of one or more gene transcripts that together classify 25 R ( s , g ) greater than 0 ; if this does not hold true , then the value samples into sensitive and resistant groups ( or responsive of R ( s . g ) is set to 0 . and non - responsive ) or predict TTP , in the context if a We used the following parameters in our application of particular classifier algorithm , is referred to as a “ model. ” this algorithm : The gene transcripts are referred to as " features. ” Determin ing which combination of gene transcript( s ) best classifies 30 samples into sensitive and resistant groups is referred to as Parameter " model selection .” The following section describes the pro cess of how the models of the present invention were Q f T A k identified . An exemplary model is set forth in Table 4 . The methods provided herein along with the single marker 35 Value in run 1 0. 9 0. 3 1. 2 5 0 . 25 identification or predictive markers can be used to identify additional models comprising markers of the invention . Markers using the 7 ,017 probe sets were analyzed for Feature Ranking and Filtering differential expression across the 224 patient samples using The first step in predictive model selection is to filter the the t -test and PFC methods described above. Probe sets 7 ,017 features down to a smaller number which show a 40 found to be significant by t- test with a p -value less than 0 .01 , correspondence with the sample classifications. Filtering or having a PFC score other than 0 , are reported in Table 1A , involves first ranking the features by a scoring method , and Table 1B , Table 2A , Table 2B and Table 3 . These probe sets then taking only the highest ranking features for further can be used in building marker sets as exemplified below . analysis. The filtering algorithms used in the present inven - A Cox proportional hazard analysis was performed to tion were : ( 1 ) t -test , and ( 2 ) Pooled Fold Change (“ PFC ) . In 45 determine predictors of time until disease progression (TTP ) certain aspects, the t - test was used to identify genes showing in patients with relapsed and refractory multiple myeloma a small but consistent change in levels , and PFC was used after treatment. This methodology is designed to analyze to identify genes that were “ off ” in one class , but “ on ” in a time to event data where some of the data may be censored fraction of the other class . For time to progression data , Cox (see E . T . Lee, Statistical Methods for Survival Data Analy proportional hazards modeling was used to determine a 50 sis , 2nd ed . 1992 , John Wiley & Sons , Inc . ) . The median time p - value for the association of a feature with time to pro - to disease progression in the bortezomib group was 6 . 2 gression . month ( 189 days ) ; and the in the dexamethasone group was The t- test is a standard statistical method to test for 3. 5 months (106 days ) ( hazard ratio 0 .55 , P < 0 .0001 ) . The significant difference of means between two sets of points date of progression was determined by computer algorithm . presumed to have normal distribution . It is closely related to 55 The statistical package SAS was used to perform the analy the more ad hoc measure of differential expression SNR sis ; what parameters used to assess, etc . ( signal to noise ratio ) , which is the difference of the class We estimated Cox proportional hazard models for each of means divided by the sum of the class standard deviations , the 7017 transcripts passing the variance filter . That is , 7 ,017 and has been used to analyze expression data before ; for models were estimated where each model contained 1 example , see the definition of P ( g , c ) , a measure of correla - 60 transcript. From each model, we obtained estimates of tion between expression of gene g and class vector c , in relative risk , 95 % confidence intervals and p - values for the Golub et al ., “Molecular Classification of Cancer : Class association of each transcript to TTP . From the 7017 models , discovery and class prediction by marker expression moni we found 294 transcripts which had p -values of less than toring, ” Science , 286 : 531 - 537 ( 1999 ) , the contents of which 0 .01 in analyzing the 162 patients treated with bortezomib . are incorporated herein by reference . 65 We found 187 transcripts which had p - values of less than The Pooled Fold Change (“ PFC ” ) method is a measure of 0 .01 in analyzing the 63 patients treated with dexametha differential expression between two groups of samples , sone. That is , these transcripts were significantly associated US 9 , 963 ,747 B2 57 58 with TTP . These probe sets are listed in Table 1A , Table 1B , selection methods were utilized for determining the best Table 2A , Table 2B and Table 3 . classifier , and rank determination : ( 1 ) t -test , ( 2 ) Pooled The rank reported in Tables 1A , 1B , 2A , 2B and 3 is Fold Change (“ PFC ” ) , and ( 3 ) the Wilcoxon Rank -Sum determined by independently ranking the different scores of Test. the markers . Ranks are generated for TTP , for PD vs R , for 5 Predictive markers of the invention are provided in Tables PD vs NC + R , for NR vs R . For the response comparisons, 1A , 1B , 2A , 2B , and 3 . Table 1 sets forth predictivemarkers both T - test and digital scores are ranked . Thus there can be identified which are specific identifiers of response or non up to 7 different # 1 ranks for proteasome inhibitor -specific response to proteasome inhibition therapy ( e . g . , bort predictive markers of treatment outcome. ezomib ) . Table 1A provides predictive markers which are Summary of the Data Provided in the Tables 10 upregulated indicators of non - response and / or correlate with The following terms are used throughout the Tables : shorter time to progression . Marker nos. 1 - 547 in Table 1A “ No .” or “ Number ” corresponds to an identification number are newly associated predictive markers , and predictive for the predictive markers. markers no . 548 - 657 have been previously identified as “ Probeset ID ” corresponds to the Affymetrix ( Santa Clara , associated markers predictive of non - response and / or cor Calif. ) identifier from the Human Genome U133A , B set 15 relation with shorter time to progression . See , International oligonucleotide arrays which were used ; Patent Publication No. W004053066 , published Jun . 24 , “ Rep Public ID ” refers to a Representative Public identifier 2004 . Table 1B provides predictive markers which are for the gene corresponding to the probe set , and was taken upregulated indicators of response and / or correlate with from HG -U133A and HG -U133B annotation files , dated longer time to progression . Marker nos. 658 - 876 in Table 1B Apr. 12 , 2005 which was available and downloaded from 20 are newly associated predictive markers , and predictive the GeneChip support area of the Affymetrix web site markers no. 877 - 911 have been previously identified as ( accessible in the Human Genome U133 Set- Support associated markers predictive of response and /or correlation Materials in the Support section of the website maintained with longer time to progression . See , International Patent by Affymetrix , Inc ., Santa Clara , Calif . ) ; Publication No . WO04053066 , published Jun . 24 , 2004 . " SEQ ID NO ” is the identification number in the sequence 25 Table 2 sets forth predictive markers identified which are listing of the sequence corresponding to the sequence in specific identifiers of response or non - response to glucocor the GenBank record identified by the Representative ticoid therapy ( e . g ., dexamethasone ) . Table 2A provides Public Identifier. predictive markers which are upregulated indicators of non “ Title” corresponds to a common description , where avail Presponse " and / or correlate with shorter time to progression . able , and was also taken from the Affymetrix annotation 30 Marker nos . 912 - 1062 in Table 2A are newly associated files ; predictive markers , and predictive markers no . 1063 - 1070 “ Gene symbol” corresponds to a symbol the gene is com - have been previously identified as associated markers pre monly known by , and was also taken from the Affymetrix dictive of non - response and / or correlation with shorter time annotation files; to progression related to advanced stage patient' s non “ Entrez Gene ID ” corresponds to the NCBI Unigene unique 35 response to bortezomib treatment. See, International Patent gene identifier; Publication No . W004053066 , published Jun . 24 , 2004 . “ TTP Marker” represents indication of predictive marker Table 2B provides predictive markers which are upregulated which is significantly upregulated in samples with a indicators of response and /or correlate with longer time to correlation to longer time to progression ( + ) , or are progression . Marker nos . 1071 - 1185 in Table 2B are newly significantly upregulated in samples with a correlation to 40 associated predictive markers , and predictive markers no . shorter time to progression (- ) ; 1186 - 1202 have been previously identified as associated “ Response Marker ” represents indication of predictive markers predictive of response and / or correlation with lon marker which is significantly upregulated in samples ger time to progression related to advanced stage patient ' s which are responsive to therapy ( + ) , or are significantly response to bortezomib treatment. See, International Patent upregulated in samples which are non - responsive to 45 Publication No . W004053066 , published Jun . 24 , 2004 . therapy ( - ) ; Table 3 sets forth predictive markers identified which are not “ Type of Specificity ” indicates the significance of TTP specific to proteasome inhibition therapy or glucocorticoid and /or response indicator as significant indicator of the therapy , rather are indicator predictive markers of responsel predictive marker ; longer time to progression ( + ) or non - response / shorter time “ Rank ” corresponds to the process of determining which 50 to progression ( - ) with regard to either therapy, and are individual markers may be used in combination to group indicators of general disease aggressiveness . Marker nos . or classify a sample , for example , as responsive or non - 1203 - 1423 in Table 3 are newly associated predictive mark responsive . Rank is indicated as the lowest rank score ers , and predictive markers no . 1424 - 1474 have been pre identified among all the methods for each of the predictive viously identified as associated markers predictive of non markers . In Table 3 where predictive markers are indica - 55 response / correlation with shorter time to progression and / or tive of responsive or non -responsive for proteasome inhi- response / correlation with longer time to progression related bition or glucocorticoid therapy, the rank indicates the to advanced stage patient' s response to bortezomib treat lowest rank across various methods for bortezomib or ment. See , International Patent Publication No . dexamethasone treated samples. Three different feature W004053066 , published Jun . 24 , 2004 . US 9, 963, 747 B2 5959 09

OvW Dauw ?? ?? ?? ?? ?? tinin ON 4 Rank 2222 30 35 36 48

resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp

TTPmarkerResponseTypeofspecificity | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Ill

Entrez GeneID 6146 6201 4673 6233 10161 27230 6235 9167 55342 689 689 8667 6207 84798 9045 6723 6165 23204/ 6210 91689 4736 1350 962 6146 6230 6136 6146 6135 1350 91689 4736 6128 1975 127253 128338 3646 8664 8721 1350

Gene Symbol RPL22 RPS7 NAP1L1 RPS27A P2RY5 SERP1 RPS29 COXTA2L STRBP BTF3 BTF3 EIF3S3 RPS13 MGC13170 RPL14 SRM RPL35A RPS15A IIIARLÓIP LOC91689 RPL10A COX7C CD48 RPL22 RPS25 RPL12 RPL22 RPL11 COX7c LOC91689 RPL10A RPL6 EIF4B LOC127253 MGC54289 EIF3S6 EIF3S7 EDF1 COX7C TABLE1 PROTEASOMEINHIBITORPREDICTIVEMARKERIDENTIFICATION 1APredictiveMarkersUpregulatedIndicatorsofNon-Responseand/orShortTimetoProgression

coupledprotein,5receptor-purinergicP2YG 3nucleosomeassemblyprotein1-like 12eukaryotictranslationinitiationfactor3, 18ribosomalproteinS15a/ADP-ribosylation 33eukaryotictranslationinitiationfactor3, 34eukaryotictranslationinitiationfactor3, 9spermatidperinuclearRNAbindingprotein 19hypotheticalgenesupportedbyAL449243 22CD48antigen(B-cellmembraneprotein) 30eukaryotictranslationinitiationfactor4B 35endothelialdifferentiation-relatedfactor1 8cytochromecoxidasesubunitVIIa subunit3gamma,40kDa 14multidrugresistance-relatedprotein factor-likeinteracting6protein 21cytochromecoxidasesubunitVIIC 28cytochromecoxidasesubunitVIIC 19hypotheticalgenesupportedbyAL449243 subunit7zeta,66/67kDa 36cytochromecoxidasesubunitVIIC 6stress-associatedendoplasmicreticulum likepolypeptide2 10basictranscriptionfactor3 11basictranscriptionfactor3 31hypotheticalproteinBC009514 32hypotheticalproteinMGC54289 subunit648kDa 1ribosomalproteinL22 2ribosomalproteinS7 4ribosomalproteinS27a protein1 7ribosomalproteinS29 13ribosomalproteinS13 15ribosomalproteinL1416spermidinesynthase 17ribosomalproteinL35a 20ribosomalproteinL10a 23ribosomalproteinL22 24ribosomalproteinS25 25ribosomalproteinL12 26ribosomalproteinL22 27ribosomalproteinL11 20ribosomalproteinL10a 29ribosomalproteinL6 NO:Title SEQ ID

RepPublic NM_001207 NM_003756 NM_003753 ID AK024960 X74070 HG-U133ANM_001032 HG-U1334NM_001017 HG-U133ANM_001019 HG-U133ANM_001867 HG-U133ANM_004718 HG-U133BBC006151 HG-U133BU16738 HG-U133ANM_003132 NM_0029544200017atHG-U133A HG-U133ABE968801 HG-U133A HG-U133AAB002282 chip 1220960_xatHG-U133ANM000983 5218589_atHG-U133ANM005767 6200971SatHG-U133ANM014445 NM20200036_satHG-U133B007104 22204118_atHG-U133ANM001778 27200010_atHG-U133ANM000975 30200036_satHG-U133ANM007104 31200034_satHG-U133BNM000970 2213941xatHG-U133A1970731 3208752_xatHG-U133AA1888672 19225794_satHG-U133BAV751709 21217491XatHG-U133AAF042165 23221775_xatHG-U133ABG152979 25200088_xatHG-U133BAK026491 26208768_xatHG-U133AD17652 28213846_atHG-U133AAA382702 29225795_atHG-U133BAV751709 32211938atHG-U133ABF247371 34225230_atHG-U133BA1735261 35208697_satHG-U133ABC000734 9233252_satHG-U133B10208517_xatHG-U133A 11211939_xatHG-U133A 12201592_atHG-U133A 24200091_satHG-U133A 33227141atHG-U133BAW205739 201094_at 201256_at 14224468_sat 15200074_sat 17213687_sat 18200781_sat 200005_at 38201134_xat No.ProbeSetID 13200018_at 16201516_at 37209058_at AWN au 7 8 36 US 9, 963, 747 B2 62

aaa Rank 49 50 55 56au NNNNN 80 91 93 94 97 99 100 101 102

resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp TTPmarkerResponseTypeofspecificity

Entrez GeneID 440309 6138 nie6143 23394 6202 8664 it2778 23480 9045 51611 3338 4666 1933 6135 6152 6189 6155 6230 10480 28973 9782 27316 6194 6217 6189 11224 6156 6152 6207 27044 6143 83939 9782 7385 6128 6129 5935 64960 55342

Gene Symbol RPL15 RPL19 ADNP RPS8 EIF3S7 GNAS SEC616 RPL14 CGI-30 DNAJC4 NACA EEF1B2 RPL11 RPL24 RPS3A RPL27 RPS25 GA17 MRPS18B MATR3 RBMX RPS6 RPS16 RPS3A RPL35 RPL30 RPL24 RPS13 SND1 RPL19 eIF2A MATR3 UQCRC2 RPL6 RPL7 RBM3 MRPS15 STRBP 1APredictiveMarkersUpregulatedIndicatorsofNon-Responseand/orShortTimetoProgression TABLE1-continued PROTEASOMEINHIBITORPREDICTIVEMARKERIDENTIFICATION

subunit7zeta,66/67kDa 15ribosomalproteinL14 member4 alphapolypeptide 59ribosomalproteinL30 proteinII 2A NO:Title 46DnaJ(Hsp40)homolog,subfamilyC50228622SatHG-U133BAW071239 SEQID initiation(eIF)translationfactoreukaryotic6171223015_atHG-U133BAF212241 55RNAbindingmotifprotein,X-linked61213762_xatHGU133AA1452524 staphylococcalnucleasedomaincontaining1HG-U133ANM_0143906069201622at 76208319_satHG-U133ANM00674365RNAbindingmotif(RNP1,RRM)protein3 45200005_atHG-U133BNM00375334eukaryotictranslationinitiationfactor3, 52200705_satHG-U133ANM00195948eukaryotictranslationelongationfactor1beta2 63ubiquinol-cytochromecreductasecore73212600_satHGU133AAV727381 66mitochondrialribosomalproteinS1577226296SatHG-U133BAKO21626 78223245_atHG-U133BAK02428567spermatidperinuclearRNAbindingprotein PublicRep 43201773_atHG-U133ANM01533941activitydependentneuroprotector 51208635_xatHG-U133ABF97626047nascentpolypeptideassociatedcomplex 59217408_atHG-U133AAL05036153mitochondrialribosomalproteinS18B HG-U133BNM_00101713ribosomalproteinS13200018at ID 39ribosomalproteinL1541221475_satHG-U133ANM002948 42200029_atHG-U133BNM00098140ribosomalproteinL19 47203484_atHG-U133ANM01430244Sec61gammasubunit HG-U133BNM00097527ribosomalproteinL1153200010_at 54200013_atHG-U133ANM00098649ribosomalproteinL24 58202231_atHG-U133ANM00636052dendriticcellprotein 62200081satHG-U133BBE74175456ribosomalproteinS6 67200013_atHG-U133BNM00098649ribosomalproteinL24 HG-U133ANM_00098140ribosomalproteinL1970200029at 44200858_satHG-U133ANM00101242ribosomalproteinS8 46200981_xatHG-U133ANM01659243GNAScomplexlocus 49224196_xatHG-U133BAF16149245CGI30protein 55200099_satHG-U133BAL35611550ribosomalproteinS3A 56200025_satHG-U133BNM00098851ribosomalproteinL27 57200091_satHG-U133BAA88838824ribosomalproteinS25 63213890_xatHG-U133AA120058957ribosomalproteinS16 50ribosomalproteinS3A64200099_satHG-U133AAL356115 65200002_atHG-U133BNM00720958ribosomalproteinL35 74200034_satHG-U133ANM00097029ribosomalproteinL6 64ribosomalproteinL775212042_xatHG-U133ABG389744 60200626_satHG-U133ANM01883454matrin3 62matrin372200624_satHG-U133AAA577695 chip 39225951_satHG-U133BAV75602637LOC440309 40216342_xatHG-U133AAL12191638 48200074SatHG-U133AU16738 200062_SatHG-U133AL05095

No.ProbeSetID 682000 NNNNN US 9, 963, 747 B2 63 64

Rank 103 109 109 111 E 113 114 115 117 119 123 125 126 127 127 128 128 130??131 ??131 ?? ?? 133?? 134 135 136 136 137 139 140 140 141 144

resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp

TTPmarkerResponseTypeofspecificity | | | | | | | | | | | | | | | | | | | | | | | | | | | . . | |

GeneID 293 4711 689 1968 3178 7388 1739 4673 746 Entrez 4666 11331 10954 6160 6181 51637 6132 6159 7178 6767 51253 2778 6156 2778 6881 56947 55830 28972 10399 6745 201725 /54543 5018

Gene Symbol NACA REA PDIR SLC25A6 NDUFB5 RPL31 RPLP2 C14orf166 RPL8 BTF3 EIF2S3 RPL29 TPT1 ST13 MRPL37 HNRPA1 UQCRH GNAS DLG1 NAP1L1 Cilorf10 RPL30 GNAS TAF10 C2orf33 GLT8D1 SPCS1 GNB2L1 SSR1 TOMM7 LOC201725 OXA1L 1APredictiveMarkersUpregulatedIndicatorsofNon-Responseand/orShortTimetoProgression TABLE1-continued PROTEASOMEINHIBITORPREDICTIVEMARKERIDENTIFICATION

carrier;adeninenucleotidetranslocator), bindingprotein(TBP)-associatedfactor, carcinoma)(Hsp70interactingprotein protein),betapolypeptide2-like1 7homolog(yeast)/hypotheticalprotein subcomplex,516kDa subunit3gamma,52kDa associatedproteinalpha) NM_01603975chromosome14openreadingframe166 77basictranscriptionfactor3 59ribosomalproteinL30 alphapolypeptide member6 (S.cerevisiae) protein 30kDa LOC201725 NO:Title ID NM_00097376ribosomalproteinL8 86discs,largehomolog1(Drosophila)97202515atHG-U133ABG251175 SEQ 110208717_atHG-U133ABC00166998oxidase(cytochromec)assembly1like 83203621_atHG-U133ANM00249272NADHdehydrogenase(ubiquinone)1beta 89224935_atHG-U133BBG16581578Eukaryotictranslationinitiationfactor2, 91216520_satHG-U133AAF07209880tumorprotein,translationallycontrolled1 98212967_xatHG-U133AAW14880187nucleosomeassemblyprotein1like 103200055_atHG-U133BNM00628491TAF10RNApolymeraseII,TATAbox 92chromosome2openreadingframe33104222832_satHG-U133BAA746206 106217927_atHG-U133ANM01404194signalpeptidasecomplexsubunit1homolog 107200651_atHG-U133ANM00609895guaninenucleotidebindingprotein( 108201894_satHG-U1334NM00192096signalsequencereceptor,alpha(translocon PublicRep 79200735_xatHG-U133ANM00559468nascentpolypeptideassociatedcomplex 80201600_atHG-U133ANM00727369repressorofestrogenreceptoractivity 81203857_satHG-U133ANM00681070forproteindisulfideisomeraserelated 82212826_satHG-U133AA196122471solutecarrierfamily25(mitochondrial 92207040_satHG-U133ANM00393281suppressionoftumorigenicity13(colon 93222993_atHG-U133BAF32570782mitochondrialribosomalproteinL37 95202233_satHG-U133ANM00600484ubiquinolcytochromecreductasehinge 99218213_satHG-U133ANM01420688chromosome11openreadingframe10 105218147_satHG-U133ANM01844693glycosyltransferase8domaincontaining1 ID 85200909_satHG-U1334NM00100474ribosomalprotein,largeP2 94200016_xatHG-U133BNM00213683heterogeneousnuclearribonucleoproteinA1 109201812_satHG-U133ANM01905997translocaseofoutermitochondrialmembrane 84200963_xatHG-U133ANM00099373ribosomalproteinL31 90200823_xatHG-U133ANM00099279ribosomalproteinL29 96217673_xatHG-U133AAA65055885GNAScomplexlocus 90GNAScomplexlocus102212273_xatHG-U133AA1591100 chip 89100228095atHG-U133BAA608749 88214800XatHG-U133AR83000 101200062_satHG-U133BL05095

No.ProbeSetID 86217768at 87200936at US 9, 963, 747 B2 65 99

Rank 145 147 147 149 149 150151 152 153 154 156 157 157 161 162 163 164 166 167 168 169 169 171 172 173 174 174 175 178 178 180 181 182

resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp TTPmarkerResponseTypeofspecificity

Entrez GeneID 80097 1936 513 5793 81502 51611 6146 1938 6233 6144 3178 84319 1938 6155 83443 328 29091 4200 57222 55342 9616 6122 4718 114049 3094 2879 55854 10128 8725 8666 293

Gene Symbol FLJ14346 EEF1D ATP5D PTPRG HM13 CGI-30 RPL22 EEF2 RPS27A RPL21 HNRPA1 MGC4308 EEF2 RPL27 SF3B5 APEX1 STXBP6 ME2 KIAA1181 STRBP RNF7 RPL3 NDUFC2 WBSCR22 HINT1 GPX4 LEREPO4 LRPPRC C19orf2 EIF354 SLC25A6 1APredictiveMarkersUpregulatedIndicatorsofNon-Responseand/orShortTimetoProgression TABLE1-continued PROTEASOMEINHIBITORPREDICTIVEMARKERIDENTIFICATION

delta(guaninenucleotideexchangeprotein) mitochondrialF1complex,deltasubunit102proteintyrosinephosphatase,receptor typeG 119NADHdehydrogenase(ubiquinone)1, subcomplexunknown,214.5kDa 126eukaryotictranslationinitiationfactor3, carrier;adeninenucleotidetranslocator), 100eukaryotictranslationelongationfactor1 103histocompatibility(minor)13 107eukaryotictranslationelongationfactor2 83heterogeneousnuclearribonucleoproteinA1 110eukaryotictranslationelongationfactor2 111splicingfactor3b,subunit510kDa 112APEXnuclease(multifunctionalDNArepair 113syntaxinbindingprotein6(amisyn) 114malicenzyme2,NAD(+)-dependent 115endoplasmicreticulum-golgiintermediatecompartment32kDaprotein 116spermatidperinuclearRNAbindingprotein 121histidinetriadnucleotidebindingprotein1 122glutathioneperoxidase4(phospholipid 123likelyorthologofmouseimmediateearly 125chromosome19openreadingframe2 101ATPsynthase,H+transporting 104U87HGmRNA,completesequence 120WilliamsBeurensyndromechromosome response,erythropoietin4 124leucine-richPPRmotifcontaining subunit4delta,44kDa 128solutecarrierfamily25(mitochondrial 99hypotheticalproteinFLJ14346 106ribosomalproteinL22 ribosomalproteinS27a4 108ribosomalproteinL21 109hypotheticalproteinMGC4308 1ribosomalproteinL27 enzyme)1 117ringfingerprotein7 118ribosomalproteinL3 hydroperoxidase) 105CGI-30protein region22 member6 NO:Title mitochondrial SEQID 127

PublicRep NM_001960 NM_002841 002954NM_ NM_000982 NM_002136 NM_001961 NM_000988 NM_031287 NM_014178 NM_004549 NM_017528 NM_002085 ID BG255188 BE798517 AL110115 BG169443 AF248965 AW071997 A1004246 BC006475 M80261 BC000147 AF267855 BC002693 BC005966 L22453 N32864 AV716798 A1653608 AW514900 BC000733 AL096829 AA916851

chip HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A U133BHG- HG-U133A HG-U133B HG-U133A 114204944_atHG-U1334 115224615_xatHG-U133B 116225547_atHG-U133B 117223671XatHG-U133B 118214042SatHG-U133A 119200094SatHG-U133B 120200017atHG-U133B 121200012_xatHG-U133B 122200016_xatHG-U133A 123224523_satHG-U133B 124204102_satHG-U133A 125200025_satHG-U133A 126221263_satHG-U133A 127210027_satHG-U133A 131223246_satHG-U133B 132224439_xatHG-U133B 133211666XatHG-U133A 134218101_satHG-U133A 138201593_satHG-U133A 139211971_satHG-U133A 140214173_xatHG-U133A HG-U133A141208887at 142216570_xatHG-U133A 143212085atHG-U133A 111212995_xat 112203113_sat 113213041_sat 128220994_sat 223847at_s130 135207628_sat 136200093_sat No.ProbeSetID 129209397_at 137201106_at US 9, 963, 747 B2 67 89

VN ?? ?? ?? Rank 183 184185 186 186 188 190 191 192 193 194 196 200 202 205 207 207 208 210 213 213 214 215 218 219 221 223 228 230 231 232 NNN

resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp TTPmarkerResponseTypeofspecificity

GeneID 1938 7417 8535 8721 8409 5479 4673 6166 6136 6125 6881 6138 3184 1452 1452 6217 6139 Entrez i 55501 196403 55830 28960 10944 55144 29789 28974 10632 145853 400055 441073 /6154 400916 51447 11165 10899 11315

RPL26/ Gene Symbol CHST12 DTX3 GLT8D1 EEF2 VDAC2 DCPS CBX4 EDF1 SMAP UXT LRRC5 PPIB PTD004 NAPILI HSPC023 ATP5L LOC145853 RPL36AL RPL12 RPL5 LOC400055 LOC441073 C22orf16 TAF10 RPL15 IHPK2 NUDT3 JTB HNRPD CSNK1A1 PARK7 CSNK1A1 RPS16 RPL17 1APredictiveMarkersUpregulatedIndicatorsofNon-Responseand/orShortTimetoProgression TABLE1-continued PROTEASOMEINHIBITORPREDICTIVEMARKERIDENTIFICATION

mitochondrialF0complex,subunitg bindingprotein(TBP)-associatedfactor, 148CDNAcloneIMAGE:4184613,partialcds (AU-richelementRNAbindingprotein1, 155Parkinsondisease(autosomalrecessive,early A1985751139nucleosomeassemblyprotein1-like 147chromosome22openreadingframe16 91TAF10RNApolymeraseII,TATAbox 150inositolhexaphosphatekinase2 151Nudix(nucleosidediphosphatelinkedmoiety 153heterogeneousnuclearribonucleoproteinD Drosophila) ribosomalproteinL26 149ribosomalproteinL15 152jumpingtranslocationbreakpoint 154caseinkinase1,alpha 156caseinkinase1,alpha 157ribosomalproteinS16 158ribosomalproteinL17 X)-typemotif3 37kDa) onset)7 12 30kDa NO:Title NM_004182135ubiquitously-expressedtranscript SEQ ID 150227558_atHG-U133BA1570531133chromoboxhomolog4(Pcclass, 131voltage-dependentanionchannel2148211662SatHGU133AL08666 155200967_atHG-U133ANM000942137peptidylprolylisomeraseB(cyclophilin) 164222229_xatHG-U133AAL121871146ribosomalproteinL26/similarto60S 130deltex3homolog(Drosophila)145235721_atHG-U133BN62126 Public 144218927_satHG-U133ANM018641129carbohydrate(chondroitin4)sulfotransferase 146218146_atHG-U133ANM01844693glycosyltransferase8domaincontaining1 151209059_satHG-U133AAB00228235endothelialdifferentiationrelatedfactor1 154218684_atHG-U133ANM018103136leucinerichrepeatcontaining5 159208746_xatHG-U133AAF070655141ATPsynthase,H+transporting 147200094_satHG-U133AA1004246107eukaryotictranslationelongationfactor2 149218774_atHG-U133ANM014026132decappingenzyme,scavenger NM_013341138GTP-bindingproteinPTD004156219293satHGU133A NM_006284 NM_006694 NM_007262 NM_000985 ID 161207585_satHG-U133ANM001001143ribosomalproteinL36alike A1814909 D55674 AW268585 BF341845 AA583817 152201784_satHG-U133ANM014267134smallacidicprotein 158217926_atHG-U133ANM014047140HSPC023protein 160229742_atHG-U133BAA420989142hypotheticalLOC145853 162214271_xatHG-U133AAA281332144ribosomalproteinL12 163213080_xatHG-U133ABF214492145ribosomalproteinL5 chip HG-U133B HG-U133A HG-U133A HG-U133B HG-U133A 167225220_atHG-U133BBF340290 168221476SatHG-U133AAF279903 157213864SatHG-U133AA1985751 169223165SatHG-U133BBC004469 170229803_SatHG-U133BA1347000 171200048_satHG-U133B 172209330_satHG-U133A 173213860_xatHG-U133A174200006 _atHG-U133A 175213086_sat 176226131_sat 177200038_sat No.ProbeSetID 153218495at 165224932_at 166200055at US 9, 963, 747 B2 69 70

Rank 238 239 240 244 247 248 250 252 254 254 256 257 258 259 260 263 265 269 271 275 275 276 = = = 280 282 283

resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp TTPmarkerResponseTypeofspecificity | | | | | | | | | | | | | | | | II

GeneID 5093 1153 52 9551 3094 2098 6223 7323 6139 6747 3178 6747 4701 3178/ 5501 9049 Entrez 23741 10899 388524 /3921 84146 387522 /7335 10548 84993 23521 27335 10155 57222 441507 121355

Gene Symbol PCBP1 CIRBP CRI1 ACP1 ATP5J2 JTB HINT1 ESD RPS19 RPSAIII LOC388524 ZNF644 UBE2D3 RPL17 SSR3 UBE2V1 /Kua UEV TMOSF1 HNRPA1 BMSC UbP SSR3 RPL13A EIF3S12 TRIM28 KIAA1181 NDUFAT HNRPA1 LOC441507 PPP1cc AIP FLJ32942 1APredictiveMarkersUpregulatedIndicatorsofNon-Responseand/orShortTimetoProgression TABLE1-continued PROTEASOMEINHIBITORPREDICTIVEMARKERIDENTIFICATION mitochondrialF0complex,subunitfisoform2 176eukaryotictranslationinitiationfactor3, 180NADHdehydrogenase(ubiquinone)1alpha ribonucleoproteinA1(Helix-destabilizing 121histidinetriadnucleotidebindingprotein1 166ribosomalproteinSA/similartoLaminin 169signalsequencereceptor,gamma(translocon 170ubiquitin-conjugatingenzymeE2variant1 174signalsequencereceptor,gamma(translocon protein)(Single-strandbinding (hnRNPcoreproteinAl)HDP 182proteinphosphatase1,catalyticsubunit 168ubiquitin-conjugatingenzymeE2D3 associatedproteingamma) 171transmembrane9superfamilymember1 172heterogeneousnuclearribonucleoproteinAl 173bonemarrowstromalcell-derivedubiquitin associatedproteingamma) 179endoplasmicreticulum-golgiintermediate compartment32kDaprotein subcomplex,714.5kDa181heterogeneousnuclearribonucleoproteinA1 183arylhydrocarbonreceptorinteractingprotein 160coldinducibleRNAbindingprotein 163ATPsynthase,H+transporting (UBC4/5homolog,yeast) 159poly(IC)bindingprotein1 161CREBBP/EP300inhibitor1 164esteraseD/formylglutathionehydrolase 177tripartitemotif-containing28 /similartoHeterogeneousnuclear 162acidphosphatase,soluble 152jumpingtranslocationbreakpoint 165ribosomalproteinS19 receptor1 167zincfingerprotein644 158ribosomalproteinL17 175ribosomalproteinL13a subunit12 gammaisoform 184hypotheticalproteinFLJ32942 like NO:Title SEQID 178

PublicRep NM_001280 NM_004889 NM_006694 NM_001022 NM_003340 NM_000985 NM_007107 NM_005762 NM_005001 NM_002710 ID U24223 AF109873 BG035989 N32864 BC001169 AW304232 AK023596 AW150923 BG164064 U94831 AL050348 BF568780 BF437161 AB033007 AL568186 AL558532 BG150433

hip HG-U133B HG-U133A HG-U133B HG-U133B HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133B HG-U133B HG-U133A HG-U133A HG-U1334 HG-U133A HG-U133B 198212716_satHG-U133AAW083133 178208620_atHG-U133A 179200811_atHG-U133A 180208669_satHG-U133A 181215227_xatHG-U133A 182202961_satHG-U133A 183200048_satHG-U133A 184200093satHG-U133A 185209009_atHG-U133A 186202649_xatHG-U133A 187213801_xatHG-U133A 197211942_xatHG-U133ABF979419 189200669_sat 190200038_sat 191222412_sat 192201001_sat 193209150_sat 194216559_xat 196217790_sat 203213356_xat 205201781_sat No.ProbeSetID 188222580_at 195225063_at 199200990_at 200239082at 201224577at 202202785_at 204200726_at 206227711at US 9, 963, 747 B2

Rank 284 285 288 292 293 294 297 298 299 300 301 303 304 307 308 309 311 311 312 313 313 314 315 317 318 318323 323 324 327 328 330 331??332 ?? 332 ?? 333 ?? 335 ??

resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp

TTPmarkerResponseTypeofspecificity | | | | | | | | | | | | | | | | | | | | | | | |

GeneID 6133 2653 6141 6136 6194 4697 6218 6161 4736 4841 6232 7982 6745 5479 6223 6150 Entrez 387522 /7335 84958 113263 11224 26355 113263 10473 26355 192286 27258 10972 51150 115416 219854 22890 84273 10018 150678 171546 64776 25873

Gene Symbol UBE2V1 /Kua UEV RPLE SYTL1 GCSH GLCCI1 RPL35 E2IG5 RPL18 RPL12 GLCCI1 HMGN4 RPS6 E2IG5 MGC2198 NDUFA4 RPS17 RPL32 LSM3 RPL10A NONO TMP21 Cab45 C7orf30 RPS27 LOC219854 ST7 ZBTB1 SSR1 C4orf14 BCL2L11 PPIB RPS19 LOC150678 C14orf147 Cilorf1 RPL36 MRPL23 1APredictiveMarkersUpregulatedIndicatorsofNon-Responseand/orShortTimetoProgression TABLE1-continued PROTEASOMEINHIBITORPREDICTIVEMARKERIDENTIFICATION

185ubiquitin-conjugatingenzymeE2variant1 associatedproteinalpha) (aminomethylcarrier) subcomplex,49kDa associated(S.cerevisiae) domain4 binding NO:Title SEQID 219224345XatHG-U133BAF107495194growthandtransformationdependentprotein 234213376_atHG-U133AA1656706209zincfingerandBTBdomaincontaining1 236223157_atHG-U133BBC004894211chromosome4openreadingframe14A1949179212 BCL2-like11apoptosis(facilitator237225606_atHGU133B) NM_000942137peptidylprolylisomeraseB(cyclophilin) 238200968satHG-U133A BE738425215chromosome14openreadingframe147241212460_atHG-U133A 190growthandtransformation-dependentprotein213223193_xatHGU133BAF201944 216225700atHG-U133BAC006042192glucocorticoidinducedtranscript1 221217773_satHG-U133ANM002489196NADHdehydrogenase(ubiquinone)1alpha 224202209_atHG-U133ANM014463199LSM3homolog,U6smallnuclearRNA 229226386_atHG-U133BBG397444204chromosome7openreadingframe30 230200741_satHG-U133ANM001030205ribosomalproteinS27(metallopanstimulin1) 235200891_satHG-U133ANM003144210signalsequencereceptor,alpha(translocon 242231530_satHG-U133BBG150085216chromosome11openreadingframe1 PublicRep 210213129_satHG-U133AA1970157188glycinecleavagesystemproteinH 211225706_atHG-U133BA1761989189glucocorticoidinducedtranscript1 L12proteinribosomal25215200088XatHG-U133AAK026491 193highmobilitygroupnucleosomalbinding217209787_satHG-U133ABC001282 226200057_satHG-U133BNM007363201nonPOUdomaincontaining,octamer 228221972_satHG-U133AAL571362203calciumbindingproteinCab45precursor ID U39361 209227134_atHG-U133BA1341537187synaptotagminlike1 233207871_satHG-U133ANM018412208suppressionoftumorigenicity7 240226845_satHG-U133BAL036350214helicase/primasecomplexprotein 244213897_satHG-U133AAI832239218mitochondrialribosomalproteinL23 208200032_satHG-U133BNM000661186ribosomalproteinL9 212200002_atHG-U133ANM00720958ribosomalproteinL35 214200022_atHG-U133BNM000979191ribosomalproteinL18 56ribosomalproteinS6218200081_satHG-U133ABE741754 220209329_xatHG-U133ABC000587195hypotheticalproteinMGC2198 222201665_xatHG-U133ANM001021197ribosomalproteinS17 223200674_satHG-U133ANM000994198ribosomalproteinL32 225229563_satHG-U133BBG231561200ribosomalproteinLioa 227212352_satHG-U133ABE780075202transmembranetraffickingprotein 231226073_atHG-U133BBE857362206hypotheticalproteinLOC219854 239213414_satHG-U133ABE259729213ribosomalproteinS19 243219762_satHG-U133ANM015414217ribosomalproteinL36 chip 232238026_atHG-U133BA1458020207 207201002_satHG-U133A No.ProbeSetID US 9, 963, 747 B2 74

AwWN uuuu NNNN Rank 342 343 343 344 345 346 346 347 349 353 353 356 356 358358 358 359 361 365 367 368 369 372?? 373?? 373??374 ??375 ?? 380 381 385 387 392 393 395

resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp

TTPmarkerResponseTypeofspecificity I | | | | | | | | | | | | | | | | | | | | | | | | | | | | | |

Entrez19 GeneID 7332220 5529 1968 126328 6208 25804 23521 5589 84153 51241 11258 28973 4528 7311 6204 3094 388789 51504 6124 10542 4841 6167 23521 10899 6209 6222 10412 7284 5537 2067 25912 6167 166 9318 2959

Gene Symbol UBE2L3 PPP2R5E EIF2S3 NDUFA11 RPS14 LSM4 RPL13A PRKCSH AYP1 C14orf112 DCTN3 MRPS18B MTIF2 UBA52 RPS10 HINT1 LOC388789HSPC152 RPL4 HBXIP NONO RPL37 RPL13A JTB RPS15 RPS18 TINP1 TUFM PPP6C ERCC1 Clorf43 RPL37AES COPS2 GTF2B 1APredictiveMarkersUpregulatedIndicatorsofNon-Responseand/orShortTimetoProgression TABLE1-continued PROTEASOMEINHIBITORPREDICTIVEMARKERIDENTIFICATION 222NADHdehydrogenase(ubiquinone)1alpha 220proteinphosphatase2,regulatorysubunitB 221eukaryotictranslationinitiationfactor2, 232mitochondrialtranslationalinitiationfactor2 repairdeficiency,complementationgroup1(includesoverlapping antisensesequence) homologsubunit2(Arabidopsis) 219ubiquitin-conjugatingenzymeE2L3 subcomplex,1114.7kDa 224LSM4homolog,U6smallnuclearRNA 227proteinkinaseCsubstrate80K-H 229chromosome14openreadingframe112 235histidinetriadnucleotidebindingprotein1 236hypotheticalgenesupportedbyAF147354 248proteinphosphatase6,catalyticsubunit 250chromosome1openreadingframe43 subunit3gamma,52kDa associated(S.cerevisiae) 231mitochondrialribosomalproteinS18B 239hepatitisBvirusxinteractingprotein 246TGFbeta-induciblenuclearprotein1 247Tutranslationelongationfactor, 249excisionrepaircross-complementingrodent 251amino-terminalenhancerofsplit 233ubiquitinA-52residueribosomalprotein fusionproduct1 240non-POUdomaincontaining,octamer 253generaltranscriptionfactorIIB (B56),epsilonisoform 223ribosomalproteinS14 230dynactin3(p22) 237hypotheticalproteinHSPC152 238ribosomalproteinL4 241ribosomalproteinL37 242ribosomalproteinL13a 243jumpingtranslocationbreakpoint 244ribosomalproteinS15245ribosomalproteinS18 241ribosomalproteinL37 252COP9constitutivephotomorphogenic 225ribosomalproteinL13a 226Transcribedlocus 228AYP1protein 234ribosomalproteinS10 binding NO:Title mitochondrial SEQ ID

Public NM_006246 NM_012321 NM_002743 NM_007234 NM_002453 NM_016404 NM_006402 NM_012423 NM_001018NM_022551 NM_014886 NM_003321 NM_002721 NM_001983 NM_001130 NM_004236 NM_001514 ID BG531983 BE252813 A1743115 AF116710 BF942308 A1857788 BF982002 AF151037 BC005373 AF348700 AA320764 U27143 BF675218 A1953886 BC003129 BF216701 AF151056 BC000152 BF216701

chip HG-U133B HG-U133A HG-U1334 HG-U133A

245200682_satHG-U133A 246203338_atHG-U133A 248228690_satHG-U133B 249208645_satHG-U133A 250202737_satHG-U133A 251212790_xatHG-U133A 252227126_atHG-U133B 253200707HG-U133Aat 254226453_atHG-U133B 255223191_atHG-U133B 256204246_satHG-U133A 257208907_satHG-U133A 258203095_atHG-U133A 259221700_satHG-U133A 260200095_xatHG-U133B 261208826_xatHG-U133A 262226236_atHG-U133B263217774_satHG-U133A 264200089_satHG-U133B 265202300_atHG-U133A 266210470_xatHG-U133A 267200092_satHG-U133B 268200716_xatHG-U133A 269210434_xatHG-U133A 270200819_satHG-U133A 271201049_satHG-U133A 272201922_atHG-U133A 273201113_atHG-U133A 276223034_satHG-U133B 277200092_satHG-U133A 278217729_satHG-U133A279202467_satHG-U133A 274206174_sat 275203720_sat 280208066_sat No.ProbeSetID 247224936_at US 9, 963, 747 B2 75

Rank 397 398 400 402 404 405 406 407 411 412 413 414 416 418 421 422 424 427 428 430 430 432 436 438 440 440 443 443 443 444 448 453 454 454 455 457 459

resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp

TTPmarkerResponseTypeofspecificity | | | | | | | | | | | | | | | | | | | | | | | |

Entrez GeneID 3094 226 6159 58528 25798 6187 1739 8668 113263 55146 51292 6189 79668 6157 6175 6167 8625 83707 4728 539 440193 51495 7381 401466 26355 51068 6139 9804 64776 51068 6191 90637 6205 6203 6175 6049

Gene Symbol HINT1 ALDOA RPL29 RRAGD BRI3 RPS2 DLG1 EIF3S2 GLCCI1 ZDHHC4 GMPR2 RPS3A PARP8 RPL27A RPLPO RPL37 RFXANK MGC11134 NDUFS8 ATP50 KIAA1509 HSPC121UQCRB LOC401466 E2IG5 CGI-07 RPL17 TOMM20 Cilorfi NMD3 RPS4X LOC90637 RPS11 RPS9 RPLPO RNF6 1APredictiveMarkersUpregulatedIndicatorsofNon-Responseand/orShortTimetoProgression TABLE1-continued PROTEASOMEINHIBITORPREDICTIVEMARKERIDENTIFICATION mitochondrialF1complex,Osubunit (oligomycinsensitivityconferringprotein) protein8,23kDa(NADH-coenzymeQ inS9 subunit2beta,36kDa containingprotein 20homolog(yeast) member8 reductase) protein NO:Title SEQ ID 299203190_atHG-U133ANM002496272NADHdehydrogenase(ubiquinone)Fe—S 290 287202514_atHG-U133AAW139131260discs,largehomolog1(Drosophila) 288208756_atHG-U133AU36764261eukaryotictranslationinitiationfactor3, 293219033_atHG-U133ANM024615266poly(ADPribose)polymerasefamily, 300200818_atHG-U133ANM001697273ATPsynthase,H+transporting 281translocaseofoutermitochondrialmembrane308212773SatHG-U133ABG165094 316203403_satHG-U1334NM005977289ringfingerprotein(C3H2C3type)6 Public 281207721_xatHG-U133ANM005340254histidinetriadnucleotidebindingprotein1 257Ras-relatedGTPbindingD284221524_SatHGU133AAF272036 290220261_satHG-U133ANM018106263zincfinger,DHHCdomaincontaining4 275butyrate-inducedtranscript1302234000SatHGU133BAJ271091 277hypotheticalgenesupportedbyBC055092304226165_atHG-U133BBF674436305220942 _xatHG-U133ANM014367278growthtransformationanddependentprotein 309222785_xatHG-U133B282chromosome11openreadingframe1AJ250229 310222497_xatHG-U133BAL520719283NMD3homolog(S.cerevisiae) NM_001007284ribosomalproteinS4,X-linked311200933xatHGU133A 282214687_xatHG-U133AAK026577255aldolaseA,fructosebisphosphate 289227525_atHG-U133BAA058770262glucocorticoidinducedtranscript1 291217990_atHG-U133ANM016576264guanosinemonophosphatereductase2265ribosomalproteinS3A 292212391xatHG-U133A1925635 295208856_xatHG-U133ABC003655268ribosomalprotein,largePO 297202758_satHG-U133ANM003721270regulatoryfactorXassociatedankyrin 298223436_satHG-U133BBC005133271tRNAsplicing2'phosphotransferase1 303205849_satHG-U133ANM006294276ubiquinolcytochromecreductasebinding 279CGI-07protein306231870SatHGU133BBG291007 315211720_xatHG-U133ABC005863288ribosomalprotein,largePO ID 280ribosomalproteinL17307212270XatHG-U133ABG168283 312226650_atHG-U133BA1984061285hypotheticalproteinLOC90637 283213969_xatHG-U133ABF683426256ribosomalproteinL29 258brainprotein13285223376_satHG-U133BAB055977 286203107_xatHG-U133ANM002952259ribosomalproteinS2 294203034_satHG-U133ANM000990267ribosomalproteinL27a 296224767_atHG-U133BAL044126269RibosomalproteinL37 313200031_satHG-U133BNM001015286ribosomalproteinS11 chip 301227228_satHG-U133BAB040942274KIAA1509 314217747_satHG-U133ANM001013287riboso 317215963XatHG-U133A298200

No.ProbeSetID US 9, 963, 747 B2 77 84

Rank 460 462 463 464 465 467 470 471 480 481484 492 494 495 502 504 506 508 509 509 510 513 514 515 518 520 521 533 541 541 548 548 558 558 561 566 566

resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp TTPmarkerResponseTypeofspecificity

Entrez GeneID 51024 51082 6124 5499 4247 6142 55291 7979 388796 29997 6218 6173 6139 51287 6129 51256 1725 1351 3338 79897 10944 6175 29956 6169 708 57092 23521 26996 51100 6136 55696 6133 10569 58516 6130 23528 9045 57456

Gene Symbol TTC11 POLRID RPL4 PPPICA MGAT2 RPL18A C1lorf23 SHFM1 LOC388796 GLTSCR2 RPS17 RPL36A RPL17 E2IG2 RPL7 TBC1D7 DHPS COXSA DNAJC4 RPP21 SMAP RPLPO LASS2 RPL38 C1QBP PCNP RPL13A GPR160 SH3GLB1 RPL12 RBM22 RPL9 SLUT C12orf14 RPLTA ZNF281 RPL14 KIAA1143 1APredictiveMarkersUpregulatedIndicatorsofNon-Responseand/orShortTimetoProgression TABLE1-continued PROTEASOMEINHIBITORPREDICTIVEMARKERIDENTIFICATION AK001105314LAG1longevityassurancehomolog2(S.cerevisiae) glycoproteinbeta-1,2mannosyl(alpha6)BC006390296 NM_015972292polymerase(RNA)IpolypeptideD,16kDa AK023950298chromosome11openreadingframe23 L04636316complementcomponent1,4subcomponent 320SH3-domainGRB2likeendophilinB1AF263293 BG479856324chromosome12openreadingframe14 NM_002708294proteinphosphatase1,catalyticsubunit NM_006304299splithand/footmalformation(ectrodactyly) AF296124301gliomatumorsuppressorcandidateregion AF151073307TBC1domainfamily,member7 NM_005528310DnaJ(Hsp40)homolog,subfamilyC BC000181319Gprotein-coupledreceptor160 U39402322RNAbindingmotifprotein22 alphaisoform type1 gene2 (ubiquitous) member4 NM_001002313ribosomalprotein,largePO bindingprotein AA843238323StepIIsplicingfactorSLUZ NM_016068291tetratricopeptiderepeatdomain11 N-acetylglucosaminyltransferase297ribosomalproteinL18a U52111 BC004886302ribosomalproteinS17 BE733979304ribosomalproteinL17 NM_004074309cytochromecoxidasesubunit8A NM_024839311ribonucleaseP21kDasubunit NM_020357317PEST-containingnuclearprotein BC001675318ribosomalproteinL13a BE559788325ribosomalproteinL7a AA838274327ribosomalproteinL14 NO:Title LOC388796AA827892hypothetical300 BE748698312smallacidicprotein NM_012482326zincfingerprotein281 ID NM_000968293ribosomalproteinL4 NM_021029303ribosomalproteinL36a NM_000971306ribosomalproteinL7 NM_000999315ribosomalproteinL38 NM_000976321ribosomalproteinL12 NM_000661186ribosomalproteinL9 A1745170328KIAA1143protein SEQ 295 NM_016565305E2IG2protein NM_013407deoxyhypusine308synthase PublicRep ID AC004692 BC000181

?P HG-U133AHG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A 324216383_atHG-U133A 325222467_satHG-U133B 326202276_atHG-U133A 329211487_xatHG-U133A 330201406_atHG-U133A 331212537_xatHG-U133A 332220647_satHG-U133A 333200717_xatHG-U133A 334223461_atHG-U133B335207831_xatHG-U133A 336201119_satHG-U133A 338218836_atHG-U133A 339200084_atHG-U133B 340201033_xatHG-U133A 341222212_satHG-U133A 342202029_xatHG-U133A 343208910_satHG-U133A 344217816_satHG-U133A 345210646_xatHG-U133A 346223423atHG-U133B 347209091_SatHG-U133A 348200809_xatHG-U133A 349224068_xatHG-U133B 350200032_satHG-U133A 351227990_atHG-U133B 352223038_satHG-U133B 353224930_xatHG-U133B 354218401_satHG-U133A 355213588_xatHG-U133A 356226816_satHG-U133B 320201154_xat 321200846_sat _Sat323211061 328234339_sat 337206782_sat No.ProbeSetID 318218034_at319218258_at 322217313_at 32765588_at US 9, 963, 747 B2 64 80

Rank 568 570571 573 573 577 582 588 594 594 596 598602 614 621 623 625 626 636 637 641 644 646 651 652 654 656 658 662 663 667 680 38 138 190 198

resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp

TTPmarkerResponseTypeofspecificity | | | | | | | | | | | | | | | | | | | | | | | | | | | | | |

11 | | | Entrez GeneID 5962 10944 6596 2971 1937 25870 10477 6124 6228 124540 10651 10972 57149 84311 55967 7086 6434 374395 25804 23588 54664 1153 170622 51100 60313 60496 6124 1329 81542 1485 11052 2778 2778 140823

Gene Symbol RDX SMAP SMARCA3 GTF3A EEF1G SUMF2 UBE2E3 RPL4 RPS23 MSI2 MTX2 TMP21 LOC57149 MRPL45 DAP13 ??? SFRS10 LOC374395 LSM4 KLHDC2 FLJ11273 CIRBP COMMD6 SH3GLB1 SP192 AASDHPPT RPLA COXSB TXNDC CTAGIB CPSF6 GNAS GNAS C20orf52 1APredictiveMarkersUpregulatedIndicatorsofNon-Responseand/orShortTimetoProgression TABLE1-continued PROTEASOMEINHIBITORPREDICTIVEMARKERIDENTIFICATION dependentregulatorofchromatin,subfamily 333eukaryotictranslationelongationfactor1 335ubiquitin-conjugatingenzymeE2E3 S23ein 338musashihomolog2(Drosophila) 341hypotheticalproteinA-211C6.1 345splicingfactor,arginine/serine-rich10 (transformer2homolog,Drosophila) 347LSM4homolog,U6smallnuclearRNA 359cleavageandpolyadenylationspecificfactor 362Chromosome20openreadingframe52 330SWI/SNFrelated,matrixassociatedactin 331generaltranscriptionfactorIIIA 334sulfatasemodifyingfactor2 (UBC4/5homolog,yeast) 342mitochondrialribosomalproteinL45 34313kDadifferentiation-associatedprotein 344transketolase(Wernicke-Korsakoff associated(S.cerevisiae) 160coldinducibleRNAbindingprotein 352SH3-domainGRB2likeendophilinB1 354aminoadipate-semialdehydedehydrogenase phosphopantetheinyltransferase 356cytochromecoxidasesubunitVb 340transmembranetraffickingprotein 346similartoRIKENcDNA1810059G22 348kelchdomaincontaining2 350hypotheticalproteinFLJ11273 353hypotheticalproteinSP192 355ribosomalproteinL4 357thioredoxindomaincontaining 358cancer/testisantigen1B 312smallacidicprotein a,member3 238ribosomalproteinL4 syndrome) 351COMMdomaincont 361GNAScomplexlocus gamma 339metaxin2 6,68kDa 360GNAScomplexlocus NO:Title 329radixin 337ribosomal SEQ ID 332 336 349

RepPublic NM_002097 NM_001404 NM_014315 NM_018374 NM_001280 AV704551 NM_001862 NM_030755 NM_000516 ID AL137751 BE748698 A1760760 AA243143 BE349022 AB017644 BC000451 H72927 AA112507 A1814644 AF257318 AA741071 AF302110 BC005817 AJ275978 AU149367 AF064092 BF381837

chip HG-U133A HG-U133A HG-U133A 369203517atHG-U133ANM_006554 370200929_atHG-U133ANM006827 HG-U133A 365200089_satHG-U133AA1953886 366217256_xatHG-U133AZ98950 367200926atHG-U133ANM001025 368225237_satHG-U133BBF435123 371203897_atHG-U133ABE963444 372224479SatHG-U133BBC006235 373223244SatHG-U133BAF217092 374208699_xatHG-U133ABF696840 360201338_xatHG-U133A 361214182_atHG-U133A 362200689_xatHG-U133A 363225002_satHG-U133B 364210024_satHG-U133A 376228089_xatHG-U133B 377202736_satHG-U133A 378217906_atHG-U133A 379224703atHG-U133B 380218930_satHG-U133A 381200810_satHG-U133A 382225312atHG-U133B 383210101_xatHG-U133A 384222452_satHG-U133B 385202169_satHG-U133A 386211710_xatHG-U133A 387202343_xatHG-U133A 388208097_satHG-U133A 389217339_xatHG-U133A 390202469_satHG-U133A 391214548_xatHG-U133A 392200780_xatHG-U133A 393224972_atHG-U133B 375200892_sat No.ProbeSetID 357212397_at 358200084at 359202983_at US 9, 963, 747 B2 28

Rank 280 o a 17 21 o ñ 33wa 34 Baba36 45 46 47 54 56 70 77 79 80 spö a

resp TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTPmarkerResponseTypeofspecificity

| | | | | | | | | | | | | | | | | | ! | I | I 11

Entrez GeneID 25814 4171 1434 9214 23179 4172 23246 9214 5885 7298 4175 1994 51729 4174 8243 3015 10236 7994 57570 8295 51053 27161 8907 4176 23 23225 5710 29894 5383

Gene Symbol ATXN10 MCM2 CSEIL TOSO RGL1 MCM3 BOP1 TOSO RAD21 TYMS MCM6 ELAVL1 WBP11 MCM5 SMC1L1 H2AFZ HNRPR MYST3 KLAA1393 TRRAP GMNN EIF2C2 AP1M1 MCM7 ABCF1 NUP210 PSMD4 CPSF1 PMS2L5 C6orf173 1APredictiveMarkersUpregulatedIndicatorsofNon-Responseand/orShortTimetoProgression TABLE1-continued PROTEASOMEINHIBITORPREDICTIVEMARKERIDENTIFICATION deficient6(MIS5homolog,S.pombe)cerevisiae 365CSE1chromosomesegregation1-like(yeast) deficient5,celldivisioncycle46(S.cerevisiae) 384eukaryotictranslationinitiationfactor20,2 391postmeioticsegregationincreased2-like5 374ELAV(embryoniclethal,abnormalvision Drosophila)-like1(HuantigenR chromosomes1-like(yeast) 392CDNAcloneIMAGE:4452583,partialcds deficient2,mitotin(S.cerevisiae) 379heterogeneousnuclearribonucleoproteinR 385adaptor-relatedproteincomplex1,mu 387ATP-bindingcassette,subfamilyF 389proteasome(prosome,macropain)26S 390cleavagepolyadenylationandspecificfactor deficient3(S.cerevisiae) 371RAD21homolog(S.pombe) 378H2Ahistonefamily,memberZ 380MYSTacetyltransferasehistone(monocytic deficient7(S.cerevisiae) subunit,non-ATPase4 366regulatorofFas-inducedapoptosis stimulator-like1 370regulatorofFas-inducedapoptosis 375WWdomainbindingprotein11 377SMC1structuralmaintenanceof 382transformation/transcriptiondomain 383geminin,DNAreplicationinhibitor 364MCM2minichromosomemaintenance 367ralguaninenucleotidedissociation 368MCM3minichromosomemaintenance 369blockofproliferation1 373MCM6minichromosomemaintenance 376MCM5minichromosomemaintenance leukemia)3 associatedprotein 386MCM7minichromosomemaintenance (GCN20),member1 388nucleoporin210kDa 372thymidylatesynthetase 1,160kDa 363ataxin10 subunit NO:Title 381KIAA1393 SEQ ID

Public NM_004526 NM_002388 NM_006265 NM_001071 NM_005915 NM_003496 NM_015895 NM_001090 NM_002810 NM_013291 ID AF119662 AF053640 AI084226 AF186779 BG491842 AF057557 BC003376 AF118023 AA807529 D80000 BF718636 BC001449 A1817830 AB037814 A1832074 AL562950 AF279900 AA502912 AA161026 BG492359

chip HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133B HG-U133B HG-U133B HG-U1334 394208833_satHG-U133A 395202107_satHG-U133A 396210766_satHG-U133A 397221601SatHG-U133A 398209568SatHG-U133A 406217821_satHG-U133A 407216237_satHG-U133A 412221952XatHG-U133A 413202642_satHG-U133A 417210983_satHG-U133A 419212316_atHG-U133A 420200882_satHG-U133A 422213893_xatHG-U133A 423226936_atHG-U133B 401221602_sat 402200608_sat 409213911_sat 410208766_sat 414218350_sat 421201639_sat No.ProbeSetID 399201555_at 400212563_at 403202589_at 404201930_at 405201726_at 408201589_at 411226547_at 415225827_at 416223024_at 418200045_at US 9, 963, 747 B2 €8 84

Rank 89 100 102 105106 107 108 110 112 121 126 131 136 140 146 148 152 155 159 160 164 166 174 180 182 184 187 193 197 199 203

TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP? TTPmarkerResponseTypeofspecificity

| | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | |

Entrez GeneID 408186 440080 29128 29964 47 6628 2339 54585 1656 2773 389835 51433 5591 1663 56660 55771 252839 6241 790 7716 6857 27346 27338 5425 3015 51433 116841 85453 203068 23216 6597

OVOSIII Gene Symbol LOC440080 UHRF1 C6orf49 ACLY SNRPB FNTA LZTFL1 DDX6 GNAI3 MGC57827 ANAPC5 PRKDC DDX11 KCNK12 FLJ11029 TMEMO RRM2 CAD ZNF161 SYT1 MAC30 UBEZS POLD2 H2AFZ ANAPC5 IMAGE3451454 TSPYL5 TUBB TBC1D1 SMARCA4 1APredictiveMarkersUpregulatedIndicatorsofNon-Responseand/orShortTimetoProgression TABLE1-continued PROTEASOMEINHIBITORPREDICTIVEMARKERIDENTIFICATION

polypeptide11(CHL1-likehelicasehomolog, 407potassiumchannel,subfamilyKmember12 dependentregulatorofchromatin,subfamily 398farnesyltransferase,CAAXboxalpha 399leucinezippertranscriptionfactor-like1 400DEAD(Asp-GluAla)boxpolypeptide6 401guaninenucleotidebindingprotein(G protein),alphainhibitingactivitypolypeptide3 411carbamoyl-phosphatesynthetase2,aspartate 417polymerase(DNAdirected),delta2 394ubiquitin-like,containingPHDandRING 395chromosome6openreadingframe49 404anaphasepromotingcomplexsubunit5 405proteinkinase,DNA-activatedcatalytic 406DEAD/H(Asp-GluAlaHis)box transcarbamylase,anddihydroorotase 404anaphasepromotingcomplexsubunit5 423TBC1(tre-2/USP6,BUB2cdc16)domain 393ovostatin/similarto-2 402CDNAFLJ41375fis,cloneBRCAN2007700403SimilartoRIKENDNA2700049P18gene 410ribonucleotidereductaseM2polypeptide 416ubiquitin-conjugatingenzymeE2S kDasubunit50regulatory 419H2Ahistonefamily,memberZ 424SWI/SNFrelated,matrixassociatedactin fingerdomains,1 397smallnuclearribonucleoproteinpolypeptides 408HypotheticalproteinFLJ11029409transmembraneprotein9 415hypotheticalproteinMAC30 family,member1 cerevisiae).S 412zincfingerprotein161 413synaptotagminI 421TSPY-like5 422tubulin,betapolypeptide member4,a 396ATPcitratelyase BandB1 polypeptide 420SVAP1protein NO:Title SEQ ID 414 418

RepPublic NM_001096 NM_020347 NM_022055 NM_004341 ID AW594320 AK025578 AF216754 J04564 BG168896 BF129093 JO3198 BE739287 AL135396 T33068 U34994 A1983033 BG165011AF151020 BC001886 AF254822

HG-U133B HG-U133B HG-U133A HG-U133B HG-U133A HG-U133B HG-U133B HG-U133B HG-U133A HG-U133A HG-U133A HG-U133BHG-U133B HG-U133A HG-U133A HG-U133A chip HG-U133AAU146275443202171at 447202779_satHG-U133ANM014501 448201115_atHG-U133ANM006230 450200853_atHG-U133ANM002106 444203999atHG-U133AAV731490 445214526XatHG-U133ANM005394 446212282_atHG-U133ABF038366 449214756_xatHG-U133AAB017004 451200098_satHG-U133AT33068 452225244_atHG-U133BAA019893 455212350_atHG-U133AAB029031 424228245_satHG-U133B 426223516_satHG-U133B 427201128_satHG-U133A 428208821atHG-U133A 453213122atHG-U133AA1096375 454211714XatHG-U133ABC005838 430218437_sat 432201180_sat 435200098_sat 436210543_sat 437213378_sat 440222988_sat 441209773_sat 442202715_at 456215714_sat No.ProbeSetID 425225655_at 429200090_at 431225549_at 433235242at 434225834_at 438220448_at 439228273_at US 9, 963, 747 B2 85 98

ONAva Rank 206 208 212 213 215 221 224 228 230 232 233 237 241 242 245 249 251 255 256 257 263 267 269 270 275 NNNN286 291

? TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTPmarkerResponseTypeofspecificity | | |

| | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Entrez GeneID 5728 23350 5937 27346 1870 3837 10987 9343 5781 252839 84677 203068 5757 9319 59349 10535 6597 55536 51593 83930 3014 57493 3184 10921 4207 7884 23528 5906 81611 23225 1810 8703

SembolGene Symbol PTEN SR140 RBMS1 MAC30 E2F2 KPNB1 COPS5 U5-116KD PTPN11 TMEM9 DSCR8 TUBB PTMA TRIP13 KLHL12 RNASEH2A SMARCA4 RAM2 ARS2 STARD3NL H2AFX HEG HNRPD RNPS1 MEF2B SLBP ZNF281 RAPIA ANP32E NUP210 DR1 B4GALT3 1APredictiveMarkersUpregulatedIndicatorsofNon-Responseand/orShortTimetoProgression TABLE1-continued PROTEASOMEINHIBITORPREDICTIVEMARKERIDENTIFICATION dependentregulatorofchromatin,subfamily polypeptideB(myocyteenhancerfactor2B) homologsubunit5(Arabidopsis) (AU-richelementRNAbindingprotein1, polypeptide3galactosyltransferase, multipleadvancedcancers1) type11(Noonansyndrome1) 453RAP1A,memberofRASoncogenefamily binding(negativecofactor2) interactingprotein1 32family,memberE a,member4 37kDa) NO:Title SEQ ID 456down-regulatoroftranscription1,TBP489209188XatHGU133ABC002809 425phosphataseandtensinhomolog(mutatedin457204053_xatHG-U133AU96180 429karyopherin(importin)beta1462208974XatHG-U133ABC003572 465212610_atHG-U133AU79291432proteintyrosinephosphatase,nonreceptor 481200060_satHG-U133BBC001659448RNAbindingproteinS1,serinerichdomain 482205124_atHG-U133ANM005919449MADSboxtranscriptionenhancerfactor2, BC002360431U5snRNP-specificprotein,116kD464222398_satHGU133B 470200773_xatHG-U133ANM002823437prothymosin,alpha(genesequence28) 474213720_satHG-U133AA1831675441SWI/SNFrelated,matrixassociatedactin 487221505_atHG-U133AAW612574454acidic(leucinerich)nuclearphosphoprotein 490210243_satHG-U133AAF038661457UDPGal:betaGlcNAcbeta1,4 427RNAbindingmotif,singlestranded459225265_atHG-U133BA1580100 428E2Ftranscriptionfactor2461228361atHG-U133BAL561296 AA770014434Downsyndromecriticalregiongene8467241224_xatHG-U133B 472225068_atHG-U133BAK024412439kelchlike12(Drosophila) 483206052_satHG-U1334NM006527450stemloop(histone)bindingprotein RepPublic 426U2-associatedSR140protein458212058_atHGU133AAI184562 415hypotheticalproteinMAC30460212281SatHG-U133ABF038366 471204033_atHG-U133ANM004237438thyroidhormonereceptorinteractor13 473203022_atHG-U133ANM006397440ribonucleaseH2,largesubunit 478205436_satHG-U133ANM002105445H2Ahistonefamily,memberX 451Zincfingerprotein281HG-U133BAU150752484222619at ID 476201680_xatHG-U133ANM015908443arsenateresistanceproteinARS2 477223065_satHG-U133BBC003074444STARD3Nterminallike 480200073_satHG-U133AM94630447heterogeneousnuclearribonucleoproteinD 463201652_atHG-U133ANM006837430COP9constitutivephotomorphogenic 466222987_satHG-U133BNM016456433transmembraneprotein9 469209026_xatHG-U133AAF141349436tubulin,betapolypeptide 475225081_satHG-U133BAK022955442transcriptionfactorRAM2 446HEGhomolog479213069atHG-U133AAI148659 488213947_satHG-U133AA1867102455nucleoporin210kDa chip 468207057atHG-U133ANM004731435 485225684_atHG-U133BBG496998452

No.ProbeSetID 486202362_at US 9, 963, 747 B2 88

Rank 170taa 10 12 14 16 O 40 42 43 48 48 52 53 59 OS 73 74 86 94 103 115 119 123

TTP/resp TTP/respTTP /respTTP /respTTP /respTTP/resp TTP/respTTP /respTTP /respTTP /respTTP /resp TTP/resp TTP/respTTP /resp TTP/resp TTP/respTTP/resp TTP/resp TTP/resp TTP/resp TTP/respTTP /respTTP /resp TTP/resp TTP/resp TTP/resp TTP/resp TTP/resp TTP/respTTP /respTTP/resp TTP /resp TTP/respTTP /resp TTP/resp TTP

TTPmarkerResponseTypeofspecificity | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | |

| | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | |

GeneID 5203 2990 5725 1537 5725 5725 1345 5725 5725 515 2029 7764 7155 5728 1810 835 5094 2778 1460 Entrez 29901 57820 219927 51497 64207 51497 26051 51497 10657 55334 221908 56947 10572 23369 26136 27346 374491 /55269

PSPC1/ Gene Symbol SHD1 CCNB11P1 PFDN4 GUSB MRPL21 TH1L C14orf4 PTBP1 CYC1 PTBP1 PTBP1 TH1L COX6C PTBP1 PPP1R16B THIL PTBP1 KHDRBS1 ATP5F1 SLC39A9 MGC22793 ENSA ZNF217 TOP2B PTEN C2orf33 SIVA DR1 CASP2 PUM2 PCBP2 TES MAC30 GNAS LOC374491 CSNK2B 1APredictiveMarkersUpregulatedIndicatorsofNon-Responseand/orShortTimetoProgression TABLE1-continued PROTEASOMEINHIBITORPREDICTIVEMARKERIDENTIFICATION mitochondrialF0complex,subunitbisoform1 developmentallydown-regulated2) 492paraspecklecomponent1/TPTEandPTEN 458Sac3homologydomain1(S.cerevisiae) 465polypyrimidinetractbindingprotein1 467polypyrimidinetractbindingprotein1 471polypyrimidinetractbindingprotein1472proteinphosphatase1,regulatory(inhibitor) 475KHdomaincontaining,RNAbindingsignal 477solutecarrierfamily39(zinctransporter), 481topoisomerase(DNA)IIbeta180kDa 482phosphataseandtensinhomolog(mutatedin multipleadvancedcancers1) 489testisderivedtranscript(3LIMdomains) 462mitochondrialribosomalproteinL21 464chromosome14openreadingframe 468polypyrimidinetractbindingprotein1 474polypyrimidinetractbindingprotein1 483chromosome2openreadingframe33 485down-regulatoroftranscription1,TBP binding(negativecofactor2)486caspase2,apoptosis-relatedcysteine protease (neuralprecursorcellexpressed, 487pumiliohomolog2(Drosophila) homologousinositollipidphosphatase 459cyclinB1interactingprotein1 470cytochromecoxidasesubunitVIC transductionassociated1476ATPsynthase ,Htransporting+ 484CD27-binding(Siva)protein 493caseinkinase2,betapolypeptide 461glucuronidase,beta 463TH1-like(Drosophila) 466cytochromec-1 469TH1-like(Drosophila) 473TH1-like(Drosophila) 478hypotheticalproteinMGC22793 480zincfingerprotein217 490hypotheticalproteinMAC30 491GNAScomplexlocus 460prefoldin4 subunit16B member9 479endosulfinealpha pseudogene NO:Title SEQ ID 488

Public NM_006559 NM_006526 NM_001068 NM_001320 ID AJ238379 AJ238374 BC004383 BC005960 AK025831 BF000655 BC000436 AF023139 AU153405

494202605_atHG-U133ANM000181 U133AHG- HG-U133B HG-U133B HG-U133A chip 491205449_atHG-U133ANM013299 492217988_atHG-U133ANM021178 499201066_atHG-U133ANM001916 500202189_xatHG-U133ANM002819 503201754_atHG-U133ANM004374 521204031SatHG-U133ANM_005016 522202720_atHG-U133ANM015641 493205361_satHG-U133AAI718295 497223474_atHG-U133BA1932310 498216306_xatHG-U133AX62006 501211270_xatHG-U133ABC002397 AJ238379502225006_xatHG-U133B 504212015_xatHG-U133ABF690062 50541577_atHG-U133AAB020630 516223354_xatHG-U133BBC003191 518209187_atHG-U133AAW516932 520201493SatHG-U133ABE778078 523212279_atHG-U133ABE779865 524211858_xatHG-U133AAF088184 525226574_atHG-U133BA1872384 495225315_atHG-U133BBF344406 496225261XatHG-U133BAJ238376 506225865_xatHG-U133B 507211271_xatHG-U133A508200040_atHG-U133B 511226434_atHG-U133B??? 512202596_atHG-U133A HG-U133A513203739_at HG-U133A514211987_at 515217492SatHG-U133A 517210792_xatHG-U133AAF033111 211755at_s509 526201390_sat No.ProbeSetID 510222445_at 519226032_at US 9, 963, 747 B2 68 06

Rank 134 134 139 149 153 168 173 76 196 209 210 219 234 236 238 244 244 272 285 289 292NNAEN PAWN14 15 16 17 23 26 27

TTP/resp TTP/resp TTP/respTTP /respTTP /respTTP /resp TTP/respTTP /resp TTP/respTTP/resp TTP/resp TTP/respTTP /resp TTP/respTTP /respTTP/resp TTP/respTTP/resp TTP/resp TTP/respTTP /resp resp 110resp resp resp 3resp resp resp resp resp resp resp resp resp resp resp resp TTPmarkerResponseTypeofspecificity | | | | | | | | | | | | | | | | | | | | | |

Entrez GeneID 3020 83940 64645 23225 5757 1964 55860 4715 7528 94239 8454 6903 8663 51320 220988 8663 56947 8663 6434 51444 9588 6201 60674 6201 4707 60674 441951 441951 6125 50854 2926 51386 441951 10480 900 8665 6227

Gene Symbol ????? TATDN1 HIAT1 NUP210 PTMA EIF1AX ACTR10 NDUFB9 YY1 H2AFV CUL1 ???? EIF358 RKHD2 HNRPA3 EIF358 C2orf33 EIF358 SFRS10 RNF138 PRDX6 RPS7 GAS5 RPS7 NDUFB1 GAS5 RPL5 C6orf48 GRSF1 EIF3S6IP GA17 CCNG1 EIF355 RPS21 1APredictiveMarkersUpregulatedIndicatorsofNon-Responseand/orShortTimetoProgression TABLE1-continued PROTEASOMEINHIBITORPREDICTIVEMARKERIDENTIFICATION

498eukaryotictranslationinitiationfactor1A,X 499actin-relatedprotein10homolog(S.cerevisiae)500NADHdehydrogenase(ubiquinone)1beta 505eukaryotictranslationinitiationfactor3, 508eukaryotictranslationinitiationfactor3, 516NADHdehydrogenase(ubiquinone)1beta 523eukaryotictranslationinitiationfactor3, 526eukaryotictranslationinitiationfactor3, 497prothymosin,alpha(genesequence28) 507heterogeneousnuclearribonucleoproteinA3 510eukaryotictranslationinitiationfactor3, 511splicingfactor,arginine/serine-rich10(transformer2homolog,Drosophila) 521chromosome6openreadingframe48 496hippocampusabundanttranscript1 subcomplex,922kDa 502H2Ahistonefamily,memberV 506ringfingerandKHdomaincontaining2 509chromosome2openreadingframe33 subcomplex,17kDa subunit5epsilon,47kDa 495TatDDNasedomaincontaining1 504tubulin-specificchaperonec 515growtharrest-specific5 522G-richRNAsequencebindingfactor1 subunit6interactingprotein 494H3histone,family3A 388nucleoporin210kDa 501YY1transcriptionfactor subunit8,110kDa subunit8,110kDa kDa110subunit8, 512ringfingerprotein138 513peroxiredoxin6 514ribosomalproteinS7 514ribosomalproteinS7 517growtharrest-specific 518SimilartoRPE-spondin 519SimilartoRPE-spondin 520ribosomalproteinL5 524SimilartoRPE-spondin 52dendriticcellprotein 527ribosomalproteinS21 linked 503cullin1 525cyclinG1 NO:Title SEQD

PublicRep NM_016626 NM_003752 NM_004593 NM_006360 NM_003754 NM_001024 ID BG505670 AF258660 AA679705 BF185165 BC000196

HG-U133A HG-U133A chip 532201019_satHG-U133ANM001412 536202487_satHG-U133ANM012412 537207614satHG-U133ANM003592 538202495atHG-U1334NM003192 546218738_satHG-U133ANM016271 551206790_satHG-U133ANM004545 555200937_satHG-U133ANM000969 556220755_satHG-U133ANM016947 558217719_atHG-U133ANM016091 527211940_xatHG-U133ABE869922 528223231_atHG-U133BAF212250 530212315_SatHG-U133AAA502912 531216515_xatHG-U133AAL121585 533222230_satHG-U133AAK022248 535201901_satHG-U133AZ14077 539210949_satHG-U133ABC000533 547200844_satHG-U133ABE869583 548200082SatHG-U133AAI805587 549224841_xatHG-U133BBF316352 552224741_xatHG-U133BBG329175 553226835SatHG-U133BBG330520 557201520_satHG-U133ABF034561 529225222atHG-U133BA1243268 534222992SatHG-U133BAF261090 540218247_satHG-U133A 541211931_satHG-U133A 542200647_xatHG-U133A 543223091_xatHG-U133B 544215230_xatHG-U133A 550200082SatHG-U133BA1805587 554224915XatHG-U133BAV756131 559226227_xatHG-U133B 560202232_satHG-U133A 561208796SatHG-U133A 562200023_satHG-U133B 563200834_sat No.ProbeSetID 545200893_at US 9, 963, 747 B2

Rank 104 105 107 112 121 122 124 1275 132 146 158 160 176 189 195 206 219 236 248 253 255 294 296

resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp esp'l resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp

TTPmarkerResponseTypeofspecificity | | | | | | | | | | | ' | | | | | | | | | | | | | | |

Entrez GeneID 6217 8665 114915 81892 6165 191 125144 738 10175 2197 5250 6633 5250 4869 1975 10284 6142 93487 7812 51002 9512 10632 10473 2197 10632 391356 51187 84973 51187 W

Gene Symbol RPS16 EIF355 TIGA1 C14orf156 RPL35A AHCY MGC40157 Cilorf2 CNIH FAU SLC25A3 SNRPD2 SLC25A3 NPM1 EIF4B SAP18 RPL18A C14orf32 UNR CGI-121 PMPCB ATP5L HMGN4 FAU ATP5L C15orf15 MGC16037 C15orf15 1APredictiveMarkersUpregulatedIndicatorsofNon-Responseand/orShortTimetoProgression TABLE1-continued PROTEASOMEINHIBITORPREDICTIVEMARKERIDENTIFICATION mitochondrialF0complex,subunitg mitochondrialF0complex,subunitg (FBR-MuSV)ubiquitouslyexpressedfox carrier;phosphate),member3 carrier;phosphate),member3 (FBR-MuSV)ubiquitouslyexpressedfox subunit5epsilon,47kDa derived);ribosomalproteinS30 kDapolypeptide.516 derived);ribosomalproteinS30 AF165521553chromosome15openreadingframe B23,numatrin) domain4 NO:Title SEQ ID 588201682_atHG-U133ANM004279549peptidase(mitochondrialprocessing)beta 565200023_satHG-U133ANM003754526eukaryotictranslationinitiationfactor3, 568221434_satHG-U133ANM031210531chromosome14openreadingframe156 576200019_satHG-U133BNM001997538FinkelBiskisReillymurinesarcomavirus 582211937_atHG-U133ANM001417543eukaryotictranslationinitiationfactor4B 544sin3-associatedpolypeptide,18kDa583208742_satHGU133A078303 585212644_satHG-U133AA1671747546chromosome14openreadingframe32 589207573_xatHG-U133ANM006476550ATPsynthase,H+transporting 591200019_satHG-U133ANM001997538FinkelBiskisReillymurinesarcomavirus 592210453_xatHG-U133AAL050277551ATPsynthase,H+transporting 596217915_satHG-U133ANM016304555chromosome15openreadingframe Public 574217969_atHG-U133ANM013265536chromosome11openreadingframe2575201653_atHG-U133ANM005776537cornichonhomolog(Drosophila) 577200030_satHG-U133ANM002635539solutecarrierfamily25(mitochondrial 579200826_atHG-U133ANM004597541smallnuclearribonucleoproteinD2 580200030_satHG-U133BNM002635539solutecarrierfamily25(mitochondrial 590209786_atHG-U133ABC001282193highmobilitygroupnucleosomalbinding 593226243_atHG-U133BBF590958552SimilartoCG14903PA ID 564201258_atHG-U133ANM001020528ribosomalproteinS16 571200903_satHG-U133ANM000687533Sadenosylhomocysteinehydrolase 581221691_xatHG-U133AAB042278542nucleophosmin(nucleolarphosphoprotein 584200869_atHG-U133ANM000980545ribosomalproteinL18a 587219030_atHG-U133ANM016058548CGI121protein 569225190_xatHG-U133BAW402660532ribosomalproteinL35a 573225065_xatHG-U133BAI826279535hypotheticalproteinMGC40157 NRAS586222975_SatHG-U133BA1423180upstreamof547 595229050_satHG-U133BAL533103554hypotheticalproteinMGC16037 566225698_atHG-U133BBF314746529TIGA1 567200024_atHG-U133BNM001009530 chip 570200024_atHG-U133ANM001009530 572234875atHG-U133BAJ224082534 540578216380_xatHG-U133AAC005011

No.ProbeSetID 594222465_at US 9, 963, 747 B2 93 94

Rank 309 322 325 326 357 360 368 376 389 424 495 496 503 516 524 526 529 3 266 283 1 N w

TTP/resp TTP/resp TTP/resp TTP/resp TTP/resp TTP/resp TTP/respTTP/resp TTP/resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp TTP TTP TTP/resp TTPmarkerResponseTypeofspecificity

Entrez GeneID 5684 442534 6392 6146 10054 3615 11335 23478 6175 498 58533 6158 51520 23521 2992 7528 30848 10598 4111 1485/ 246100 1485/ 246100 4893 2079 2079 64093 23658 5150 694 11335

Gene Symbol PSMA3 SDHD RPL22 UBA2 IMPDH2 CBX3 SEC11L1 RPLPO ATP5?1 SNX6 RPL28 LARS RPL13A GYG YY1 CTAG2 AHSA1 MAGEA12 CTAG1B CTAGIA CTAGIB / CTAGIA NRAS ERH ERH SMOC1 LSM5 PDE7A BTG1 CBX3 1APredictiveMarkersUpregulatedIndicatorsofNon-Responseand/orShortTimetoProgression TABLE1-continued PROTEASOMEINHIBITORPREDICTIVEMARKERIDENTIFICATION 558succinatedehydrogenasecomplex,subunitD 556proteasome(prosome,macropain)subunit mitochondrialF1complex,alphasubunit 578NeuroblastomaRASviral(v-ras)oncogene 574AHA1,activatorofheatshock90kDaprotein 576cancer/testisantigen1B 580SPARCrelatedmodularcalciumbinding1 560SUMO-1activatingenzymesubunit2 561IMP(inosinemonophosphate)dehydrogenase2 562chromoboxhomolog3(HP1gamma 564ribosomalprotein,largePO ATPasehomolog1(yeast) 575melanomaantigenfamilyA,12 577cancer/testisantigen1B 581LSM5homolog,U6smallnuclearRNA 583B-celltranslocationgene1,antiproliferative 562chromoboxhomolog3(HP1gamma integralmembraneprotein homolog,Drosophila)563SEC11 -like1(cerevisiaeS.) 565ATPsynthase,H+transporting isoform1,cardiacmuscle 579enhancerofrudimentaryhomolog 579enhancerofrudimentaryhomolog associated(S.cerevisiae) homolog,Drosophila) alphatype,3 559ribosomalproteinL22 568ribosomalproteinL28 nin 573cancer/testisantigen2 (Drosophila) (Drosophila) 582Phosphodiesterase7A 566sortingnexin6 569leucyl-tRNAsynthetase 570ribosomalproteinL13a 572YY1transcriptionfactor antigen1A antigen1A homolog NO:Title 557LOC442534 571gly SEQID 567

PublicRep NM_002788 NM_003002 NM_005499 NM_000884 NM_016587 NM_000991 NM004130NM004130 NM_003403 NM_012111 NM_004450 NM_004450 NM_001731 NM_016587 ID AI798822 BE250348 AF121856 AJ249900 AK021413 BC000514 AJ012833 BC003408 U87459 AF038567 BE964484 BF516292 BC005938 AW269834

chip HG-U133A HG-U133BHG-U133A HG-U133A HG-U133B HG-U133A HG-U133B HG-U133B HG-U133A 604216274_satHG-U133AN99438 605214167_SatHG-U133AA555113 600221726_atHG-U133A 601201177_satHG-U133A 602201892_satHG-U133A 603200037_satHG-U133B 606213738SatHG-U133AA1587323 607222410_satHG-U133B 608222784_atHG-U133B 609200003_satHG-U133B 610222427_satHG-U133B 611200715_xatHG-U133A 612201554xatHG-U133A 613200047_satHG-U133B 614215733_xatHG-U133A 615201491_atHG-U133A 616210467_xatHG-U133A 617210546XatHG-U133A 624223358_satHG-U133B625200921 _satHG-U133A 626200037_satHG-U133A 618211674_xat 622222783_sat 623211747_sat No.ProbeSetID 597201532_at 598239237_at 599202026_at 619224985_at 620200043_at 621200043_at US 9, 963, 747 B2 95 96

Rank 118 143 240 248 271 82 118 170 179 187 298 349 362 379 451 599 683 67 127 226 264 Ar zu 222

TTP/resp TTP/respTTP /respTTP /respTTP /resp TTP/resp TTP/respTTP/resp TTP /resp TTP/resp TTP/resp resp resp resp resp resp resp resp resp resp resp resp resp resp TTP TTP TTP TTP TTP TTP TTP

TTPmarkerResponseTypeofspecificity | | | | | | | II | | | | | | | |

| | | | | | | | | | | | | | | | |

Entrez GeneID 7529 51097 7528 6742 4191 6224 7812 6224 517 6184 10914 7381 6272 6430 5859 10473 3476 6430 3151 10914 29964 56943 8994 163259 1810 9556 22827 9112 4738 694 54665

CGI-49 Gene Symbol YWHAB YY1 SSBP1 MDH2 RPS20 UNR RPS20 ATP5G2 RPN1 PAPOLA UQCRB SORT1 SFRS5 QARS HMGN4 IGBP1 SFRS5 HMGN2 PAPOLA C6orf49 e(y)2 LIMD1 FLJ37099 DR1 C14orf2 SIAHBP1 MTA1 NEDD8 BTG1 RSBN1 1APredictiveMarkersUpregulatedIndicatorsofNon-Responseand/orShortTimetoProgression TABLE1-continued PROTEASOMEINHIBITORPREDICTIVEMARKERIDENTIFICATION mitochondrialF0complex,subunitc( monooxygenaseactivationprotein,beta 599immunoglobulin(CD79A)bindingprotein1 developmentallydown-regulated8 584tyrosinemonooxygenase3-/tryptophan5 596splicingfactor,arginine/serine-rich5 600splicingfactor,arginine/serine-rich5 603chromosome6openreadingframe49 607down-regulatoroftranscription1,TBP binding(negativecofactor2) 612B-celltranslocationgene1,antiproliferative 586single-strandedDNAbindingprotein1 587malatedehydrogenase2,NAD 591ATPsynthase,H+transporting 594ubiquinol-cytochromecreductasebinding 598highmobilitygroupnucleosomalbinding 608chromosome14openreadingframe2 609fuse-bindingproteininteractingrepressor 611neuralprecursorcellexpressed, 613roundspermatidbasicprotein1 597glutaminyl-tRNAsynthetase 601high-mobilitygroupnucleosomalbinding 605LIMdomainscontaining1 588ribosomalproteinS20 proteinS20ribosomal590 593poly(A)polymerasealpha 602poly(A)polymerasealpha 610metastasisassociated1 585CGI-49protein 572YY1transcriptionfactor (mitochondrial) 589upstreamofNRAS 9),isoform2 domain4 domain2 polypeptide 592ribophorinI protein 595sortilin1 604e(y)2protein 606FLJ37099protein NO:Title SEQ ID

RepPublic NM_001023 NM_007158 NM_002950 NM_005051 NM_006353 NM_001551NM_006925 NM_017601 NM_020189 NM_014240 1916261 NM_001938 NM_006156 ID BC001359 BF184532 D13119 A1984479 M26700 BF447105 AW084582 BC003689 BF797555 AF116639 AF217197 BC006177 HG-U133AAL535380HG-U133BR45958 chip HG-U133A HG-U133A HG-U133A HG-U133AHG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A 629200047_satHG-U133ANM003403 630202591_satHG-U133ANM003143 628201825_satHG-U133AAL572542 631209036_satHG-U133ABC001917 632200949_xatHG-U133A 633219939_satHG-U133A 634214003_xatHG-U133A 635208764_satHG-U133A 636201011_atHG-U133A 637222035_satHG-U133A 638209066XatHG-U133A 646212718_atHG-U133A 647218233_satHG-U133A 648218482_atHG-U133A 649218850_satHG-U133A 650230769atHG-U133B 651207654_xatHG-U133A 627208743_sat 639212807_sat 640212266_sat 642202579_xat 644203380_xat 645208668_xat 652210532_sat 653209899_sat 654211783_sat 656200920_sat No.ProbeSetID 641217846_at 643202105_at 655201840_at 657222789_at US 9, 963, 747 B2 97 86

a Rank 116 12 13 18 20 29 31 33 34 36 8ã 42 46 56 57 64 65 66 N N ofType speci ficity resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp

Response marker + + + + + + + + + + + + + ++ + + ++ + + + + + + + + + + + + + + TTP marker

330 493 687 199 Entrez Gene ID 114884 440823 7305 1490 8732 30061 4818 1783 7940 10875 23180 51312 10475 3281 8717 28639 3117/3118 22980 7088 1508 22980 1545 197259

DQA1/ Gene Symbol OSBPL10 LOC440823 TYROBP CTGF RNGTT SLC40A1 NKG7 DNCLI2 LST1 FGL2 RAFTLIN MSCP BIRC3 TRIM38 HSBP1 TRADD ATP2B4 TRBC1 HLA HLA DQA2 KLAA1049 TLE1 KLF9 CTSB KIAA1049 CYP1B1 AIF1 MLKL 1BPredictiveMarkersUpregulatedIndicatorsofResponseand/orLongTimetoProgression

histocompatibilitycomplex,class 642cytochromeP450,family1subfamily 614oxysterolbindingprotein-like10 618RNAguanylyltransferaseand5' intermediatepolypeptide2 626BaculoviralIAPrepeat-containing3 629heatshockfactorbindingprotein1 TNFRSF1A-associatedviadeath classII,DQalpha1/major 637transducin-likeenhancerofsplit1 644mixedlineagekinasedomain-like 615hypotheticalgenesupportedby kinaseproteintyrosineTYRO factorgrowthtissueconnective617 (iron-regulatedtransporter), 622leukocytespecifictranscript1 628tripartitemotif-containing38 632ATPase,Ca+transportingplasma 634majorhistocompatibilitycomplex, Drosophila)homolog,(splE 643allograftinflammatoryfactor1 family40solutecarrier619 620naturalkillercellgroup7 621dynein,cytoplasmiclight 625Mitochondrialsolutecarrier 633Tcellreceptorbetaconstant1 638Kruppel-likefactor9 bindingprotein member1 623fibrinogen-like2 624raft-linkingprotein membrane4 II,DQalpha2 polypeptide1,B Title AK098833 phosphatase sequence protein domain 636KIAA1049protein 639CathepsinB 641KIAA1049protein SEQ ID NO: 627 630 631 635 640

RepPublic NM017784 NM003332 NM003800 NM_005601 NM007161 NM_006682 NM021039 NM006355 NM002122 ID BF475488 M92934 AL136944 BG110975 D42043 H69701 AA805622 AK026575 AW972351 L41690 AW517686 AL559122 AW070877 AF322111 A1758763 A1690205 W47179 N55205 BF000251 AU144855 BF213829 AA706818

HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133B HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B chip HG

204122_at 224616_at 214574_xat 231078_at 230499_at 208540_xat 203568s_at 212136_at 213193_xat 203290_at 229147_at 221495sat 203221_at 203542_sat 213275_xat 216063_at 213311sat 202436_sat 215051_xat 658219073_sat 659227168_at 661209101_at 662204208_at 223044at 213915at 204834at 212646at 200941at 222368at 1729at 238025at ProbeSetID No 660 663 664 665 666 667 668 669 670 671 672 673 674 675 676 677 678 679 680 681 682 683 684 685 686 687 688 US 9, 963, 747 B2 99 100

Rank 74 79 ooooooWO 86 90 91 93 96 98 101 102 105 108 111 117 119 124 129 146 148 153 ofType speci ficity resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp

Response marker + + + ++ ++ + + + ++ ++ + + + + ++ + + + + + ++ + + + + TTP marker

493 358 Entrez Gene ID 23019 2934 5713 11107 11153 1282 2222 79901 8732 1508 7846 3815 6515 1191 6818 1052 23190 81671 3837 3003 3423 7204

Gene Symbol CNOT1 GSN PSMD7 PRDM5 HYPE COL4A1 FDFT1 ATP2B4 CYBRD1 RNGTT CTSB TUBA3 KIT SLC2A3 CLU SULT1A3 CEBPD AQP1 UBXD2 VMP1 KPNB1 GZMK IDS TRIO 1BPredictiveMarkersUpregulatedIndicatorsofResponseand/orLongTimetoProgression continued- prosomemacropain)26Sproteasome,(647 646gelsolin(amyloidosis,Finnishtype) subunit,non-ATPase7(Mov34 653RNAguanylyltransferaseand5' message2,apolipoproteinJ)660sulfotransferase family,cytosolic 1A,phenol-preferringmember3 kDa)protein,28integral 665Karyopherin(importin)beta1 granzyme3;tryptaseII) 645CCR4-NOTtranscriptioncomplex, 649HuntingtininteractingproteinE 650collagen,typeIValpha1 farnesyltransferase1632ATPase,Ca+transportingplasma 655CDNAFLJ46553fis,clone sarcomaviraloncogenehomolog member3transporter), inhibitor,SP-40sulfated glycoprotein,2testosterone-repressedprostate 661CCAATenhancer/bindingprotein 662aquaporin1(channel-forming protein1membrane 666granzymeK(serineprotease, 667iduronate2-sulfatase(Hunter 668triplefunctionaldomain(PTPRF 648PRdomaincontaining5 652cytochromebreductase1 657v-kitHardyZuckerman4feline 658solutecarrierfamily2 659clusterin(complementlysis 663UBXdomaincontaining2 664likelyorthologofratvacuole syndrome) interacting) subunit1 homolog) 651farnesyl-diphosphate membrane4 656tubulin,alpha3 (facilitatedglucose (C/EBP),delta Title phosphatase 654cathepsinB THYMU3038879 SEQ ID NO:

RepPublic NM000177 NM002811 NM_018699 NM_007076 NM001845 NM024843 NM_000222 NM005195 NM002104 ID BC000779 BC003573 AW517686 AB012142 AA020826 AW575863 AF141347 A1631159 M25915 U08032 AL518391 A1927512 BF674052 AA861608 AF050145 AF091395

chip HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A

200860_sat 200696_sat 201705_at 220792_at 219910_at 211981at 210950s_at 212135_sat 217889_sat 204207sat 213274_sat 243780_at 209118_sat 205051_sat 202497_xat 208791_at 209607_xat 203973_sat 209047_at 213574_sat 206666_at 210666_at 209013xat ProbeSet 708212007_at 709224917_at No.ID 689 690 691 692 693 694 695 696 697 698 699 700 701 702 703 704 705 706 707 710 711 712 713 US 9, 963, 747 B2 101 102

Rank 153 155 155 159 160 161 164 168 172 175 177 188 191 199 200 203 205 206 209 211 215215 216 217 218 219 220 220 221 ofType speci ficity resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp respresp resp resp resp resp resp resp resp

Response marker + + + + + + + + + + + + + + + + + + + + + + + + + + + + TTP marker

390 939 975 Entrez Gene ID 6526 8717 85462 1488 253981 11216 65018 10347 201799 11313 9114 7474 64968 89122 166824 5788 5176 404636/55855 10479 8837 1236 7204 55824

FAM45B/ Gene Symbol SLC5A3 TRADD KIAA1727 CTBP2 LOC253981 AKAP10 PINK1 RND3 ABCAT TNFRSF7 FLJ32028 LYPLA2 ATP6VOD1 WNT5A MRPS6 TRIM4 RASSF6 PTPRC SERPINF1 FAM45A SLC9A6 CD81 CFLAR CCR7 TRIO PAG 1BPredictiveMarkersUpregulatedIndicatorsofResponseand/orLongTimetoProgression continued- 686Rasassociation(RalGDS/AF-6)domain antiproliferativeantibody1) 678ATP-bindingcassette,subfamilyA inhibitor,cladeFalpha(-2 689familywithsequencesimilarity45, transporters),member3 670TNFRSF1A-associatedviadeathdomain 674Akinase(PRKA)anchorprotein10 38kDa,VOsubunitdisoform1 684MitochondrialribosomalproteinS6 antiplasmin,pigmentepitheliumderivedfactor),member1 memberB/familywithsequence similarity45,memberA (sodiumhydrogenexchanger), 694chemokine(C-motif)receptor7 669solutecarrierfamily5(inositol 675PTENinducedputativekinase1 RhofamilyGTPase3 member7superfamily, 682ATPase,H+transportinglysosomal 683wingless-typeMMTVintegrationsite 685tripartitemotif-containing4 687proteintyrosinephosphatase, receptortype,C 692CASP8andFADD-likeapoptosis PTPRFdomainfunctional(triple695 enrichedglycosphingolipid- 672C-terminalbindingprotein2 (ABC1),member7 679tumornecrosisfactorreceptor 680hypotheticalproteinFLJ32028 family,member5A 688serine(orcysteine)proteinase 690solutecarrierfamily9 691CD81antigen(targetof 696phosphoproteinassociatedwith 673hypotheticalproteinLOC253981 681lysophospholipaseII family6 isoform6 interacting) Title 671KIAA1727protein regulator microdomains SEQ ID NO: 676 693

RepPublic NM003789 NM019112 NM001242 NM007260 NM_003392 NM002615 NM006359 NM004356 NM001838 NM_007118 ID A1867198 AA527080 AF222711 AI394438 AA456929 BF432478 A1634046 BG054844 AA806283 AL566172 AK024896 AF220023 A1167789 Y00062 BE565675 AF005775 A1358867 AK000680

chip HG-U133A HG-U133A HG-U133B HG-U133A HG-U133B HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B

202292x_at 205990_sat 224159xat 221804_sat 209939_xat 212884_xat 208178xat 213164_at 715205641_sat 717210835_sat 719213396_sat 720209018_sat 723219577s_at 206150_at 238063_at 212041at 212944_at 235638at 212588_at 202283_at 203909_at 200675at 206337_at 225626_at ProbeSet 716226599_at 718226430_at 721239629at 722212724at No.ID 714 724 725 726 727 728 729 730 731 732 733 734 735 736 737 738 739 740 741 US 9, 963, 747 B2 103 104

Rank 223 224 224 225 226 226 233 235 249 251 261 262 264 266 268 270272 274 291 302 305 306 311 312 314 315 316 316 ofType speci ficity resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp

Responsemarker + + + + + + + + + + + + + + + + + + + + + + + + + + + + TTP marker

915 199 Entrez Gene ID 9669 2791 5295 2776 6818 51132 79726 23390 3689 ????23392 51177 6275 79686 27071 7415 3702 1462 8933 2530 146346 3507 1534 6480 8932 8780

CKIP-1 Gene Symbol EIF5B GNG11 PIK3R1 GNAQ SULT1A3 RNF12 CD3D FLJ12270 ZDHHC17 ITGB2 KIAA0368 S100A4 C14orf139 DAPP1 VCP ITK AIF1 CSPG2 CXX1 FUT8 LOC146346 IGHM CYB561 STOGAL1 MBD2 RIOK3 1BPredictiveMarkersUpregulatedIndicatorsofResponseand/orLongTimetoProgression continued- 718fucosyltransferase8(alpha1,6) regulatorysubunit1(p85alpha) antigenp95),CD18(integrinbeta2706 710chromosome14openreadingframe139 722ST6betagalactosamide-alpha2,6 699phosphoinositide-3kinase, 701sulfotransferasefamily,cytosolic 1A,phenol-preferringmember3 lymphocytefunction-associatedantigen1;macrophage 708CK2interactingprotein1;HQ0024c 709S100calciumbindingproteinA4(calciumprotein,calvasculin 715allograftinflammatoryfactor1716chondroitin sulfateproteoglycan2 720immunoglobulinheavyconstantmu 723methyl-CpGbindingdomainprotein2 698guaninenucleotidebindingprotein (Gprotein),gamma11 700guaninenucleotidebindingprotein 703CD3Dantigen,deltapolypeptide metastasin,murineplacental 711dualadaptorofphosphotyrosineand fucosyltransferase) sialyltranferase1 724RIOkinase3(yeast) 697eukaryotictranslationinitiation (Gprotein),2polypeptide 702Ringfingerprotein12 (TiT3complex) 705zincfinger,DHHCdomain containing17 (mac-1)betasubunit homolog) 713valosin-containingprotein 714?L2-inducibleTcellkinase 719HypotheticalproteinLOC146346 721cytochromeb-561 Title 5Bfactor 704FLJ12270protein protein 3-phosphoinositides (versican) 717CAAXbox1 NO: 707KIAA0368 SEQ ID 712

RepPublic NM004126 NM000732 NM024673 NM_000211 NM016274 NM002961 NM024633 NM014395 NM003928 NM004480 ID BG261322 A1679268 BF222895 L25275 AW973232 AI621223 BF059292 A1912583 W60953 D13720 AF299327 BF218922 A1742940 BF002659 BC002976 A1743792 AF072242 AK024958

chip HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HGU133A- HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A

201024_xat 204115_at 212240_sat 202615_at 210580_xat 242121at 213539_at 218505_at 212982_at 202803_sat 236368_at 218223_sat 203186sat 219563_at 219290_Xat 214085_xat 208648_at 211339sat 213095xat 221731x_at 201828_xat 203988_sat 225918_at 215949_xat 209164_sat 201998_at 202484sat 215588_xat ProbeSet ID No. 742 743 744 745 746 747 748 749 750 751 752 753 754 755 756 757 758 759 760 761 762 763 764 765 766 767 768 769 US 9, 963, 747 B2 105 106

Rank 317 319 320 326328 333 334 336 337 340 342 342 344 348 349 355 355 356 369 375 377 381 382 398 407 408 410 412 415 ofType specificity resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp ZEEResponse111 marker + + + + + + ++ + + + + + + ++ + + + + + + ++ ++ + + *+ ++ + TTP marker

301 118 !Entrez op Gene ID at399948 9780 til165324 54498 9056 9683 time7412 115352 7474 6687 401105 7466 3790 6256 9459 111157187 57674 8848 7091 1992 23041 4891

Gene Symbol FLJ45803 FAM38A UBXD4 SMOX SLC7A7 N4BP1 VCAM1 ANXA1 FCRH3 WNT5A SPG7 FLJ42393 WFS1 KCNS3 RXRA ARHGEF6 THOC2 C17orf27 TGFB114 TLE4 SERPINB1 KIAA1040 SLC11A2 ADD1 1BPredictiveMarkersUpregulatedIndicatorsofResponseand/orLongTimetoProgression continued- aminoacidtransporter,y+system) transducin-likeenhancerofsplit4 727familywithsequencesimilarity38, 739potassiumvoltage-gatedchannel,delayed-rectifier,subfamilyS GEF)6factor(exchange chromosome17openreadingframe27745 746transforminggrowthfactorbeta1 inhibitor,cladeB(ovalbumin) 730solutecarrierfamily7(cationic 732vascularcelladhesionmolecule1 735wingless-typeMMTVintegrationsite 736spasticparaplegia7,paraplegin 738Wolframsyndrome1(wolframin) 743CDNA:FLJ23566fis,cloneLNG10880 (proton-coupleddivalentmetalionmember2transporters ), 725CDNAFLJ42313fis,clone 728UBXdomaincontaining4 731Nedd4bindingprotein1 734Fcreceptor-likeprotein3 family,member5A (pureandcomplicatedautosomal 740retinoidXreceptor,alpha inducedtranscript4 (Espl)homolog,Drosophila 750serinecysteine(or)proteinase 752solutecarrierfamily11 726FLJ45803protein memberA 729spermineoxidase member7 recessive) 737FLJ42393protein member3 741Rac/Cdc42guaninenucleotide THOcomplex2744 749Transcribedlocus member1 751KIAA1040protein Title TRACH2019425 733annexinA1 753adducin SEQ ID NO: 742 748

RepPublic NM014745 NM003982 NM001078 NM000700 NM005200 NM006005 NM002252 NM002957 NM007005 NM_030757 ID BG288330 BE176566 BG111015 BC000669 AB014515 BF514552 A1968085 AL080232 D25304 A1821721 AK027219 BE543527 AA233374 AK027071 AI703496 A1554300 A1760249 AU154469 AL556041

chip HG-U133B HG-U133B HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HGU133A- HG-U133A HG-U133A HG-U133A HG-U133B HG-U133B HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A

238701_xat 235327_xat 210357_sat 204588_sat 214494_sat 214902_xat 202449s_at 241223_xat 234675xat 225929_sat 215111_sat 208082_xat 213572_sat 212754_sat 203123sat 214726_xat 235028_at 202771_at 32069_at 203868sat 201012_at 231093_at 213425_at 202908_at 205968_at 209539_at 212994at 204872_at 227749_at ProbeSet ID No. 770 771 772 773 774 775 776 777 778 779 780 781 782 783 784 785 786 787 788 789 790 791 792 793794 795 796 797 798 US 9, 963, 747 B2 107 108

Rank 434 437 439 442 445 456 457 461 465 469 472 474 476 478 483 484 497 499 507 511 515 519 523 524 528 529 534 537 540 549 551 552 ofType speci ficity resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp Response marker TTP marker

203 Entrez Gene ID 6509 114782 159090 9813 84981 10723 8837 2804 11067 84166 25957 25957 3500/390714 285231 57730 6277 8621 8165 10013 196515 114876 56987 55720 134147 50619 10457 150759 5199

IGHG1/ Gene Symbol SLC1A4 KIAA1881 LOC159090 KIAA0494 MGC14376 SLC12A7 CFLAR GOLGB1 C10orf10 NOD27 Coorf111 C6orf111 LOC390714 FBXW12 KIAA1641 S100A6 CDC2L5 AKAP1 HDAC6 AK1 FLJ30092 OSBPLIA BBX FLJ10534 LOC134147 DEF6 GPNMB LOC150759 PFC 1BPredictiveMarkersUpregulatedIndicatorsofResponseand/orLongTimetoProgression continued- neutralaminoacidtransporter), 767immunoglobulinheavyconstantgamma1 754solutecarrierfamily1(glutamate/ (potassiumchloridetransporters), 763chromosome10openreadingframe 765chromosome6openreadingframe111766chromosome6openreadingframe111 (Glmmarker)/similartoIgheavy (cholinesterase-relatedcelldivision 777oxysterolbindingprotein-like1A 782differentiallyexpressedinFDCP6 783glycoprotein(transmembrane)nmb 761CASP8andFADD-likeapoptosis 762golgiautoantigen,golginsubfamily chainV-IIIregionVH26precursor 768F-boxandWD40domainprotein12 771S100calciumbindingproteinA6 772Celldivisioncycle2-like5 773Akinase(PRKA)anchorprotein1 778bobbysoxhomolog(Drosophila) 781similartomouse2310016A09Rikgene 759hypotheticalproteinMGC14376 760solutecarrierfamily12 b,macrogolgin(withtransmembrane 764nucleotide-bindingoligomerization 776AF-1specificproteinphosphatase 779hypotheticalproteinFLJ10534 homolog(mouse) LOC150759proteinHypothetical784 785properdinPfactor,complement 757KIAA0494geneproduct 1signal), 774histonedeacetylase6 775adenylatekinase1 member4 756similartohypothetical MGC17347protein member7 regulator domains27 (calcyclin) controller) Title 755KIAA1881 770KLAA1641 SEQ ID NO: 758 769 780

RepPublic NM017932 NM014774 NM006598 NM004487 NM_025190 NM014624 NM006044 NM001915 NM_002510 NM_002621 ID W72527 AI348001 AL080112 AF070569 U97075 AL136653 AA005023 BF591408 AW081113 AB035175 AK022174 AF238870 AK024379 BC000729 BC001116 BC006270 W19983 AF283775 AK026565 BE537881 Z97832 A1983904

chip HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A U133BHG- HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133B HG-U133B HG-U133A HG-U133A HG-U133A

212810_sat 207730_xat 225361_xat 201778_sat 216858xat 214696_at 218066_at 210563x_at 201057s_at 209183_sat 226474_at 225507_at 212177_at 216510_xat 215600_xat 215811_at 217728_at 232266_xat 201674_sat 206846_sat 202587_sat 211034_sat 209485_sat 232008_sat 218155_xat 207986_xat 234981xat 226659_at 220940at 828201141_at 829221973_at 830206380_sat ProbeSet ID No. 799 800801 802 803 804 805 806 807 808 809 810 811 812 813 814 815 816 817 818 819 820 821 822 823 824 825 826 827 US 9, 963, 747 B2 109 110

Rank 554 556 557 561 562 563 565 567 569 574 578 586 591 592 596 603 605 609 612 614 616 619 622 631 633 640 645 648 667 668 675 681 ofType speci ficity resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp Response marker + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + TTP marker

Entrez Gene ID 5228 7940 55256 28639 5576 2810 2207 22839 11152 391733 79890 4782 10146 7084 374618 65125 3492/3500 1043 5927 26137 57714 23039 1043 1513 9324

MTCBP-1 IGH@/ Gene Symbol PGF LST1 TRBC1 PRKARA SFN FCERIG DLGAP4 WDR45 RIN3 NFIC G??? TK2 TEX9 WNK1 IGHG1 CD52 JARIDA ZBTB20 KIAA1618 XPO7 CD52 CTSK HMGN3 1BPredictiveMarkersUpregulatedIndicatorsofResponseand/orLongTimetoProgression continued- metalloproteinasecytoplasmictail 805immunoglobulinheavylocus/ 806CD52antigen(CAMPATH-1) 807Homosapiens,cloneIMAGE:5218355 815CD52antigen(CAMPATH-1) endothelialgrowthfactor-related 791proteinkinase,CAMP-dependent 793FcfragmentofIgE,highaffinityI 798nuclearfactorI/C(CCAAT-binding 804WNKlysinedeficientproteinkinase1 immunoglobulinheavyconstant 786Placentalgrowthfactor,vascular 787leukocytespecifictranscript1 bindingprotein-1 Tcellreceptorbetaconstant1 790CDNA:FLJ20946fis,cloneADSE01819 regulatory,typeIIalpha receptorfor;gammapolypeptide794discs,large(Drosophila)homolog associatedprotein4 transcriptionfactor) 801Ras-GTPaseactivatingproteinSH3 802Thymidinekinase2,mitochondrial gamma1(Glmmarker) 808Jumonji,ATrichinteractivedomain 816cathepsinK(pycnodysostosis) 788membrane-type1matrix 795WDrepeatdomain45796Similar toMicronemeantigen 797RasandRabinteractor3 domain-bindingprotein 803testisexpressedgene9 1A(RBBP2-like)809zincfingerandBTBdomain containing20 817highmobilitygroupnucleosomal bindingdomain3 protein 810Transcribedlocus 813exportin7 Title 792stratifin mRNA 812KIAA1618 SEQ ID NO: 799 800 811 814

RepPublic NM018269 NM004106 NM_005597 NM_017618 NM005056 NM_015024 NM013344 NM_001803 NM_000396 ID AK023843 AF000424 M15564 AK024599 BC002763 X57348 AK024674 AK024315 A1907083 AA625133 AF130054 H09533 X78262 AA020920 A1445745 U80164 N90866 AL117474 AW974823 AI130690 BF056139 AA936632 AF274949

chip HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HGU133A- HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133B HG-U133B HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A

211582x_at 210915_xat 233702xat 204842_xat 233056_xat 215553_xat 222380_sat 206929_sat 211452_xat 239748xat 222187_xat 202040_sat 236715_xat 208459_sat 208238xat 202450_sat 209377_sat 831215179_xat 217761_at 33323rat 204232_at 60471_at 847208246_xat 849211992_at 850217198_xat 34210at 231828_at 222357_at 238668_at 241347_at 204661_at ProbeSet ID 848243198at No. 832 833 834 835 836 837 838 839 840 841 842 843 844 845 846 851 852 853 854 855 856 857 858 859 860 861 862 US 9, 963, 747 B2 111 112

Rank 688 92 156 194 204 207 246 247 250 254 274 111 122 150 132 198 272 350 374 447 482 505 537 581 589 604 657 253 16 37 ofType speci ficity resp TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP/ resp TTP/ resp TTP/ resp resp resp resp resp resp resp resp resp resp resp resp resp resp TTP/ resp resp resp

Response marker + + + + + + + + + + + + + + + + + + + TTP marker + + + + + + + + + + + + + +

567 825 37 330 Entrez Gene ID 7324 81567 64764 283970 4833 5954 26524 2213 6585 4641 1028 79966 55361 55729 7001 5606 4983 55567 5480654806 15022 150271

Gene Symbol UBE2E1 TXNDC5 B2M CREB3L2 LOC283970 NMEA RCN1 LATS2 FCGR2B SLITI CAPN3 MYO1C CDKNIC ACADVL SCD4 PI4KII ATF7IP PRDX2 MAP2K3 OPHN1 DNAH3 AHI1 LOC150271LOC150271 BIRC3 1BPredictiveMarkersUpregulatedIndicatorsofResponseand/orLongTimetoProgression continued- Ubiquitin-conjugatingenzymeE2E1 FcfragmentofIgG,lowaffinityIIb 834phosphatidylinositol4-kinasetypeII baculoviralIAPrepeat-containing3 thioredoxindomaincontaining5 reticulocalbin1,EF-handcalcium 832acyl-CoenzymeAdehydrogenase,very factor7transcriptionactivating 839Dynein,axonemalheavypolypeptide3 840Homosapiens,cloneIMAGE:4829003 842Homosapiens,cloneIMAGE:4822875 (UBC4/5homolog,yeast) CAMPresponsiveelementbinding HypotheticalproteinLOC283970 non-metastaticcells4,protein LATS,largetumorsuppressor homolog2(Drosophila) slithomolog1(Drosophila) 831cyclin-dependentkinaseinhibitor10 hypotheticalproteinLOC150271 protein3-like2 expressedin CD32receptor)( calpain3,(+94) 833stearoyl-CoAdesaturase4 interactingprotein 837Mitogen-activatedproteinkinase 844Abelsonhelperintegrationsite beta-2microglobulin bindingdomain myosinIC (p57,Kip2) longchain 836peroxiredoxin2 kinase3 838oligophrenin1 Title mRNA mRNA SEQ ID NO: 818 819820 821 822 823 824 825 826 827 828 829 830 841 843 845846

PublicRep NM_030810 NM002901 BioC-3 NM000018 NM024906 NM002547 NM025095 ID AU146791 AW188940 AA827878 BG236289 A1925734 AL523860 A1535735 AFFX M31933 AB011537 BC003169 X98507 N33167 AL561930 AK025060 AU147942 AA780381 A1151104 AA601031 AF131777 AL049260 AL136797 R54042 U37546

chip HG-U133A HG-U133A HG-U133A HG-U133B HG-U133B HG-U133A HG-U133A HG-U133A HG-U133B HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133B HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A

221253_sat 216231_sat 223577xat 210889_sat 210944_sat 209345s_at 231825_xat 215067_xat 206323xat 220725x_at 237475_xat 215504_xat 216524_xat 210538_sat 215577_at 228759_at 869212739s_at 201063_at 227013at BioC-3_at 213601_at 32811_at 213348at 200710_at 220232_at 215499_at 228919_at 221569_at 228658_at ProbeSet 868221992_at AFFX No.ID 863 864 865 866 867 870 871 872 873 874 875 876 877 878 879 880 881 882 883 884 885 886 887 888 889 890 891 892 US 9, 963, 747 B2 113 114114

Rank 107 165 196 318 359 421 439 474 475 487 611 151 168 63 30 46 201 329 189 ofTypeinterne speci ficity resp resp resp resp resp resp resp resp resp resp resp TTP TTP TTP TTP/ resp TTP/ resp TTP/ resp resp TTP Response marker + + + + + + + + + + + + + + + + +

TTP marker +

695 Entrez Gene ID 3423 3423 8837 9100 9467 55754 4779 9467 80205 4779 2745293 3459 7082 2745 9958 6509 64115

Gene Symbol IDS IDS CFLAR USP10 SH3BP5 TMEM30A NFE2L1 SH3BP5 CHD9 BTK NFE2L1 GLRX IFNGR1 TJP1 GLRX USP15 SLC1A4 PP2135 1BPredictiveMarkersUpregulatedIndicatorsofResponseand/orLongTimetoProgression continued- chromodomainhelicaseDNAbinding Solutecarrierfamily1(glutamate/ neutralaminoacidtransporter), iduronate2-sulfatase(Hunter iduronate2-sulfatase(Hunter CASP8andFADD-likeapoptosis ubiquitinspecificprotease10SH3-domainbindingprotein5 transmembraneprotein30A nuclearfactor(erythroid-derived SH3-domainbindingprotein5 Brutonagammaglobulinemiatyrosine nuclearfactor(erythroid-derived glutaredoxin(thioltransferase) interferongammareceptor1 tightjunctionprotein1(zona glutaredoxin(thioltransferase) ubiquitinspecificprotease15 occludens1) syndrome) syndrome) (BTK-associated) (BTK-associated) protein9 Transcribedlocus member4PP2135protein Title regulator 2)-like1 kinase 2)-like1 NO: SEQ ID 847 848 849 850 851 852 853 854 855 856 857 858 859 860 861 862 863 864 865

RepPublic NM000202 AV703259 NM004844 NM_000061 NM003204 NM000416 NM_003257 NM002064 ID AF009616 BG390445 AL080250 H93013 AL562152 BF668950 AF162769 AF106069 T99553 BF510711 BE271644

chip HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133B HG-U133B

202439_sat 212221_xat 211316_xat 209136_sat 201811x_at 899214179s_at 201810sat 212616_at 205504_at 200759_xat 209276_sat 202727_sat 202011_at 206662at 209475_at 235661_at 235875_at 225373_at ProbeSet 898222391_at No.ID 893 894 895 896 897 900 901 902 903 904 905 906 907 908 909 910 911 US 9, 963, 747 B2 115 116

lenkeRank wnWN bouou 12 ã 28 36 38 39 48 50 51 52 57 65 66 84 98 104 Typeof specificity resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp respecterResponsemarker LILIT I TILL TIL LITT I TTP marker

2577/ 841 Entrez Gene ID 65220 2579/26748 376267 29793 7317 80336 60559 5831 29793 27101 27101 6789 6678 4830 81873 112770 26010 8826 4638 6731 29922 7074 57136 e 111 100

GAGE5/ GAGE7/ Gene Symbol FLJ13052 GAGE7B RAB15 TI-227H UBE1 C20orf119 SPCS3 CASP8 PYCR1 TI-227H CACYBP CACYBP STK4 SPARC NME1 ARPC5L MGC31963 DNAPTP6 IQGAP1 MYLK SRP72 NMEZ TIAM1 C20orf3 TABLE2 GLUCOCORTICOIDPREDICTIVEMARKERIDENTIFICATION 2APredictiveMarkersUpregulatedIndicatorsofNon-Reponseand/orShortTimetoProgression 881Actinrelatedprotein2/3complex, 882kidneypredominantproteinNCU-G1 889chromosome20openreadingframe3 867Gantigen5/7 869hypotheticalproteinTI-227H sensitivitycomplementing) 872signalpeptidasecomplexsubunit3 875hypotheticalproteinTI-227H 886signalrecognitionparticle72kDa kinase)diphosphatenucleoside-( 868RAB15,memberRASonocogenefamily 870ubiquitin-activatingenzymeEl 871Chromosome20openreading homolog(S.cerevisiae) 873caspase8,apoptosis-related 874pyrroline-5carboxylatereductase1 879secretedprotein,acidic cysteine-rich(osteonectin) 880non-metastaticcells1,protein activatingprotein1 887non-metastaticcells7,protein (A1S9TandBN75temperature 878serine/threoninekinase4 (NM23A)expressedin subunit5-like 883DNApolymerase-transactivated 885myosin,lightpolypeptidekinase 888T-celllymphomainvasionand Gantigen7B frame119 cysteineprotease 876calcyclinbindingprotein877calcyclinbindingprotein protein6 884IQmotifcontainingGTPase expressedin metastasis1 Title 866NADkinase NO: SEQ ID

RepPublic NM021123 NM003334 NM006907 NM_003118 NM_000269 NM003870 NM005965 ID A1334128 AA631242 N37081 AL109839 AL136660 BF439983 A1687738 BC005975 AF057356 BE222274 AU158936 BF977145 AK002064 BE856385 A1094580 U90902 BC000353

chip HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B U133BHG- HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133B HG-U133B HG-U133A HGU133A- HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A

208918_sat 208235_xat 221810_at 212725s_at 200964at 226670_sat 222753_sat 213373_sat 202148_sat 212337_at 211761_sat 201381_xat 225364at 200665s_at 201577_at 226914_at 225401_at 222154_sat 200791_sat 202555_sat 208801_at 227556_at 213135_at 206656_sat ProbeSet ID No. 912 913 914 915 916 917 918 919 920 921 922 923 924 925 926 927 928 929 930 931 932 933 934 935 US 9, 963, 747 B2 117 118

8 vw ??? ? ? ? Rank 22 21 24 28 29 30 ??? ? ? ? ? ? 60 Typeof speci ficity resp1resp respresp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp

Response marker - - TTP marker

2574/ 2576/ 2577/ 2578/ 2576/ 2577/ 2574/ 2576/ 2577/ 2578/ 2579/ 2543/ 2574/ 2575/ 2576/ 2577/ 2578/ 2579/ 135 Entrez Gene ID 2579/26748 2578/126748 26748/26749 26748/26749 2575 7045 84677 5790 126306 283029 23654 389048 23231 55683 440448 8527 27143 23218

GAGE2/ GAGE4/ GAGE5/ GAGE6/ GAGE7/ GAGE4/ GAGE5/ GAGE6/ GAGE2/ GAGE4/ GAGE5/ GAGE6/ GAGE7/ GAGE7B/ GAGE1/ GAGE2/ GAGE3/ GAGE4/ GAGE5/ GAGE6/ GAGE7/ GAGE7B/ Gene Symbol GAGE7B GAGE7B GAGES GAGES GAGE3 ADORAZA TGFBI DSCR8 PTPRCAP FLJ32416 PLXNB2 LOC389048 KIAA0746 FLJ10081 DGKD KIAA1274 KIAA0540 2APredictiveMarkersUpregulatedIndicatorsofNon-Reponseand/orShortTimetoProgression TABLE2-continued GLUCOCORTICOIDPREDICTIVEMARKERIDENTIFICATION antigen4/G21 Gantigen5/6 Gantigen4/5 Gantigen2/4Gantigen5/6 Gantigen7/7B Gantigen1III2/Gantigen3/4 Gantigen5/6 Gantigen7/7B 434Downsyndromecriticalregiongene8 proteinassociatedreceptortype,C- 905diacylglycerolkinase,delta130kDa Gantigen7/7B Gantigen6/7B 896transforminggrowthfactor, proteinsarcoplasmicreticulum beta-induced,68kDa 897proteintyrosinephosphatase, 898homologofmouseskeletalmuscle Gantigen8 Gantigen8 895adenosineA2areceptor 903hypotheticalproteinFLJ10081 894Gantigen3 JP-45 899HypotheticalLOC283029 901hypotheticalLOC389048 902KIAA0746protein 907KIAA0540protein Title 900plexinB2 904LOC440448 906KIAA1274 NO: 2APredictiveMarkersOprea SEQ ID 890 891 892 893 DPublicRep RepPublic NM_001477 NM_001476 NM_001474 NM_001472 NM001473 NM000675 NM_000358 NM_005608 NM003648 ID AA770014 A1805145 AW167298 BC004542 BG285837 AA522514 AF311326 BE563152 AB033100 AB011112

chip HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133B HG-U133B HG-U133A HG-U133B HG-U133A HG-U133B HG-U133B HG-U133A HG-U133B HG-U133A

206640_xat 208155_xat 207086_xat 207739_sat 207663_xat 205013_sat 201506_at 241224xat 204960at 235863_at 228116_at 208890s_at 242881xat 212311_at 224318_sat 224806_at 208072_sat 231887_sat 212443_at ProbesetProbeSet ID No. 936 937 938 939 940 941 942 943 944 945 946 947 948 950 951 952 953 954 US 9, 963, 747 B2 119 120

RankeRank 106 117 119 133 3 16 18 19 25 ? 30 36 42 43 47 48 51 53 60 Typeof speci ficity resp resp resp resp resp resp resp resp resp resp resp resp resp resp TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP

ResponseResponse marker I III III II II I TTP marker IL IT ITIL ITIL

377 301 Entrez Gene ID 2316 3587 5664 55215 54512 283445 4046 23512 9910 23451 25861 55000 6597 286499 23344 1155 5604 3782 203411 51367 55154 10329 7837 10602 4700 2956 80305 8407 4141

oneonGene Symbol FLNA ILIORA PSEN2 FLJ10719 EXOSC4 LOC283445 LSP1 SUZ12 RABGAPIL SF3B1 DFNB31 FLJ20618 SMARCA4 FLJ37659 MBC2 ARF3 CKAP1 MAP2K1 KCNN3 LOC203411 POP5 FLJ10504 TMEM5 D2S448 ANXA1 CDC42EP3 NDUFA6 MSH6 PP2447 TAGLN2 MARS 2APredictiveMarkersUpregulatedIndicatorsofNon-Reponseand/orShortTimetoProgression TABLE2-continued GLUCOCORTICOIDPREDICTIVEMARKERIDENTIFICATION RABGTPaseactivatingprotein1-like NADHdehydrogenaseubiquinone()1 presenilin2(Alzheimerdisease4) lymphocyte-specificprotein1 splicingfactor3b,subunit1 chromatin,subfamilyamember4 channel,subfamilyNmember3 alphasubcomplex,614kDa filaminA,alpha(actinbinding interleukin10receptor,alpha suppressorofzeste12homolog deafness,autosomalrecessive31 SWI/SNFrelated,matrixassociated likelyorthologofmousemembraneboundC2domain containingprotein 925cytoskeletonassociatedprotein1 conductancecalcium-activated 933CDC42effectorprotein(RhoGTPase 935mutShomolog6(E.coli) proteinFLJ10719hypothetical exosomecomponent4 hypotheticalproteinLOC283445 hypotheticalproteinFLJ20618 actindependentregulatorof hypotheticalproteinFLJ37659 ADP-ribosylationfactor3 926mitogen-activatedproteinkinase 927potassiumintermediate/small 929processingofprecursor5, ribonucleaseP/MRPsubunit 931transmembraneprotein5 protein280) (Drosophila) 928hypotheticalproteinLOC203411 932Melanomaassociatedgene binding)3 936hypotheticalproteinPP2447 938methionine-tRNAsynthetase 155kDa kinase1 (S.cerevisiae) 733annexinAl 937transgelin2 Title 930misato SEQ ID NO: 908 909 910 911 912 913 914 915 916 917 918 919 920 921 922 923 924

RepPublic NM_001456 NM001558 NM019037 NM002339 NM014857 012433_NM NM001659 NM003349 NM_002249 NM015918 NM014254 NM_000700 NM002490 NM000179 NM004990 ID U34349 BG403615 A1970898 BF382924 AA813332 AK000749 D26156 A1864183 BC004998 AD001527 AI571419 BF666325 BC002535 D86983 A1801777 BG399562 BC002616

chip HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A

200859_xat 201071_xat 208794_sat 208858_sat 200011_sat 201003_xat 216194_sat 205903_sat 224233_sat 204808s_at 202001_sat 221807_sat 210978_sat 201475_xat ProbeSet 204912_at 211373sat 213008_at 218695_at 43427at 203523_at 212287_at 203020_at 47553_at 222244sat 239481_at 202670_at 226760_at 204839_at 212013at 201012at 225685_at 202911_at No.ID 955 956 957 958 959 960 961 962 963 964 965 966 967 968969 970 971 972 973 974 975 976 977 978 979 980 981 982 983984 985 986 US 9, 963, 747 B2 121121 122

Rank 65 as nie toale 107 110 112 115 116 117 120 124 126 129 136 141 144 145 147 148 151 154 Typeofspeci ficity TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP Response marker TTP marker LLLL LLL TL

Entrez Gene ID 64591/7258 6051 4836 51290 10109 10440 2678 122509 55699 1155 4257 4720 2314 5160 55154 5596829095 4709 2678 116496 26470 6464 7837 54470 2052 63939 116496

TSPY1III Gene Symbol TSPY2 RNPEP NMT1 PTX1 ARPC2 TIMM17A GGT1 FAM14B FLJ10326 CKAP1 MGST1 NDUFS2 FLII PDHA1 FLJ10504 NSFLIC ORMDL2 NDUFB3 GGT1 Clorf24 SEZ6L2 SHC1 D2S448 ARMCX6 EPHX1 C20orf177 Clorf24 2APredictiveMarkersUpregulatedIndicatorsofNon-Reponseand/orShortTimetoProgression TABLE2-continued 951NADHdehydrogenase(ubiquinone)Fe—S GLUCOCORTICOIDPREDICTIVEMARKERIDENTIFICATION membrane17homologA(yeast) 957NADHdehydrogenaseubiquinone()1 944actinrelatedprotein2/3complex, 947familywithsequencesimilarity14, protein2,49kDa(NADH-coenzymeQ 953pyruvatedehydrogenase(lipoamide) betasubcomplex,312kDa 959chromosome1openreadingframe24 containing)transformingprotein1 965chromosome20openreadingframe177 966chromosome1openreadingframe24 939testisspecificprotein,Y-linked 949cytoskeletonassociatedprotein1 952flightlessIhomolog(Drosophila) 955NSFL1(p97)cofactorp47 (aminopeptidaseB) mitochondrialinnertranslocaseof 956ORMl-like2(S.cerevisiae) 963armadillorepeatcontaining, 964epoxidehydrolase1,microsomal 1/testisspecificprotein, kDasubunit2,34 946gamma-glutamyltransferase1 948mitochondrialisoleucinetRNA 958gamma-glutamyltransferase1 960seizurerelated6homolog 961SHC(Srchomology2domain 941N-myristoyltransferase1 memberB S-transferase1 reductase) likemouse2)-( 962Melanomaassociatedgene (xenobiotic) Title Y-linked2940arginylaminopeptidase 942PTX1protein synthetase 950microsomalglutathione alpha1 X-linked6 NO: 954misato SEQ ID 943

RepPublic NM003308 NM020216 NM016570 NM_013421 NM018060 NM001281 NM_020300 NM004550 NM_000284 NM018116 NM_014182 NM002491 NM_022083 NM_012410 NM000120 ID AF020500 AA824298 AF279893 AK023063 AL039706 U80184 AK023585 L20490 A1091079 BF342851 AK000818 AI627538 AF288391

chipchip HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HGU133A- HG-U133B HG-U133A

207918_sat 988208270_sat 989201157_sat 992208679_sat 993215171_sat 994208284_xat 217900_at 998231736_xat 1000212024_xat 1001200980_sat 1002218296_xat 1003232520_sat 1005203371_sat 1006209919_xat 1008218720_xat 1009214853_sat ProbeSet 990218135_at 991222606_at 995230172at 997201804xat 999201966_at 1004218556_at 1007217966sat 1010212012_at 1011214749sat 1012202017_at 1013225313at 1014217967sat No.ID 987 996 US 9, 963, 747 B2 123 124

Rank 161 162 171 180 183 185 186 187 21 22 43 53 70 71 N 13 15 47 Typeofspeci ficity TTP PSTTPEEEEEEE TTP TTP TTP TTP TTP TTP resp resp resp resp resp resp resp resp resp resp resp resp TTP TTP TTP TTP TTP TTP Response marker TTP marker . ILI TITULLITILL IT

100 100 516 Entrez Gene ID 3782 9761 7345 8407 7003 3782 51029 80306 30827 9898 55752 387882 81893 1163 9368 55249 257106 1278 10109 10109 6757 79581 10564

ScanbodGene Symbol KCNN3 KLADIS2KIAA0152 UCHLI TAGLN2 TEAD1 KCNN3 PNAS-4 MED28 ADA CXXC1 UBAP2L ADA SEPT11 LOC387882 LAT1-3TM CKSIB SLC9A3R1 YY1AP1 ATP5G1 LOC257106 COL1A2 ARPC2 ARPC2 SSX2 GPR172A ARFGEF2 2APredictiveMarkersUpregulatedIndicatorsofNon-Reponseand/orShortTimetoProgression TABLE2-continued GLUCOCORTICOIDPREDICTIVEMARKERIDENTIFICATION mitochondrialF0complex,subunito channel,subfamilyNmember3 971TEAdomainfamilymember1(SV40 transcriptionalenhancerfactor) channel,subfamilyNmember3 977ubiquitinassociatedprotein2-like hydrogenexchanger),isoform3 988actinrelatedprotein2/3complex, 989actinrelatedprotein2/3complex, conductancecalcium-activated conductancecalcium-activated transcription,subunit28homolog 976CXXCfinger1(PHDdomain) 983solutecarrierfamily9(sodium/ 985ATPsynthase,Htransporting+ 990synovialsarcoma,Xbreakpoint2 nucleotide-exchangefactor2 smallintermediate/potassium967 972Potassiumintermediate/small 974MediatorofRNApolymeraseII 987collagen,typeIalpha2 991Gprotein-coupledreceptor172A (brefeldinA-inhibited) 969ubiquitincarboxyl-terminal esteraseL1(ubiquitin thiolesterase) 982CDC28proteinkinaseregulatory regulator1 984YY1associatedprotein1 (subunit9),isoform1LOC257106proteinhypothetical 986 subunit2,34kDa subunit2,34kDa 992ADP-ribosylationfactorguanine 973CGI-146protein (yeast) 980hypotheticalprotein 981LAT1-3TMprotein subunit1B Title 968KIAA0152 970transgelin2 975adenosinedeaminase 978adenosinedeaminase 979septin11 NO: 2APredictiveMarkersU SEQ ID

RepPublic NM004181 NM003564 NM_014593 NM_014847 NM000022 NM_001826 NM_004252 NM_018253 AW575123 NM005731 NM003147 NM024531 ID AJ251016 BC000371 A1590088 N47474 AL049397 BE645241 X02189 AL534972 BF969397 AF060511 AL080089 AA788711 BG034239 BF525399

chip HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B U133BHG- HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B

201349_at 226219_at 202403_sat 213513_xat 207988_sat 1015205902_at 1016200616_sat 1017201387_sat 1018200916_at 1019224955_at 1020244040_at 1021212371_at 1022238761_at 1023216705_sat 1024218058_at 1025201377_at 1026204639_at 1027201307_at 1028225105_at 1029209836_xat1030201897_sat 1032217836_sat 1033208972s_at 1038207493_xat1039218151 _xat ProbeSet ID 1040222518at No. 1031 1034 1035 1036 1037 US 9, 963, 747 B2 125 126

Rank 53 108 W O 21 26 31 34 42 45 46 49 56 a 66 75 112 Typeofspeci ficity TTP TTP TTP/ resp TTP/ resp TTP/ resp TTP/ resp TTP/ resp TTP/ resp TTP/ resp TTP/ resp TTP/ resp TTP/ resp TTP/ resp TTP/ resp TTP/ resp TTP/ resp TTP/ resp TTP/ resp TTP/ resp TTP/ resp TTP/ resp TTP/ resp Response marker LLLLL LLL LLL L L 1 TTP marker -

2678/91227 50 Entrez Gene ID 54407 4282 85668566 3514 2678 9557 23603 25926 9784 27440 9744 1368 1155 2678 6731 308 79902 51602

Symbol GGT1/ NOP5/NOP58 Gene SLC38A2SLC38A2 MIF GGTL4 PDXK IGKC GGT1 WhiliCHDIL COROIC DKFZP586L0724 SNX17 CECR5 CENTB1 CPM CKAP1 mulACO2 GGT1 SRP72 ANXA5 PCNT1 2APredictiveMarkersUpregulatedIndicatorsofNon-Reponseand/orShortTimetoProgression TABLE2-continued GLUCOCORTICOIDPREDICTIVEMARKERIDENTIFICATION 995gamma-glutamyltransferase1/glutamyltransferaselike4- gamma 993solutecarrierfamily38,member2 996pyridoxal(pyridoxine,vitaminB6) similartoNP_055301.1neuronal 1001coronin,actinbindingprotein1C 1004cateyesyndromechromosomeregion, factor(glycosylation-inhibiting 997Transcribedlocus,moderately NTPAD7Cprotein-thread 998immunoglobulinkappaconstant gammaglutamyltransferase-1 1000chromodomainhelicaseDNAbinding CarboxypeptidaseM 1007cytoskeletonassociatedprotein1 1011signalrecognitionparticle72kDa 1014nucleolarproteinNOP5/NOP58 994macrophagemigrationinhibitory [Homosapiens] protein1-like 1005centaurin,beta1 1010gamma-glutamyltransferase1 pericentrin1 factor) 1002DKFZP586L0724protein 1003sortingnexin17 candidate5 mitochondrialaconitase10092, Title kinase 1012annexinA5 NO: SEQ ID 999 1006 1008 1013

RepPublic NM_018573 NM002415 NM003681 NM014748 NM017829 NM_014716 NM_014827 NM001098 NM001222 NM001154 NM_024844 ID AI344075 BE883167 X72475 A1422099 BC002342 AU158148 AA902480 BC005969 L20493 AF161469 HG-U133ANM_013430 chip HG-U133A HG-U133A HGU133A- HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HGU133A- HG-U133A HG-U133B

205788_sat 1041218041_xat 1042217871_sat 1043215603_xat 1044202671_sat 243606at 1046216829_at 1047207131_xat 1048212539_at 1049221676_sat 1050221970_sat 1051200991_sat 1052218592_sat 1053205213_at 1054229711_sat 1055211759_xat 1057200793_sat 1058211417_xat 1059208095_sat 1060200782_at 1062223096_at ProbeSet ID 1061218014at No. 1045 1056 US 9, 963, 747 B2 127 128

Rank 27 14 105 13 46 127 177 Typeofspeci ficity resp TTP TTP resp resp resp TTP TTP/ resp

Response marker | | | TTP marker III III II

824 377 Entrez Gene ID presente56884 4257 126731 23231 4726

Gene Symbol FSTL5 MGST1 LOC126731 KIAA0746 NDUFS6 CAPN2 ARF3 2APredictiveMarkersUpregulatedIndicatorsofNon-Reponseand/orShortTimetoProgression TABLE2-continued GLUCOCORTICOIDPREDICTIVEMARKERIDENTIFICATION 1020NADHdehydrogenase(ubiquinone)Fe—S 1016MesenchymalstemcellproteinDSC96 protein6,13kDa(NADH-coenzymeQ 1021calpain2,(m/II)largesubunit 1022ADP-ribosylationfactor3 1015follistatin-like5 1017microsomalglutathione S-transferase1 1019KIAA0746protein reductase) Title 1018LOC126731 NO: SEQ ID

RepPublic NM004553 ID AA129444 AW511319 AI220117 N64686 A1887866 M23254 BG341906

chip HG-U133B HG-U133B HG-U133B HG-U133B HG-U133B HG-U133A HG-U133A HG-U133A

1064227167_sat 1070200734_sat ProbeSet 1063232010_at 1065224918xat 1066225904_at 1067235353at 1068203606at 1069208683_at No.ID US 9, 963, 747 B2 129 130

Rank 222 16 19 22 23 24 24 34 34 35 38 40 42 42 46 46 47 50 56 57 Typeof specificity resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp

Response marker + + + + + + + + + + + + + + + + + + + + + + + + + + + + TTP marker 212 847 Entrez Gene ID 10718 118429 3725 3725 9445 1316 387914 55681 10826 1991 8724 8743 10240 7314 1316 55069 4778 2099 26137 2994 220972 10661

Gene Symbol NRG3 ANTXR2 JUN JUN ITM2B KLF6 LOC387914 SCYL2 ALAS2 C5orf4 CAT ELA2 SNX3 TNFSF10 MRPS31 UBB KLF6 FLJ10099 NFE2 ESR1 ZBTB20 GYPB MARCH8 KLF1

2BPredictiveMarkersUpregulatedIndicatorsofResponseand/orLongTimetoProgression integralmembraneprotein2B aminolevulinate,delta- chromosome5openreading tumornecrosisfactor(ligand) Homosapiens,cloneIMAGE: glycophorinB(includesSs membrane-associatedringfinger anthraxtoxinreceptor2 v-junsarcomavirus17oncogene homolog(avian) V-junsarcomavirus17oncogene homolog(avian) Kruppel-likefactor6 SCY1-like2(S.cerevisiae) synthase2(sideroblastic/ hypochromicanemia) superfamily,member10 Kruppel-likefactor6FLJ10099 Hypotheticalprotein (erythroid-derived2), estrogenreceptor1 zincfingerandBTBdomain Kruppel-likefactor1 neuregulin3 elastase2,neutrophil sortingnexin3 mitochondrialribosomal ubiquitinB 4096273,mRNA containing20 bloodgroup) frame4 proteinS31 nuclearfactor (C3HC4)8 (erythroid) Title WGAR9166 catalase 45kDa NO: SEQ ID 10231024 1025 1026 1027 1028 1029 1030 1031 1032 1033 1034 1035 1036 1037 1038 1039 1040 1041 1042 1043 1044 1045 1046 1047 Hs18SrRNA-5 Hs18SrRNA-5 RepPublic NM_021999 NM_017988 AFFX-12 NM_016348 NM001752 NM001972 NM_005830 M100985 NM_000125 AFFX-r2 NM015642 NM_002100 ID H05240 AU152178 BC002646 BG491844 AB017493 AW664964 NIVI AF130113 AF062483 U57059 X04801 AFFX HUMRGE BE675435 BE962299 L13974 BG250721 AA770170 U65404

chip HG-U133B HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133B HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A

Hs18SrRNA-5_at Hs18SrRNA-5_at 201465_sat 201464_xat 217731_sat 221220_sat 220751_sat 208781_xat 202687_sat M100985at 208960_sat 209930_sat 205383_sat 207459_xat 221824_sat 229233_at 225524_at 208961Sat 230493_at 211560_sat AFFX-12 201432_at 206871at 212603_at 217144_at HUMRGE/ 224688_at 224606_at 205225_at AFFX-12 210504_at ProbeSet ID AFFX No. 1071 1072 1073 1074 1075 1076 1077 1078 1079 1080 1081 1082 1083 1084 1085 1086 1087 1088 1089 1090 1091 1092 1093 1094 1095 1096 1097 1098 US 9, 963, 747 B2 131 132

bu Rank 57 58 69 74 76 88 97 116 10 11 I 16 1818 26 27 Typeof speci ficity resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp respresp resp resp resp resp respresp

Responsemarker | + + + + + + + + + + + + + + + + + + + + + + + + ++ + + + + + TTP marker

286 Entrez Gene ID 51092 2994 130576 3106 2993 80344 4601 50640 53335 3045 8724 2993 7430 3048 9445 10170 3048 3047/3048 55214 221810 10170 283650 10170 9314

HBG1/ Gene Symbol SIDT2 GYPB LOC130576 ANK1 HLA-B GYPA WDR23 MXI1 IPLA2 (GAMMA) BCL11A HBD SNX3 GYPA VIL2 HBG2 ITM2B DHRS9 HBG2 HBG2 LEPREL1 LOC221810 DHRS9 MGC27165 DHRS9 KLF4

2BPredictiveMarkersUpregulatedIndicatorsofResponseand/orLongTimetoProgression dehydrogenase/reductase(SDR hemoglobin,gammaA/ dehydrogenase/reductase(SDR dehydrogenase/reductase(SDR SID1transmembranefamily, glycophorinB(includesSs glycophorinA(includesMN B-cellCLL/lymphoma11A(zinc glycophorinA(includesMN integralmembraneprotein2B Homosapiens,cloneIMAGE: IGrearrangedH-chainmRNA Kruppel-likefactor4(gut) continued- proteinLOC130576hypothetical complex,classIB WDrepeatdomain23 associatedcalcium-independent phospholipaseA2gamma hemoglobin,gammaG hemoglobin,gammaG hemoglobin,gammaG hypotheticalproteinLOC221810 hypotheticalproteinMGC27165 ankyrin1,erythrocytic MAXinteractor1 fingerprotein) hemoglobin,delta sortingnexin3 villin2(ezrin) family)member9 leprecan-like1 family)member9 family)member9 member2 bloodgroup) majorhistocompatibility bloodgroup) intracellularmembrane bloodgroup) 4513167,mRNA Title V-region

NO: SEQ ID 1048 1049 1050 1051 1052 1053 1054 1055 1056 1057 1058 1059 10601061 1062 10631064 1065 1066 1067 10681069 1070 1071 1072 1073 10741075 1076

RepPublic NM_002099 HUMRGE/ M100985 NM_005962 NM_022893 NM_000519 NM_000184 NM_000559 NM018192 NM005771 ID AA150165 AI240545 BE327172 BF060747 A1659683 L42024 AF283773 AFFX BG025248 AB047360 U00179 BF663141 AI133353 AF092128 AF240698 BE881590 AF240697 BF032500 AJ275401 Z27446 AJ275355 BF514079

chip HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B U133BHG- HG-U133B HG-U133B HG-U133A HG-U133A HG-U133A

214407_xat 205389_sat 209140_xat M10098_5at 219497_sat 210648_xat 211820_xat 208621_sat 213515_xat 217732_sat 223952_xat 204419_xat 204848_xat 218717_sat 224009Xat 234419_xat 234390_xat 216542_xat 219799_sat 221841_sat 56256_at 213281at 228360_at 205838_at 1106216389_sat HUMRGE/ 1109223309_xat 206834_at 235278_at 1108202364_at 221911at ProbeSet ID 1107AFFX No. 1099 1100 1101 1102 1103 1104 1105 1110 1111 1112 1113 1114 1115 1116 1117 1118 1119 1120 1121 1122 1123 1124 1125 1126 1127 1128 US 9, 963, 747 B2 133133 134

wNNA Rank 29 30 31 38 44 of aaaaa 54 55 58any 61 74 82 86 91 o 93 94 98 102 Typeof speci ficity resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp

Responsemarker | + + + + + + + + + + + + + + + + + + + + + + + + + TTP marker

669 664 596 Entrez Gene ID 6504 64762 8905 6622 3727 26228 55819 10123 4684 3106 55353 3108 2994/2996 23645 10370 8795 9474 3106/3107 5217 116151 3135 4929

HLA-B/ Gene Symbol SLAMF1 C18orf11 AP1S2 SNCA JUND BRDG1 BPGM RNF130 ARL7 NCAM1 HLA-B LAPTM4B HLA-DMA BNIP3 GYPBIII GYPE PPP1R15A CITED2 TNFRSF10B APG5L HLA-C PFN2 BCL2 C20orf108 HLA-G NR4A2

2BPredictiveMarkersUpregulatedIndicatorsofResponseand/orLongTimetoProgression componentofamyloidprecursor) histocompatibilitycomplex, nuclearreceptorsubfamily4, moleculefamilymember1chromosome18openreading Adaptor-relatedproteincomplex synuclein,alpha(nonA4 BCRdownstreamsignaling1 ADP-ribosylationfactorlike7 neuralcelladhesionmolecule1 alphaclassDMII,complex BCL2/adenovirusE1B19kDa glycophorinB(includesSsbloodgroup)/glycophorinE proteinphosphatase1, transactivator,withGlu/ APG5autophagy5-like complex,classIB/major chromosome20openreading continued- signalinglymphocyticactivation ringfingerprotein130 complex,classIBlysosomalassociatedprotein transmembrane4beta interactingprotein3 regulatory(inhibitor) tumornecrosisfactorreceptor superfamily,member10b GclassI,antigen junDproto-oncogene 2,3-bisphosphoglyceratemutase Cbp/p300-interacting Asp-richcarboxyterminal B-cellCLL/lymphoma2 member2groupA, 1,sigma2subunit majorhistocompatibility majorhistocompatibility subunit15A domain,2 (S.cerevisiae) majorhistocompatibility classI,C profilin2 HLA-Ghistocompatibility Title frame11 frame108

NO: SEQ ID 1077 1078 1079 1080 1081 1082 1083 10841085 1086 1087 1088 1089 1090 1091 1092 1093 1094 1095 1096 1097 10981099 1100 1101

RepPublic NM_003037 NM_022751 NM_012108 NM_001724 NM_018434 NM_018407 NM004052 NM002628 NM_000633 ID AA205444 BG260394 AI762296 AW450363 U63041 D83043 X76775 U05255 U83981 AF109161 AF016266 AK001899 BC004489 AI133137 M90686 A1935096

chip HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A

204466_sat 203751_xat 209968_sat 208729_xat 208029_sat 217478_sat 216833_xat 202511_sat 208812_xat 204992_sat 211530_xat 204621_sat 1129206181_at 219377_at 228415_at 220059_at 203502at 217865_at 202206_at 201849_at 37028_at 209357_at 209295_at 203685_at 224693_at ProbeSetID No. 1130 1131 1132 1133 1134 1135 1136 1137 1138 1139 1140 1141 1142 1143 1144 1145 1146 1147 1148 1149 1150 1151 1152 1153 US 9, 963, 747 B2 135 136

Rank 104 105 108 110 116 117 124 130 135 64 68 69 85 130 131 138 139 146 167 170 174 182 Typeof speci ficity resp resp resp resp resp resp resp resp resp TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP TTP resp TTP TTPTTP TTP TTP

Response marker + + + + + + + + + + + + + + TTP marker + + + + + + + + + + + + + + + + ++ + +

Entrez Gene ID 7314 81618 25842 64798 4929 7453 4929 5611 3133 55030 1974 3133 23171 23365 7320 5589 219402 80344 2788 8087 25482548 51035 85450 8428 2330423304 4217 8555

GeneSymbol UBB ITM2C ASF1A DEPDC6 NR4A2 WARS NR4A2 DNAJC3 HLA-E FBXO34 EIF4A2 HLA-E GPDIL ARHGEF12 UBE2B PRKCSH MTIF3 WDR23 GNG7 FXR1 GAA LOC51035 KLAA1754 STK24 UBR2 MAP3K5 CDC14B

2BPredictiveMarkersUpregulatedIndicatorsofResponseand/orLongTimetoProgression nuclearreceptorsubfamily4, nuclearreceptorsubfamily4, isoform2factor,initiation4A proteinkinaseCsubstrate80K-H glucosidase,alpha;acid(Pompe integralmembraneprotein2C ASF1anti-silencingfunction1cerevisiae) homologA(S. DnaJ(Hsp40)homolog,subfamily dehydrogenase1-like gamma7protein),protein(G ubiquitinproteinligaseE3 componentn-recognin2 CDC14celldivisioncycle14homologB(S.cerevisiae) continued- DEPdomaincontaining6 tryptophanyl-tRNAsynthetase complex,classIE complex,classIE Rhoguaninenucleotideexchange ubiquitin-conjugatingenzyme fragileXmentalretardation, autosomalhomolog1 serine/threoninekinase24 (STE20homolog,yeast) kinase5 2membergroup,A member2groupA, glycerol-3phosphate factor(GEF)12 E2B(RAD6homolog) initiationfactor3WDrepeatdomain23 guaninenucleotidebinding disease,glycogenstorage diseasetypeII) mitogen-activatedprotein ubiquitinB F-boxprotein34 eukaryotictranslation mitochondrialtranslational C,member3 majorhistocompatibility majorhistocompatibility Title ORF KIAA1754 NO: SEQ ID 11021103 1104 1105 1106 1107 11081109 1110 1111 11121113 1114 1115 1116 1117 1118 1119 1120 1121 1122 1123 1124 1125 1126 1127 1128 1129

RepPublic NM_018955 NM_030926 NM_022783 NM_006186 NM_005516 NM_017943 NM001967 NM_025230 NM005087 NM_000152 015853NM_ ID AW665227 AB028628 M61715 S77154 AL119957 M31183 AA135522 AB002380 AA877765 A1815793 BG529919 AL039870 AA425726 AF083420 AB002347 D84476 AU145941

chip HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A

221555_xat 200633_at 221004_sat 229713_at 203428_sat 218858_at 204622_xat 200628_sat 216248_sat 235341_at 200905_xat 218539_at 200912_sat 217456_xat 212510_at 201334_sat 1169202333_sat 214080_xat 223356_sat 201886_at 228831_sat 201637_sat 202812_at 201871_sat 225582_at 208855_sat 212760_at 203836_sat ProbeSet ID No. 1154 1155 1156 1157 1158 1159 1160 1161 1162 1163 1164 1165 1166 1167 1168 1170 1171 1172 1173 1174 1175 1176 1177 1178 1179 1180 1181 US 9, 963, 747 B2 137137 138

Rank 14 23 34 80 33 35 40 83 15 36 43 t 50 59 83 93 107 113 118 28 14 59 Typeof speci ficity TTP/ resp TTP/ resp TTP/ resp TTP/ resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp resp TTP TTP/ resp

Response marker | + + + + + + + + + + + + + + + + + + + + + TTP marker + + + + ++ + +

214 Entrez Gene ID 2104 53335 3725 26100 7468 53335 64167 1176 55788 7128 55507 3135 3135 3106/3107 6925 283131 64744 8850 80853

HLA-B/ WIPI-2 HLA-G HLA-G HLA-C Gene Symbol ESRRG BCL11A JUN WHSC1 BCL11A LRAP AP3S1 C6orf209 TNFAIP3 GPRC5D TCF4 TncRNA LOC64744 ALCAM PCAF KIAA1718

2BPredictiveMarkersUpregulatedIndicatorsofResponseand/orLongTimetoProgression complex,classIB/majorhistocompatibility complex, continued- estrogenrelated-receptorgamma B-cellCLL/lymphoma11A(zinc B-cellCLL/lymphoma11A(zinc adaptor-relatedproteincomplex chromosome6openreading Gprotein-coupledreceptor, familyC,group5memberD hypotheticalproteinAL133206 tumornecrosisfactor, antigen,classIG antigen,classIG factor4transcription trophoblast-derivednoncoding fingerprotein)v-junsarcomavirus17oncogene homolog(avian)WIP149-likeprotein2 Wolf-Hirschhornsyndrome fingerprotein)leukocyte-derivedarginine alpha-inducedprotein3 Activatedleukocytecell p300/CBP-associatedfactor candidate1 3,sigma1subunit frame209 HLA-Ghistocompatibility HLA-Ghistocompatibility majorhistocompatibility classI,C adhesionmolecule KIAA1718protein Title aminopeptidase RNA NO: SEQ ID 1130 1131 1132 1133 1134 1135 1136 1137 1138 1139 1140 1141 1142 1143 1144 1145 1146 1147 1148 1149 1150

RepPublic NM002228 NM_016003 NM_022350 NM_001284 NM_018368 NM_018654 ID AF094518 AF080216 AF083389 A1912275 A1738896 M90684 M80469 M90685 L07950 AK026674 AI042152 AL137764 BF242905 AV727449 BE217882

chip HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133B HG-U133A HG-U133A HG-U133A

209966_xat 210347_sat 201466_sat 204710_sat 209054_sat 222891_sat 219759_at 202442_at 218191_sat 1191202643_sat 221297_at 211529_xat 217436_xat 211528_xat 211911_xat 222146_sat 224566_at 225282_at 201951_at 1201203845_at 1202221778_at ProbeSetID No. 1182 1183 1184 1185 1186 1187 1188 1189 1190 1192 1193 1194 1195 1196 1197 1198 1199 1200 US 9, 963, 747 B2 139139 140

Rank 11 12 13 15 16 17 18 19 19 ?? ?? ?? 24 36 37 39 39 40 41 corticoid Response marker ! ! ! + ++ ++ 11L ++ ' '1 ++ ++ ++ ++ ++ ++ ++ ++ ++ corticoid TTP marker I . proteasome inhibitor Response marker proteasome inhibitor TTP marker

283650/ 682 Entrez Gene ID 4580 3507 388078 3514 51296 57515 1293 23549 3500/3507 54865 122618 6733 10365 22837 5788 3049 10537 7070/94105 1122 6773 9645 26268 7852 10985

IGHG1/ IGHM/ THY1{} Gene Symbol MTX1 IGHM LOC388078 IGKC SLC15A3 TDE2 COL6A3 DNPEP MGC27165 GPATC4 C14orf175 SRPK2 KLF2 COBLL1 PTPRC HBQ1 UBD LOC94105 CHML STAT2 BSG MICAL2 FBXO9 CXCR4 GCN1L1 TABLE3 AGRESSIVENESSPREDICTIVEMARKERIDENTIFICATION

variabledomain(CLL-L3B)/ gamma1(Glmmarker)/ 1177GCN1generalcontrolofamino-acid Immunoglobulinkappaconstant immunoglobulinheavyconstant 1176chemokine(C-Xmotif)receptor4 synthesis1-like(yeast) 1154HRVFab027-VL/ HRVFabN27-VL/ 1165Kruppel-likefactor2(lung) 1171choroideremia-like(Rabescort kDa113transcription2, 1152ImmunoglobulinheavyconstantmuL23518 1153similartoIgheavychainV-IL23516 026-VL/Iglightchaingene 1167proteintyrosinephosphatase, receptortype,C 1170Thy-cellsurfaceantigen/ 1172signaltransducerandactivatorof 1173basigin(OKbloodgroup) regionHG3precursor NM_0165821155solutecarrierfamily15,member3 mu/hypotheticalprotein BE6767031162chromosome14openreadingframe 1163SFRSproteinkinase2 1168hemoglobin,theta1 Thy-1cotranscribed protein2) 1174flavoproteinoxidoreductaseMICAL2 AF1647941156tumordifferentiallyexpressed2 NM_0043691157collagen,typeVIalpha3 AJ2393831160immunoglobulinheavyconstant A19115181161Gpatchdomaincontaining4 1166COBL-like1 1175F-boxprotein9 MGC271 175 1169ubiquitinD NO:Title SEQ ID 1151metaxin1BC001906 1164 NM_0121001158aspartylaminopeptidase AF1035291159 RepPublic NM_005331 NM_006398 ID L14458 AL558479 AU155565 H98105 AL550657 BE965029 AL031178 AF348491 D86973

HG-U133A HG-U133B HG-U133B HG-U133B HG-U133B chip 1215203182_satHG-U133ANM003138 1219207238_satHG-U133ANM002838 1216211643_xatHG-U133AL14457 1217226646_atHG-U133BA1831932 1203210386_satHG-U133A 1204211639xatHG-U133A 1205211637_xatHG-U133A 1207219593_atHG-U133A 1208208671_atHG-U133A1209201438 _atHG-U133A 1210 201937_satHG-U133A 1211216576_xatHG-U133A 1212217281_XatHG-U133A 1218203641SatHG-U133ABF002844 1220220807_atHG-U133A 1221205890_satHG-U133A 1222208850_satHG-U133A 1225208677_satHG-U133A 1226212473_satHG-U133A 1227212987_atHG-U133A 1228211919_satHG-U133A 1229212139_atHG-U133A 1206211644Xat 1213224634_at 1214235802_at 1223226350_at 1224225636_at ProbeSet ID No. US 9, 963, 747 B2 141 142

NN 2. Rank 4242 43 45 46 47 47 48 49 aaaaa 63 64 64 78 79 81 82 o 91 92 gluco corticoid Response marker + + ++ + + ++ + 1 ++ + + + + L + + + + + + gluco- corticoid TTP marker | | proteasomeinhibitor Response marker proteasomeinhibitor TTP marker

966 Entrez Gene ID 6929 6005 7852 8613 9258 66008 9450 2778 23158 10365 10644 7170 4217 25793 6929 2487 91319 83593 2040 5925 51101 665 2271 2171 4635

IMP-2 CGI-62 Gene Symbol TCF3 RHAG CXCR4 PPAP2B MFHAS1 ALS2CR3 LY86 GNAS KIAA0882 KLF2 ???? MAP3K5 FBX07 TCF3 FRZB DERL3 CD59 RASSF5 STOM RB1 BNIPUL FH FABP5 MYL4 TABLE3-continued AGRESSIVENESSPREDICTIVEMARKERIDENTIFICATION

immunoglobulinenhancerbinding 1180chemokine(C-Xmotif)receptor4 1196Rasassociation(RalGDS/AF-6) interactingprotein3-like 1178transcriptionfactor3(E2A 1181phosphatidicacidphosphatasetype (juvenile)chromosomeregion, 1187Kruppel-likefactor2(lung) 1192transcriptionfactor3(E2A immunoglobulinenhancerbinding 1194Der1-likedomainfamily,member3 1195CD59antigenp18-20( antibodies16.3A5,EJ16 1203myosin,lightpolypeptide4 factorsE12/E47) 1179Rhesusbloodgroup-associated amplifiedsequence11183 amyotrophiclateralsclerosis2 1188IGF-IImRNAbindingprotein2 1190mitogen-activatedproteinkinase factorsE12/E47) EJ30,EL32andG344) 1198retinoblastoma1(including 1200BCL2/adenovirusE1B19kDa 1202fattyacidbindingprotein5 1182malignantfibroushistiocytoma candidate3 1184lymphocyteantigen86 1185GNAScomplexlocus kinase5 1193frizzled-relatedprotein identifiedbymonoclonal domainfamily5 osteosarcoma) (psoriasis-associated) alkali;atrial,embryonic glycoprotein 1186KIAA0882protein 1189tropomyosin3 1191F-boxprotein7 1199CGI-62protein 1201fumaratehydratase 2B NO:Title 1197stomatin SEQ ID 1091

RepPublic NM_004271 NM_016270 NM_006548 NM_005923 NM_012179 NM_000321 NM_016010 NM_001444 ID BE962186 U05255AF031549 AJ224869 AA628586 BF739959 AV705253 AA401492 A1348094 BC000771 M31222 BC004270 M81635 AL132665 AA669797 M36172

chip HG-U133AHG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133AHG-U133A 1246203697_atHG-U133AU91903 1247229721_xatHG-U133BA1655697 1248200984_satHG-U133AX16447 1230213730_xatHG-U133A 1233217028_atHG-U133A 1234212226_satHG-U133A 1250201061_SatHG-U133A 1251203132_atHG-U133A 1256210088_xatHG-U133A 1232211254_xat 1236202124_sat 1240219371_sat 1242222976_sat 1245210776_xat 1254214170_xat 1255202345_sat 1231216398_at 1235213457_at 1237205859_at 1238214157_at 1239212956_at 1241218847_at 1243203837_at 1244201178_at 1249223322_at 1252205308_at 1253221478_at ProbeSet ID No. US 9, 963, 747 B2 143143 144

Rank 101 101 102 103 106 114 114 121 122 122 123 137 140 142 149 156 158 159 gluco corticoid Response marker + + gluco- corticoid TTP marker . IL TUTT proteasomeinhibitor Response marker proteasomeinhibitor TTP marker

286 7070/94105 Entrez Gene ID 8683 8644 22883 5431 441347/7110 10312 2547 3192 6281 91768 2534 4635 5034 5216 55154 23029 29920

THY1III Gene Symbol SFRS9 ANK1 AKR1C3 CLSTN1 POLR2B TMF1 TCIRG1 G22P1 HNRPU S100A10 CABLES1 FYN MYL4 P4HB LOC94105 PFN1 FLJ10504 KIAA0117 PYCR2 TABLE3-continued AGRESSIVENESSPREDICTIVEMARKERIDENTIFICATION sequencesimilarity9,memberC dehydrogenase,typeII) calpactinI,ligandannexinII( ofHeLacellsCot25-normalized memberC3(3-alphahydroxysteroid polypeptideB,140kDa H+transporting,lysosomalVO U(scaffoldattachmentfactorA) lightpolypeptide(p11) M143331215FYNoncogenerelatedtoSRC,FGR (proline4-hydroxylase),betadisulfide proteinpolypeptide( isomerase;thyroidhormonebinding Homosapiens(human) AB0185801206aldo-ketoreductasefamily1, NM_0009381208polymerase(RNA)IIDNAdirected familywithSimilarto/1 proteinaisoform3 1214Cdk5andAblenzymesubstrate1A1889160 alkali;atrial,embryonic 2-oxoglutarate4dioxygenase AL1619581218Thy-1cellsurfaceantigen/ member2family, serine-rich9 NM_0060191210T-cell,immuneregulator1ATPase NM_0014691211thyroidautoantigen70kDa(Ku X520051216myosin,lightpolypeptide4 proteinp55) Thy-1cotranscribed AL5618681224pyrrolinecarboxylate-5reductase AL0501361209TATAelementmodulatoryfactor antigen) ribonucleoprotein NM_0029661213S100calciumbindingproteinA10 J027831217procollagen-proline, A13576391219Full-lengthcDNAcloneCSODK008YI09 NM_0037691204splicingfactor,arginine/ YES NO:Title NM_0000371205ankyrin1,erythrocytic BC0036211212heterogeneousnuclear SEQ ID AA8874801223KIAA0117protein NM_0149441207calsyntenin1 NM_0050221221profilin1 AL5735911222misato RepPublic NM_0068421220 ID

chip HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U1334 HG-U133BHG-U133A HG-U133B 1265200593_satHG-U133A 1267225532_atHG-U133B 1268210105_satHG-U133A 1269217274_xatHG-U1334 1258205390_sat 1260201561_sat 1262214948_sat 1263204158_sat 200872_at 1271208851_sat 1257200044_at 1259209160_at 1261201803_at 1264200792_at 1270200654_at 1272225716_at 1273200619_at1274 200634_at 1275222584_at 1276212591_at 1277224855_at ProbeSet ID No. 1266 US 9, 963, 747 B2 145 146

Rank 168 175 178 6 10 11 22 24 27 33 40 41 41 44 74 81 93 103 114 125 128 137 141 gluco corticoid Response marker i II IIlll II . ii . gluco- corticoid TTP marker in willTL proteasomeinhibitor Response marker proteasomeinhibitor TTP marker IL LITTL L LL LLLLLLLLL

Entrez Gene ID 6391 51514 2224 51463 5591 51463 2339 92703 55585 51463 3065 51463 79022 8683 4085 128077 10051 51497 9262 54928 10603 3020 7514 9837 84232 150275 149951 5710 6635

Gene Symbol SDHC RAMP FDPS GPR89PRKDC GPR89 FNTA Clorf37 UBE2Q GPR89 HDAC1 GPR89 MGC5576 SFRS9 MAD2L1 LX1L SMC4L1 THIL STK17B IMPA3 APS ????? XPO1 KIAA0186 MAF1 FLJ33814 COMMD7 PSMD4 SNRPE TABLE3-continued AGRESSIVENESSPREDICTIVEMARKERIDENTIFICATION

398farnesyltransferase,CAAXbox 1232chromosome1openreadingframe37 chromosomes4-like1(yeast) 1242Serine/threoninekinase17b(apoptosis- 1246exportin1(CRM1homolog,yeast) 1251proteasome(prosome,macropain)26S 1225succinatedehydrogenasecomplex, (farnesylpyrophosphatesynthetase,dimethylallyltranstransferase, 1230proteinkinase,DNA-activated 1238MAD2mitoticarrestdeficient-like 1244adaptorproteinwithpleckstrinhomology subunitC,integralmembrane matrix-associatedprotein geranyltranstransferase)1229Gprotein-coupledreceptor89 1231Gprotein-coupledreceptor89 1233ubiquitin-conjugatingenzymeE2Q 1234Gprotein-coupledreceptor89 1236Gprotein-coupledreceptor89 1204splicingfactor,arginine/ 1239Lix1homolog(mouse)like 1240SMC4structuralmaintenanceof 1243myo-inositolmonophosphataseA3 andsrchomology2domains 1250COMMdomaincontaining7 subunit,non-ATPase4 protein,15kDa 1228farnesyldiphosphatesynthase catalyticpolypeptide 1235histonedeacetylase1 1237hypotheticalproteinMGC5576 serine-rich9 1241TH1-like(Drosophila) inducing) 1245H3histone,family3A 1247KIAA0186geneproduct 1248homologofyeastMAF1 1249hypotheticalproteinFLJ33814 polypeptideE 1226RA-regulatednuclear alpha (putative) 1(yeast) 1252smallnuclearribonucleoprotein NO:Title SEQ ID 1227

RepPublic NM_003001 NM_016448 NM_002004 NM_017582 NM_016334 NM_004964 NM_024056 NM_003769 NM_002358 AW500180 NM_016397 NM_020979 NM_021067 NM_003094 IDID D49737 AF151035 U47077 BF941168 BG168896 AF070537 AK021758 AL136877 N51102 A1302253 A1955655A1955655 D89729 AL136937 BF060776 AA148301 AB033605

chip HG-U133A HG-U133A HG-U133B HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133B HG-U133A HG-U133A 1280210131_XatHG-U133A 1281201275_atHG-U133A 1288220642_xatHG-U133A 1289201209_atHG-U133A 1290222140_satHG-U133A 1291201764_atHG-U133A1292201698_satHG-U133A 1296220607_xatHG-U133A 1297226525_atHG-U133B 1300200080_satHG-U133B 1301208775atHG-U133A 1302206102_atHG-U133A HG-U133B1304225644at HG-U133B1305224815at 1306210460_satHG-U133A 1278202004_xat 1279218585_sat 1282223531_xat 1284225463_xat 1287217978_sat 1293203362_sat 1307203316_sat 1283208694_at 1285200090_at 1286212165_at 1294225793_at 1295201664_at 1298222654_at 1299205367_at ProbeSet ID 1303222998at No. US 9, 963, 747 B2 147 148

Rank 144 154 157 157 158 163 169 186 195 205 214 216 225 236 243 281 288 293 119 171 10 12 14 18 23 24 26 33 41 63 108 gluco corticoid Response marker I gluco- corticoid TTP marker , proteasomeinhibitor Response marker TILL I TIL proteasomeinhibitor TTP marker .. . LLL LLL L LITTL + + 471 Entrez Gene ID 55765 23215 3020 9939 3028 5932 29920 51433 8683 3020 5710 10051 81875 55147 1213 2058 2280 64764 192683 6756 55905 8428 64943 7458 6757 6187 1652 54927 6305 25864 51282

Gene Symbol FLJ10901 BAT2D1 ????? RBM8A HADH2 RBBP8 PYCR2 ANAPC5 SFRSO ????? SMC4L1 FLJ12671 RBM23 CLTC EPRS ATIC SCAMP5 SSX1 ZNF313 STK24 FLJ12442 SSX2 RPS2 DDT CHCHD3 SBF1 DKFZP5640243 SCAND1 TABLE3-continued AGRESSIVENESSPREDICTIVEMARKERIDENTIFICATION

chromosomes4-like1(yeast) ribonucleotideformyltransferase/ dehydrogenase,typeII subunit,non-ATPase4 family,member2 protein3-like2 homolog,yeast) domaincontaining3 5subunit serine-rich9 IMPcyclohydrolase 5protein region1 NO:Title SEQ ID 1324200709_atHG-U133ANM0008011267FK506bindingprotein1A,12kDaFKBP1A 12685-aminoimidazole4carboxamide1325208758atHGU133AD89976 1336217972_atHG-U133ANM0178121279coiledcoilhelix 1318211609_xatHG-U133AU510071261proteasome(prosome,macropain)26SPSMD4 1322200614_atHG-U133ANM0048591265clathrin,heavypolypeptide(HC) 1271synovialsarcoma,Xbreakpoint11328206626_xatHG-U133ABC001003 1330208854_satHG-U133AAA5867741273serine/threoninekinase24(STE20 1283SCANdomaincontaining11340232652XatHG-U133BAF207829 RepPublic 1314231715_satHG-U133BNM0133281258pyrroline5carboxylatereductase 1323200843_satHG-U133ANM0044461266glutamylprolyltRNAsynthetase CREB3L21326212345_satHG-U133ABE6751391269CAMPresponsiveelementbinding 1332206621_satHG-U133ANM0221701275WilliamsBeurensyndromechromosomeWBSCR1 1333216471_xatHG-U133AX792001276synovialsarcoma,Xbreakpoint2 IDID 1312202282_atHG-U133ANM0044931256hydroxyacylCoenzymeA 1321219816_satHG-U133ANM0181071264RNAbindingmotifprotein23 1308219010_atHG-U133ANM0182651253hypotheticalproteinFLJ10901 1309211946_satHG-U133AAL0968571254BAT2domaincontaining1 1310200080_satHG-U133AA19556551245H3histone,family3A 1311222443_satHG-U133BAF1824151255RNAbindingmotifprotein8A 1313203344_satHG-U133ANM0028941257retinoblastomabindingprotein8 1316200044_atHG-U133BNM0037691204splicingfactor,arginine 1317208755_xatHG-U133ABF3123311260H3histone,family3A 1319201663_satHG-U133ANM0054961262SMC4structuralmaintenanceof 1272zincfingerprotein3131329211678_satHG-U133AAF090934 133739835_atHG-U133AU931811280SETbindingfactor1 1282DKFZP5640243protein1339210006atHG-U133ABC002571 1315211036_xatHG-U133ABC0063011259anaphasepromotingcomplex 1320212766_satHG-U133AAW2945871263hypotheticalproteinFLJ12671 1327212699_atHG-U133ABE2228011270secretorycarriermembrane 1331218051_satHG-U133ANM0229081274hypotheticalproteinFLJ12442 1334212433_xatHG-U133AAA6303141277ribosomalproteinS2 1335202929_satHG-U133ANM0013551278Ddopachrometautomerase chipchip 1338213166XatHG-U133ABG3324621281

ProbeSet ID No. US 9, 963, 747 B2 149 150

Rank 144165 202 212 225 242 253 257 285 351 361 386 415 448 467 487 19 28 61 99 113 129 171 178 2 gluco corticoid Response marker gluco- corticoid TTP marker | |

proteasomeinhibitor Response markercle" IIIIIIIIIIIIIIII III II I I 1 + proteasomeinhibitor TTP marker LLL LLL L L I

52 226 2023/6636 Entrez Gene ID 57128 6746 131118 5955 54480 389541 10589 2132 26135 2098 57222 11124 81704 55013 65005 8733 54460 1965 7203 5202 7818 4841 5692 8634 1154

CSGICA-T ENO1/ Gene Symbol ACP1 ALDOA Coorf149 SSR2 TIM14 RCN2 LOC389541 DRAP1 EXT2 PAI-RBP1 ESD KIAA1181 FAF1 DOCKS FLJ20647 MRPL9 GPAA1 MRPS21 EIF281 CCT3 PFDN2 SNRPF DAP3 NONO PSMB4 RTCD1 CISH TABLE3-continued AGRESSIVENESSPREDICTIVEMARKERIDENTIFICATION

(translocon-associatedproteinbeta) intermediatecompartment32kDa factor2,subunit1alpha35kDa ribonucleoproteinpolypeptideF subunit,betatype4 cofactor2alpha) homolog(yeast) ma) bindingdomain glucuronyltransferase hydrolase protein 3(gamma) binding domain1 protein NO:Title SEQ ID 1363203832_atHG-U133ANM0030951306enolase1,(alpha)/smallnuclear HG-U133ANM_0029021289reticulocalbin2,EFhandcalcium1346201486at 1349203258_atHG-U1334NM0064421292DR1associatedprotein1(negative 1361200910_atHG-U1334NM0059981304chaperonincontainingTCP1,subunit 1365200057_satHG-U133ANM007363201nonPOUdomaincontaining,octamer 1366202244_atHG-U133ANM0027961308proteasome(prosome,macropain) RepPublic 1342200966_xatHG-U133ANM0000341285aldolaseA,fructosebisphosphate 1343218561_satHG-U133ANM0204081286chromosome6openreadingframe149 1344200652_atHG-U133ANM0031451287signalsequencereceptor,beta 1294PAI-1mRNAbindingprotein1351209669_satHGU133ABC003049 1354224217_satHG-U133BAF0947001297Fas(TNFRSF6)associatedfactor1 1358211060_xatHG-U133ABC0063831301GPAA1Panchorattachmentprotein1 ID 1341201630_satHG-U133ANM0043001284acidphosphatase1,soluble 1348224890_satHG-U133BBE7276431291similartoCG14977PA 1295esteraseD/formylglutathione1352215096_satHG-U133AAU145746 1353224576_atHG-U133BAK0007521296endoplasmicreticulumgolgi 1367203594_atHG-U133ANM0037291309RNAterminalphosphatecyclase 1369223377_xatHG-U133BAF0359471311cytokineinducibleSH2containing 1345225359_atHG-U133BBF6669611288homologofyeastTIM14 1350202012_satHG-U133AAA1962451293exostoses(multiple)2 1355225502_atHG-U133BAL1617251298dedicatorofcytokinesis8 1356218802_atHG-U133ANM0179181299hypotheticalproteinFLJ20647 1357209609_satHG-U133ABC0045171300mitochondrialribosomalproteinL9 1359222997_satHG-U133BBC0045661302mitochondrialribosomalproteinS21 1364208822_satHG-U133AU183211307deathassociatedprotein3 134755093_atHG-U133AAA5341981290chondroitinsulfate 1360201144_satHG-U133ANM0040941303eukaryotictranslationinitiation HG-U133ANM_0123941305prefoldin21362218336at chip 1368215450_atHG-U133AW879011310

ProbeSet ID No. US 9, 963, 747 B2 151 152

Rank a 26 31 39 41 93 115 121 142 143 144 158 167 170 176 182 192 197 199199 205 222 233 246 gluco corticoid Response marker *+ * **+ gluco- corticoid TTP marker proteasomeinhibitor Response marker + + + + + + + + + + + + + + + + + + + + + + + + + + + + + proteasomeinhibitor TTP marker

6983445347/ 3500/3507 953 996 706 Entrez Gene ID 3932 7280 7111 51176 6282 4064 2335 5934 8837 2752 10475 10125 84669 6955 23468 23094 6095 7168 80008 9958 3500 23338 o

TRGV9/ IGHM/ Gene Symbol TARP IGHG1 LCK TUBB2 TMOD1 LEF1 S100A11 LY64 FN1 RBL2 CFLAR GLUL TRIM38RASGRP1 USP32 TRAC ENTPD1 CRCBX5 SIPA1L3 RORA TPM1 CDC27 FLJ23235 USP15 IGHG1 BZRP PHF15 TABLE3-continued AGRESSIVENESSPREDICTIVEMARKERIDENTIFICATION marker)gammaGlmconstant1( radioprotective105kDa(mouse) 1337CloneA9A2BRB5(CAC)n/GTG 1339benzodiazapinereceptor(peripheral) 1317lymphoidenhancer-bindingfactor1 1319lymphocyteantigen64homolog, 1321retinoblastoma-like2(p. 1323glutamate-ammonialigase(glutamine (calciumandDAG-regulated) 1329Chromoboxhomolog5(HP1alpha 1318S100calciumbindingproteinA11 1324tripartitemotif-containing38 1325RASguanylreleasingprotein1 1326ubiquitinspecificprotease32 diphosphohydrolase1 homolog,Drosophila) associated1like3 1331RAR-relatedorphanreceptorA 1332Tropomyosin1(alpha) 1335ubiquitinspecificprotease15 repeat-containingmRNA. gamma1(Glmmarker) 1312Tcellreceptorgammavariable 9/TCRgammaalternatereading 1313Immunoglobulinheavyconstant mu/Immunoglobulinheavy 1314lymphocyte-specificprotein 1322CASP8andFADD-likeapoptosis 1327Tcellreceptoralphalocus 1333celldivisioncycle271334hypotheticalproteinFLJ23235 constantheavyimmunoglobulin1338 frameprotein tyrosinekinase 1315tubulin,beta2 1316tropomodulin1 (calgizzarin) 1320fibronectin1 regulator synthase) 1328ectonucleosidetriphosphate 1330signal-inducedproliferation 1340PHDfingerprotein15 NO:Title SEQ ID 1336

RepPublic NM_005356 NM_001069 NM_005620 NM_005582 NM_005739 NM_000714 ID M16768 U80139 BC002660 AF288571 U08626 AU157590 A1148567 M15565 AV717590 AA181060 AB011117 U92706 A1735639

chip HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A 1392220169_atHG-U133ANM024943 HG-U133A HG-U133A HG-U133A 1378211719_xatHG-U133ABC005858 1380214486_xatHG-U133AAF041459 1389210426_xatHG-U133AU04897 1390210987_xatHG-U133AM19267 1394213327SatHG-U133AAI820101 1395211994_atHG-U133AAI742553 1370209813_xatHG-U133A 1371216491_xatHG-U133A 1373204141_atHG-U133A 1374203661_satHG-U133A 1375221558_satHG-U133A 1376200660_atHG-U133A 1379212332atHG-U133ABF110947 1384226505_xatHG-U133B 1385210972_xatHG-U133A 1386209473_atHG-U133A 1391217878SatHG-U133AAI203880 1393210681satHG-U133AAF153604 1372204891_sat 1381217202_sat 1382203567_sat 1396216557_xat 1397202096_sat 1377206206_at 1383205590_at 1387226085_at 138837831_at 1398212660_at ProbeSet ID No. US 9, 963, 747 B2 153 154

Rank 249 273 281 286 299 306 309 315 329 329 333 340 357 368 377 382 406 433 522 525 535 547 627 664 686 gluco corticoid Response marker gluco- corticoid TTP marker proteasomeinhibitor Response marker +- *+ *+ *+ *+ *+ ++ + ++ ++ * * * * * * * * * * * * * * - proteasomeinhibitor TTP marker

341 994 199 195 645 Entrez Gene ID 79567 9473 65125 283666 6338 2634 1645 9848 9935 6095 7756 7184 28232 5886 10000 6583 6039 1488 91663 6675

Gene Symbol FLJ13725 Clorf38 WNK1 LOC283666list SCNN1B GBP2 AKR1C1 MFAP3Lin MAFB RORA ZNF207 TRA1 APOC1 SLCO3A1 CDC25B RAD23A ???3 AIF1 SLC22A4 RNASE6 AHNAK CTBP2 BLVRB MYADM UAP1 TABLE3-continued AGRESSIVENESSPREDICTIVEMARKERIDENTIFICATION homolog3(proteinkinaseB,gamma) dehydrogenase1;20-alpha fibrosarcomaoncogenehomologB transporterfamily,member3A1 cationtransporter),member4 ofPlacentaCot25-normalized Homosapiens(human) AK0258621353tumorrejectionantigen(gp96)1 BF5729381357RAD23homologA(S.cerevisiae) reductase(NADPH) NM_0003361346sodiumchannel,nonvoltage-gated1 beta(Liddlesyndrome) memberC1(dihydrodiol 1358v-aktmurinethymomaviraloncogene BG2878621362AHNAKnucleoprotein(desmoyokin) pyrophosphorylase1 NM_0048481343chromosome1openreadingframe38 interferon-inducible (3-alpha)hydroxysteroid dehydrogenase) L146111351RAR-relatedorphanreceptorA 1359allograftinflammatoryfactor1U19713 NM_0056151361ribonuclease,RNaseAfamilyk6 BG4328871341Full-lengthcDNAcloneCSODIO20Y119 A17685121344WNKlysinedeficientprotein kinase1 NM_0041201347guanylatebindingprotein2, NM_0013531348aldo-ketoreductasefamily1, (avian) NM_0030591360solutecarrierfamily22(organic NM0007131364biliverdinreductaseB(flavin AA4002061342hypotheticalproteinFLJ13725 AW0061851345hypotheticalproteinLOC283666 3-like A11256461352Zincfingerprotein207 NM_0013291363C-terminalbindingprotein2 AA9090441365myeloid-associateddifferentiation 1366UDP-NacteylglucosamineS73498 NO:Title NM_0216471349Microfibrillar-associatedprotein NM_0016451354apolipoproteinC-I NM_0132721355solutecarrierorganicanion NM_0218731356celldivisioncycle25B marker SEQ ID NM_0054611350v-mafmusculoaponeurotic RepPublic ID N32526

chip HG-U133B HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133B HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U133A HG-U1334 HG-U133A HG-U133B HG-U133A 1401207571_xatHG-U133A 1417209901XatHG-U133A 140045749_atHG-U133A 1402211993_atHG-U133A 204151_xat 1418205896_atHG-U133A 1408218559_sat 1409210479_sat 1411216449_xat 1412204416_xat 1414201853_sat 1415201039_sat 1421201220_xat 1423224920_xat 209340_at 1399243699_at 1403226682_at 1404205464_at 1405202748_at 1407205442_at 1410228157_at 1413219229_at 1416212607_at 1419213566_at 1420211986_at 1422202201at ProbeSet ID No. 1406 1424 US 9, 963, 747 B2 155155 156

Rank 38 15 177200 289 2 104 135 14 32 4545 343 364 32 35 36 89 184 35 100 gluco corticoid Response marker III + I gluco- corticoid TTP marker + | proteasomeinhibitor Response marker TILL L + + +

proteasomeinhibitor TTP marker | | | | I L I

860 831 831 894 Entrez Gene ID 7520 9582 4113 4105 9503 2826 84527 22919 6757 57050 6187 2731 26051 5269 57610 22837 7167 3107 51514 1786 1029 8733 7167

Gene Symbol XRCC5 ???????? MAGEB2 MAGEA6 RUNX2 XAGE1 CCR10 ZNF559 MAPRE1 SSX2 SAS10 RPS2 GLDC PPP1R16B SERPINB6 RANBP10 COBLL1 CAST CAST TPI1 CCND2 HLA-C RAMP DNMT1 CDKN2A GPAA1 TPI1 TABLE3-continued AGRESSIVENESSPREDICTIVEMARKERIDENTIFICATION rejoining;Kuautoantigen,80kDa) defectiverepairinChinesehamster polypeptide-likecatalyticenzyme, inhibitor,cladeB(ovalbumin) (melanoma,p16inhibitsCDK4) cells5(double-strandbreak decarboxylase,glycinecleavage RP/EBfamily,member1 (decarboxylating;glycine systemproteinP) (inhibitor)subunit16B matrix-associatedprotein homolog(yeast) member6 classI,C transferase1 ??3B NO:Title SEQ ID 1431220565_atHG-U1334NM0166021373chemokine(Cmotif)receptor10 1427206218_atHG-U133ANM0023641369melanomaantigenfamilyB,2 1439225239_atHG-U133BA13554411380CDNAFLJ26120fis,cloneSYN00419 1428214612_xatHG-U133AU106911370melanomaantigenfamilyA,6 1429232231_atHG-U133BAL3539441371Runtrelatedtranscriptionfactor2 1430220057_atHG-U133ANM0204111372Xantigenfamily,member1 1450201697_satHG-U133ANM0013791391DNA(cytosine5)methyl RepPublic 1433200713_satHG-U133ANM0123251375microtubuleassociatedprotein, 1434210497_xatHG-U133ABC0028181376synovialsarcoma,Xbreakpoint2 1438212750_atHG-U133AAB020630472proteinphosphatase1,regulatory 1440211474_satHG-U133ABC0049481381serine(orcysteine)proteinase 1451209644_xatHG-U133AU389451392cyclindependentkinaseinhibitor2A 1452215690_xatHG-U133AAL1574371393GPAA1Panchorattachmentprotein1 IDID 1426206632_satHG-U133ANM0049001368apolipoproteinBmRNAediting 1435209486_atHG-U133ABC0045461377disrupterofsilencing10 144153987_atHG-U133AAL0418521382RANbindingprotein10 1447216526_xatHG-U133AAK0248361388histocompatibilitymajorcomplex, 1425208642_satHG-U133AAA2058341367Xrayrepaircomplementing 1432224518_satHG-U133BBC0064361374zincfingerprotein559 1378ribosomalproteinS21436217466_xatHG-U133AL48784 1437204836_atHG-U1334NM0001701379glycinedehydrogenase 1442203642_satHG-U133ANM0149001383COBLlike1 1445213011_satHG-U133ABF116254triosephosphate1386isomerase1 1449222680_satHG-U133BAK0012611390RAregulatednuclear 1453200822_xatHG-U1334NM0003651394triosephosphateisomerase1 1444207467_xatHG-U133ANM0017501385calpastatin 1446200953_satHG-U133ANM0017591387cyclinD2 chip 1443208908_satHG-U133AAF3274431384calpastatin 1448201952atHG-U133AAA1567211389

ProbeSet ID No. US 9, 963, 747 B2 157 158

Rank 287 43 158 339 460 12 39 16 95 52 78 111 149 234 254 354 32 37 25 gluco corticoid Response marker + + + + + gluco- corticoid TTP marker + proteasomeinhibitor Response marker LILITII + + + + + + + +

proteasomeinhibitor TTP marker 1 + IIII

Entrez Gene ID 3020 51696 51205 26986/5042 1794 6929 8733 8836 84320 2752 54537 1193 5796 8837 8837 8837 2057 57162 57162 57162 140465

PABPC3/ Gene Symbol ???? HECA ACP6 PABPC1 DOCK2 TCF3 GPAA1 GGH ACBD6 GLUL FAM35A CLIC2 PTPRK CFLAR CFLAR CFLAR EPOR PELI1 PELI1 PELI1 MLCISA TABLE3-continued AGRESSIVENESSPREDICTIVEMARKERIDENTIFICATION

cytoplasmic3/poly(A)binding immunoglobulinenhancerbinding protein,cytoplasmic1 factorsE12/E47) homolog(yeast) (conjugase,folylpolygammaglutamyl receptortype,K hydrolase) containing6 synthase) memberA regulator regulator regulator NO:Title SEQID HG-U133BAU1475061412Pellinohomolog1(Drosophila)1471232213_at HG-U133ANM_0024751415myosinlightchain1slowa1474204173at 1398poly(bindingA)protein,1457215823_xatHG-U133AU64661 1462225317_atHG-U133BAL5746691403acylCoenzymeAbindingdomain HG-U133AA17686281406chlorideintracellularchannel21465213415at 1467210564_atHG-U133AAF0096191408CASP8andFADDlikeapoptosis HG-U133BAK0267141413Pellinohomolog1(Drosophila)1472232304_at 1473218319_atHG-U133ANM0206511414pellinohomolog1(Drosophila) RepPublic 1455218603_atHG-U133ANM0162171396headcasehomolog(Drosophila) 1456218795_atHG-U133ANM0163611397acidphosphatase6,lysophosphatidic 1399dedicatorofcytokinesis21458213160_atHG-U133AD86964 1460201618_xatHG-U133ANM0038011401GPAA1Panchorattachmentprotein1 1463215001_satHG-U133AAL1619521404glutamateammonialigase(glutamine 1464220547_satHG-U133ANM0190541405familywithsequencesimilarity35, 1466203038_atHG-U133ANM0028441407proteintyrosinephosphatase, 1468209508_xatHG-U133AAF0057741409CASP8andFADDlikeapoptosis 1469208485_xatHG-U133ANM0038791410CASP8andFADDlikeapoptosis ID 1454213828_xatHG-U133AAA4776551395H3histone,family3A 1459213811_XatHG-U133AAW0623411400transcriptionfactor3(E2A 1461203560_atHG-U133ANM0038781402gammaglutamylhydrolase 147037986_atHG-U133AM604591411erythropoietinreceptor chip

ProbeSet ID No. US 9 , 963, 747 B2 159 160 Classification Methods The k -nearest neighbor classification method computes Various algorithms are currently available that can be the similarity between a query profile and each of the used to classify patient samples using a given set of features . profiles in the training set [Introduction to Machine Learning Therefore , the combination of markers selected through the feature selection process may be used in any of the available 5 by0 2 Ethem ALPAYDIN , The MIT Press, October 2004 , ISBN algorithms in order to derive a prediction equation as to 5 0 - 262- 01211 - 1 ] . The k most similar profiles are selected , whether the patient sample is sensitive or resistant. The and a vote is taken amongst their class labels to determine classification methods used to illustrate the use of multiple the prediction for the query profile . Here , we used k = 1 . markers for patient sample clasdification in the present Feature Selection invention were : 1) Linear Predictive Score (“ LPS ” ); and 2 ) 10 Feature selection is the process of determining a subset of k -nearest neighbors . 10 the thousands of available features in the dataset , resulting The Linear Predictive Score was implemented as in a combination of features that form a marker set or model , described by Wright et al ., “ A gene -expression based to classify patients by treatment outcome. There are many method to diagnose clinically distinct groups of diffuse large approaches to selecting features. Here we report two B cell lymphoma. " PNAS 100 ( 17 ) :9991 - 9996 ( 2003 ) , the 16 approaches to generate example marker sets : ( 1 ) top N most significant features , and ( 2 ) a standard feature selection contents of which are incorporated herein by reference. As method , sequential forward feature selection ( See , Dash and described by Wright et al. , the LPS score for a vector X is Liu , “ Feature Selection for Classification , " Intelligent Data computed as: Analysis 1 : 131 - 156 , 1997 ) . We now describe how feature selection is applied to our dataset . 20 As a first step , only features associated with the outcome LPS( X ) = a; X ; variable are considered as candidates for a feature set. For the LPS models models , all features with multiple - test adjusted p - values less than 0 . 05 were determined . For the where X , represents the log expression value for the jth k - nearest- neighbor models , the top 100 PFC markers were feature in the set , and a , is a scaling factor representing the 25 determined In either case, sequential forward selection starts degree to which the jth feature is associated with the outcome with no markers in the set. At each iteration , a new feature to be predicted . As in Wright et al. , we used the t - statistics set is formed by adding a feature selected by an evaluation of the features for the scaling factors. Given the LPS score, function . Iteration terminates when no feature can be added the likelihood that a sample is in the first of the two classes that improves the evaluation function . The evaluation func is determined using this formula : 30 tion is the number of samples correctly predicted either ( 1 ) by the model built on all of the samples, or ( 2 ) in leave one - out cross - validation (Dash and Liu , 1997 ) . Ties are $ (LPS ( X ); Â1, 6 ) broken by using the feature that has a higher univariate P ( X ESI) = association with the outcome variable . Multiple marker sets O ( LPS( X ); Îll, ã î ) + ° ( LPS( X ) ; fl2 , ôž) ' 35 can be generated by repeated rounds of feature selection , each time removing the features already selected . where p ( x ; ? , o ? ) represents the normal density function Specific Application of Class Prediction with mean u and variance o?, and û1, 0 , ?, üzand ô , are the Linear Predictor Score (LPS ) observed means and variances of the LPS scores for cat - Using the 162 bortezomib - treated patients classified into egory 1 and category 2 . In our case , for example, category 40 Responsive or Nonresponsive groups, the table below shows 1 would be responders, and category 2 would be non the markers in the first LPS predictive set we built from our responders . Then the prediction for a new sample would be data set . Also indicated is whether the marker is more highly that it would be in the first class with probability P (XES , ) expressed in Responsive ( R ) or in Non - responsive ( N ) and in the second class with probability P (XES2 ) = 1 - P patients. The probe set annotations are those provided by ( XES ) . Affymetrix . TABLE 4

LPS Predictive Marker Set SEQ Sub Gene ID set Order Probe Set Chip Symbol Description NO : Direction 1 1 210532 _ s _ at A C14orf2 chromosome 14 open reading 608 N frame 2 -1 2 206790 _ s _ at A NDUFB1 NADH dehydrogenase 516 N (ubiquinone ) 1 beta subcomplex , 1 , 7 kDa 3 200082 _ s _ at A RPS7 ribosomal protein S7 514 N 4 217988 _ at A CCNB11P1 cyclin B1 interacting protein 1 459 N 5 200937 _ s _ at A RPL5 ribosomal protein L5 520 N 6 213941 _ x _ at A RPS7 ribosomal protein S7 2 N 7 224616 _ at A DNCLI2 dynein , cytoplasmic , light 621 R intermediate polypeptide 2 8 224985 _ at A SS18 synovial sarcoma 578 N translocation , chromosome 18 9 233252 _ s _ at A STRBP spermatid perinuclear RNA 9 Z binding protein US 9 , 963, 747 B2 161 162 TABLE 4 -continued LPS Predictive Marker Set SEQ Sub Gene ID set Order Probe Set Chip Symbol Description NO : Direction ??3 10 206621 _ s _ at A WBSCR1 Williams- Beuren syndrome 1275 N chromosome region 1 3 11 208540 _ x _ at B S100A11P S100 calcium binding protein 627 R All pseudogene ??? 12 202605 _ at A GUSB glucuronidase , beta 461 N ? 13 201637 _ s _ at A FXR1 fragile X mental retardation , 1122 N autosomal homolog 1 ? 14 209475 _ at B USP15 ubiquitin specific protease 15 862 R ? 15 200626 _ s _ at B MATR3 matrin 3 54 N ? 16 219939 _ s _ at UNR upstream of NRAS 589 N

? 17 219910 _ at A HYPE Huntingtin interacting protein E 649 R 18 200034 _ s _ at A RPL6 ribosomal protein 16 29 N ? 19 201784 _ s _ at SMAP small acidic protein 134 N ? 20 211316 _ x _ at A CFLAR CASP8 and FADD - like 849 R apoptosis regulator ?5 21 213080 _ x _ at A RPL5 ribosomal protein L5 145 N

It will be appreciated that additional marker sets may be trations of aspects of the invention . Functionally equivalent obtained by employing the methods described herein , and methods and components are within the scope of the inven methods standard in the field , for identifying models . There 25 tion , in addition to those shown and described herein and are many highly correlated features that could be substituted will become apparent to those skilled in the art from the for each other in the models ; these are not all listed . Similar foregoing description , using no more than routine experi methods may be employed utilizing one or more markers mentation . Such equivalents are intended to be encompassed from the identified marker sets of the present invention in by the following claims. order to generate Predictive Marker Sets . 30 All references cited herein , including journal articles , The present invention is not to be limited in scope by the patents , and databases are expressly incorporated by refer specific embodiments described that are intended as illus ence .

SEQUENCE LISTING The patent contains a lengthy “ Sequence Listing” section . A copy of the “ Sequence Listing” is available in electronic form from the USPTO web site ( http : // seqdata .uspto . gov / ? pageRequest = docDetail & DocID = US09963747B2 ) . An electronic copy of the “ Sequence Listing ” will also be available from the USPTO upon request and payment of the fee set forth in 37 CFR 1 . 19 ( b ) ( 3 ).

45 What is claimed is : b ) comparing the level of expression of the at least one 1 . A method for treating hematological cancer in a patient nucleic acid sequence to a reference expression level of with a cancer therapy regimen comprising : that sequence to determine whether the level of expres a )measuring the level of expression of at least one nucleic sion of the at least one nucleic acid sequence is upregu acid sequence selected from the group consisting of lated in the patient sample comprising tumor cells , sequences recognized by probesets of predictive thereby identifying whether expression in the sample marker or markers numbered 1- 547 in Table 1A , 658 includes a profile of expression of the predictive marker 871 in Table 1B , 873 - 876 in Table 1B , 912 - 1062 in or markers ; Table 2A , 1071 - 1079 , 1081 - 1087 , 1089 - 1093 , 1095 - 55 c ) selecting a patient whose predictive marker profile 1106 , and / or 1108 - 1185 in Table 2B , in a patient sample indicates that the patient will respond to the cancer comprising hematological tumor cells , therapy regimen ; and wherein the sequences recognized by probesets of the predictive markers : d ) treating the patient with the cancer therapy regimen , i) numbered 1 -547 consist of SEQ ID NOs : 1 -513 , 60 wherein the hematological cancer is multiple myeloma, ii ) numbered 658 -871 and 873 - 876 consist of SEQ ID wherein the hematological tumor cells are multiple NOs: 614 - 830 , myeloma tumor cells , and wherein the cancer therapy iii ) numbered 912 - 1062 consist of SEQ ID NOs: 866 regimen is proteasome inhibition -based therapy and / or 1014 , 434 and 733 , and glucocorticoid based therapy . iv ) numbered 1071 -1079 , 1081 -1087 , 1089 -1093 , 65 2 . The method of claim 1 wherein the level of expression 1095 - 1106 and 1108 - 1185 consist of SEQ ID NOs : of the at least one nucleic acid sequence is measured by 1023 - 1133 ; detection of mRNA . US 9 , 963, 747 B2 163 164 3 . The method of claim 1 wherein thereby determining the expression profile of the predic i) upregulation of at least one nucleic acid sequence tive marker or markers ; and selected from the group consisting of sequences rec d ) continuing treatment of the hematological cancer when ognized by probesets of predictive markers numbered the expression profile of the predictive marker or markers in the tumor sample demonstrates responsive 1 -547 indicates nonresponsiveness to proteasome inhi- 5 ness ; bition therapy and the patient would not benefit from wherein the hematological cancer is multiple myeloma this cancer therapy regimen , and wherein the hematological tumor cells are multiple ii ) upregulation of at least one nucleic acid sequence myeloma tumor cells . selected from the group consisting of sequences rec 6 . The method of claim 5 wherein the level of expression ognized by probesets of predictive markers numbered " of the at least one nucleic acid is determined by detection of 658 - 871 and 873 - 876 indicates responsiveness to pro mRNA . teasome inhibition therapy and the patient would ben 7 . The method of claim 5 wherein the cancer therapy efit from this cancer therapy regimen , regimen comprises a proteasome inhibition - based regimen iii) upregulation of at least one sequence selected from the therapy . group consisting of sequences recognized by probesets 8 . The method of claim 5 wherein the expression profile of predictive markers numbered 912 - 1062 indicates is determined by a predictive marker set comprising two or nonresponsiveness to glucocorticoid therapy and the more predictive markers . patient would not benefit from this cancer therapy 9 . The method of claim 5 , wherein the proteasome inhi regimen ; and bition - based regimen for treating the tumor comprises treat iv ) upregulation of at least one sequence selected from the 20 ment with a proteasome inhibitor is selected from the group group consisting of sequences recognized by probesets consisting of a peptidyl aldehyde , a peptidyl boronic acid , a of predictive markers numbered 1071 - 1079 , 1081- peptiay?peptidyl boronicb ester, a vinyl sulfone , an epoxyketone , and 1087, 1089 - 1093 , 1095 - 1106 and 1108 - 1185 indicates a lactacystin analog . responsiveness to glucocorticoid therapy and the 35 10 . The method of claim 4 , wherein the predictive marker patient would benefit from this cancer therapy. set is identified by applying a statistical analysis method to 4 . The method of claim 1 wherein the profile of expressionin the expression level of each marker to select features asso is determined by a predictive marker set comprising two or ciated with responsiveness or non -responsiveness . 11 . The method of claim 10 , wherein the statistical more predictive markers . analysis method is the linear predictive score method or the treating5 . A method a hematological for continuing cancer a incancer a patient therapy comprising regimen : 101 30 k nearest neighbors model a ) treating the patient with a cancer therapy regimen 12 . The method of claim 1, wherein the proteasome comprising proteasome inhibition therapy and / or glu inhibition -based therapy comprises treatment with a protea cocorticoid therapy , some inhibitor selected from the group consisting of a b ) obtaining a tumor sample from the patient, wherein the 35 peptidyl aldehyde , a peptidyl boronic acid , a peptidyl tumor sample comprises hematological tumor cells ; 33 boronic ester , a vinyl sulfone, an epoxyketone, and a lacta c ) measuring the level of expression of at least one nucleic cystin analog. acid sequence selected from the group consisting of 13 . The method of claim 12 , wherein the proteasome sequences recognized by probesets of predictive inhibition - based therapy comprises treatment with bort marker or markers numbered 1 -547 in Table 1A , 658 - 10 ezomib . 871 in Table 1B , 873 - 876 in Table 1B , 912 - 1062 in 14 . The method of claim 7 , wherein the proteasome Table 2A , 1071 - 1079 , 1081 - 1087 , 1089 - 1093 , 1095 inhibition - based therapy comprises treatment with bort 1106 , 1108 - 1185 in Table 2B , and / or 1203 - 1423 in ezomib . Table 3in the tumor sample , 15 . The method of claim 4 ,wherein the predictive marker wherein the sequences recognized by probesets of the 45 set comprises at least 10 markers . predictive marker or markers : 45 16 . The method of claim 11 , wherein the predictive i) numbered 1 - 547 consist of SEO ID NOs: 1 -513 marker set comprises markers identified in Table 4 . ii ) numbered 658 - 871 and 873 -876 consist of SEQ ID 17 . The method of claim 8 , wherein the predictive marker NOs: 614 - 830 , set comprises at least 10 markers . iii ) numbered 912 - 1062 consist of SEQ ID NOs: 866 - 50 18 . The method of claim 1 wherein the tumor sample is a 1014 , 434 and 733 , and nsist of SEQ ID NOS: 000 - 50 blood sample . iv ) numbered 1071- 1079 , 1081- 1087 , 1089 - 1093 , 19 . The method of claim 5 wherein the tumor sample is a 1095 - 1106 and 1108 - 1185 consist of SEQ ID NOs: blood sample . 1023 - 1133 , * * * * *