US 2016024.3077A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2016/0243077 A1 BROWN et al. (43) Pub. Date: Aug. 25, 2016

(54) COMPOSITIONS AND METHODS TO (52) U.S. Cl. IMPROVE THE THERAPEUTIC BENEFIT OF CPC ...... A6 IK3I/404 (2013.01); A61 K3I/7056 INDIRUBIN AND ANALOGS THEREOF, (2013.01); A61K.39/3955 (2013.01); C07K INCLUDING MESOINDGO 16/40 (2013.01); C12N 15/1137 (2013.01); A61K 45/06 (2013.01); C12N 23 10/141 (71) Applicants: Dennis M. BROWN, Menlo Park, CA (2013.01); C12N 2320/31 (2013.01) (US); Ian Nisbet, Victoria (AU) (72) Inventors: Dennis M. BROWN, Menlo Park, CA (57) ABSTRACT (US); Ian Nisbet, Victoria (AU) The present invention describes methods and compositions (21) Appl. No.: 14/250,958 for improving the therapeutic efficacy of therapeutically active agents previously limited by Suboptimal therapeutic (22) Filed: Apr. 11, 2014 performance by either improving efficacy as monotherapy or reducing side effects. Such methods and compositions are Related U.S. Application Data particularly applicable to therapeutically active agents selected from the group consisting of: (i) indirubin; (ii) an (63) Continuation-in-part of application No. PCT/US2013/ analog of indirubin; (iii) a derivative of indirubin or of an 033556, filed on Mar. 22, 2013. analog of indirubin; and (iv) a pharmaceutical composition (60) Provisional application No. 61/614,724, filed on Mar. comprising indirubin, an analog of indirubin, or a derivative 23, 2012. of indirubin or of an analog of indirubin, especially meisoin digo. In particular, the therapeutically active agents selected Publication Classification from the group consisting of: (i) indirubin; (ii) an analog of indirubin; (iii) a derivative of indirubin or of an analog of (51) Int. Cl. indirubin; and (iv) a pharmaceutical composition comprising A6 IK3I/404 (2006.01) indirubin, an analog of indirubin, or a derivative of indirubin A6 IK 45/06 (2006.01) or of an analog of indirubin, especially meisoindigo, are C07K 6/40 (2006.01) effective Nek9 inhibitors. The active agent, such as meisoin CI2N 15/I 13 (2006.01) digo, can act as a Nek9 inhibitor. The active agent can be used A6 IK3I/7056 (2006.01) together with Nek9 inhibitors or agents that inhibit the A 6LX39/395 (2006.01) expression of Nek9. Patent Application Publication Aug. 25, 2016 Sheet 1 of 15 US 2016/024.3077 A1

120% 100% 80% 60% 40% 20% O% O 1% DMSO O. 1UM 1UM 1OUM OOUM -0-48 - 24 HOURS HOURS FIG. 1 Patent Application Publication Aug. 25, 2016 Sheet 2 of 15 US 2016/024.3077 A1 Patent Application Publication Aug. 25, 2016 Sheet 3 of 15 US 2016/024.3077 A1

1OO -0-HEL 8O

--- -HKG i. 60

en ? MV4; 11 > 40 2 -(-OC-AML3 de 2O

O O ? 5 10 50 CONCENTRATION OF MEISOINDIGO (UM) FIG. 3 Patent Application Publication Aug. 25, 2016 Sheet 4 of 15 US 2016/024.3077 A1

MEISOINDIGO (050000nM Strongest binding NEK9 Percent Control AISO O O% STK25 O 0.1% PAK O 0.1-1% TSSK1B O 1-5% o 5-10% ERBB3 o 10-35%

MESOINDGO 5OOOOnM

FIG. 4 Patent Application Publication Aug. 25, 2016 Sheet 5 of 15 US 2016/024.3077 A1

Sse3Scs&S.33SS.

N. CN C s Co d d

5)NNINS NOOdOld NIWNO Patent Application Publication Aug. 25, 2016 Sheet 6 of 15 US 2016/024.3077 A1

KNASEARGE ESONGO A8 GENESSBO. CFR50000nM AAK BRSK2 too ABLE255K-PHOSPHORYLATED BTK 8 ABL1(F317-NONPHOSPHORYLATED CAMK 8 ABLF37-PHOSPHORYLATED CAMKD too ABL1(F37L-NONPHOSPHORYLATED CAMKG as ABL1(F347L-PHOSPHORYATED CAMK2A 9 ABL1(H396P-NONPHOSPHORYLATED cAMKB 8 ABL1(H396P-PHOSPHORYLATED ABL (M35T-PHOSPHORYLATED 73 CAK ABLC252HNONPHOSPHORYATED ABL (252H-PHOSPHORYLATED ABL (1345-NONPHOSPHORYLATED ABLT35)PHOSPHORYLATED ABL1(Y253F-PHOSPHORYATED ABL-NONPHOSPHORYAED AB-PHOSPHORYAE) ABL ACVR ACVRB ACVRA CDK4CYCUND 8 ACVR28 CDK-CYCLIND3 too ACVR CDKs to Patent Application Publication Aug. 25, 2016 Sheet 7 of 15 US 2016/024.3077 A1

KASE ARE ABGENESYBBC %CTRG50000nM ACK3 ACK 78 AK AK A3 92 AK APKAPHA APKAPHA ANKK ARKS 85 ASK ASK AURKA ARKB ARKC 94 AX 90 CSNKA BKE is BK O) CSNKD 83 BPRA CSNKE 9 BMPRB CSNKG 95 BPR2 96 CSNKG2 93 BX CSNKG3 so BRAF 84 CSNK2A 79 BRAFVSOOE) CSNKA. 18 BRK BRSK 94 Patent Application Publication Aug. 25, 2016 Sheet 8 of 15 US 2016/024.3077 A1

KNASETARGET (NASEARGE ESENGO ABET GENESYBABO %CFR50000nM AMBIT GENESYBABO CTRL 50000ns, DAPK2 as DAPK3 93 ERN 83 DCARK FAK 83 DCARK2 is DCARK3 9 FES 95 DDR 84 FGFR too DDR2 8 FGFR as DK O FGFRs too DRPK 84 FGFRG697C) too DAPK2 92 FGFR too DRAK FGR s DRAK2 100 FLT 1: DYRK1A FT3 is DYRK1B 92 FLT8D83SH too DYRK2 84 EGFR O EGFRE746A750DEL) FLT3K630) is EGFRG79C) FT3N84) too EGFRG749S) 94 EGFRL747-El.9DEL AISOP) 8 FLT too EGFRL4-S83DEL Piss) is FRK is EGFRL747.75tDEL, SINS) is FYN to EGFRL858R) GAK 8 FIG. 6C Patent Application Publication Aug. 25, 2016 Sheet 9 of 15 US 2016/024.3077 A1

KNASEARGE ABENE SYBBC EGFR1858RT790M) 98 GCN2KNDO,2S803G) 8 EGFRL86tQ 83 GRK too EGFRS752-1759DEL) GRK too EGFRT790M) 96 EF2AK is GSK3A as EPHA 100 EPHA EPHA 9 EFHA4 too EPHAS 96 EPHA is EPHAT 100 EPHA8 94 EPHB EPHB2 100 EPHB3 90 EPHBA 98 EPHB6 ERBB2 too ERBB3 ERBB4 ERK 40 ERK too ERK3 too ERK4 JAK HIDOMAINCATALYTIC 94 ERKS JAKH2DOMAINPSEUDOKNASE) too Patent Application Publication Aug. 25, 2016 Sheet 10 of 15 US 2016/024.3077 A1

KNASETARGET KASEARGE AMBIT GENESYBREBOL JAK2(HYDOMAINCATALYTIC) MERTK 100 JAK3H DOMAINCATALYTC) MET too JNK MET(M250T) 80 JNK2 METY235D) 99 JNK3 NK KT 94 MKK KTA829P) 3 MKNK 98 KTD816H) KNK 68 KTD816V) CK too KTL576P) 98 MLK O KTVS59D) 9 MLK KITYS59D, T670) 93 MLK KTNS59D, V654A) RCKA 92 LATS 00 ARCKB LATS2 9 MST LCK 9 MSTR 92 LAK 84 MST? 99 LAK2 00 S3 LKB 100 S4 LOK MTOR 90 LRRK2 is MUSK 86 LRRK2G2019S) YK LTK 92 MYK FIG. 6E Patent Application Publication Aug. 25, 2016 Sheet 11 of 15 US 2016/024.3077 A1

KNASETARGET KNASEARGE AREBT GENESYBABOL %CRig 50000nM ABT GENESYBREBOL %CTR850000nM LYN 100 YK L2K 63 MY03A MAK Y03B OO MAP3K 83 NDR MAP3K15 NR 09 RAP3K 99 NEK 92 MAP3K3 94 NEK MAP3K4 NEK 92 RAPK 92 NEK O MAP4K3 90 NEK 86 MAP4K4 O NEKS 96 RAPAKS NEK 3 MAPKAPK2 00 NEK 59 APKAPK5 NEK9 RARK 92 NIM as MARK2 NK ARK3 98 OSR MARK4 P38APHA MAST 85 P38 BETA 09 EK P38DEA MEK2 P38-GAA 9 MEK PAK MEK4 OO PAK MEKS PAK3 MEK6 O PA 84 MELK 8 PAKs FIG. 6F

Patent Application Publication Aug. 25, 2016 Sheet 13 of 15 US 2016/024.3077 A1

KNASEARGE ARBGENESYBBC PIK3CAM1043) PIK3CAQ346K) 92 PIK3CD 9s PIK3CG RPS6KAKNDOMINTERMINAL) 82 PK4CB RPS6KAKNDOM2C-TERMINAL 89 P 3 PR PM3 79 RSKIKNDOMINTERMINAL 96 PPSKA RSKKNDOM.2CTERMINAL 84 PPSKC RSK2KINDOMANTERMINAL 94 PPSKB PPSKC is PKACAPHA 93 RSKKNDOMANTERMINAL 99 PKACBEA 9 PKRY PKN 99 PKNBMTUBERCULOSS) OO PK P 94 P3 P4 79 PRKC PRKCE PRKC PRKC GO Patent Application Publication Aug. 25, 2016 Sheet 14 of 15 US 2016/024.3077 A1

KNASEARGE ABGENESYBB0. %CTRiQ 50000nM SRPK SRPK3 00 SKS SK33 00 STK35 SK36 so SK39 78 SYK 85 AK AOK AOK? 84 AOK3 as BK so TEC 00 TESK TGFBR 00 GFBR 94 TE 83 E2 K 8 K 8 NK 00 NK 98 FIG. 6 Patent Application Publication Aug. 25, 2016 Sheet 15 of 15 US 2016/024.3077 A1

KNASEARGE ESONGO ARE GENESYBBO %CFR50000nM NK TNN3K TRKA TRKB RKC TRP6 SSKB TK XK TYK2(JHDOMAINCATALYTIC) TYK2(JHDOMAINPSEUDOKINASE) TYRO3 UK K2 K3 WEGFR2 WRK WEE WEE2 YANK YANK YANK3 YES YSK YSK4 ZAK FIG. 6J US 2016/0243077 A1 Aug. 25, 2016

COMPOSITIONS AND METHODS TO ment (angiogenesis) or gene and antisense therapies to alter IMPROVE THE THERAPEUTIC BENEFIT OF the genetic make-up of cancer cells, and other biological INDIRUBIN AND ANALOGS THEREOF, response modifiers. INCLUDING MEISOINDGO 0005. The work supported by the NCI, other governmental agencies both domestic and foreign in academic or industrial CROSS-REFERENCES research and development laboratories has resulted in an extraordinary body of biological, chemical and clinical infor 0001. This application is a continuation-in-part of PCT mation. In addition, large chemical libraries have been cre Patent Application Serial No. PCT/US2013/033556 by D. M. ated, as well as highly characterized in vitro and in vivo Brown, filed Mar. 22, 2013 and entitled “Compositions and biological Screening systems that have been Successfully Methods to Improve the Therapeutic Benefit of Indirubin and used. However, from the tens of billions of dollars spent over Analogs Thereof, Including Meisoindigo.” which PCT appli the past thirty years Supporting these programs both preclini cation in turn claimed the benefit of U.S. Provisional Appli cally and clinically, only a small number of compounds have cation Ser. No. 61/614,724 by D. M. Brown, filed Mar. 23, been identified or discovered that have resulted in the suc 2012 and entitled “Compositions and Methods to Improve the cessful development of useful therapeutic products. Never Therapeutic Benefit of Indirubin and Analogs Thereof, theless, the biological systems both in vitro and in vivo and Including Meiso indigo.” The contents of both PCT Patent the “decision trees’ used to warrant further animal studies Application Serial No. PCT/US2013/033556 and U.S. Provi leading to clinical studies have been validated. These pro sional Application Ser. No. 61/614,724 are incorporated grams, biological models, clinical trial protocols, and other herein in their entirety by this reference. information developed by this work remain critical for the discovery and development of any new therapeutic agent. FIELD OF THE INVENTION 0006 Unfortunately, many of the compounds that have 0002 The present invention relates to the general field of Successfully met the preclinical testing and federal regulatory hyperproliferative diseases including oncology with a focus requirements for clinical evaluation were either unsuccessful on novel methods and compositions for the improved utility or disappointing in human clinical trials. Many compounds of chemical agents, compounds, and dosage forms previously were found to have untoward or idiosyncratic side-effects that limited by Suboptimal human therapeutic performance were discovered during human clinical Phase I dose-escala including indirubin and analogs thereof. Such as meisoindigo, tion studies used to determine the maximum tolerated dose particularly use to inhibit the kinase Nek9. (MTD) and side-effect profile. In some cases, these toxicities or the magnitude of their toxicity were not identified or pre BACKGROUND OF THE INVENTION dicted in preclinical toxicology studies. In other cases, chemi cal agents where in vitro and in vivo studies Suggested a 0003. The search for and identification of cures for many potentially unique activity against a particular tumor type, life-threatening diseases that plague humans still remains an molecular target or biological pathway were not successful in empirical and sometimes Serendipitous process. While many human Phase II clinical trials where specific examination of advances have been made from basic Scientific research to particular cancer indications/types were evaluated in govern improvements in practical patient management, there still ment sanctioned (e.g., U.S. FDA), IRB approved clinical remains tremendous frustration in the rational and Successful trials. In addition, there are those cases where potential new discovery of useful therapies particularly for life-threatening agents were evaluated in randomized Phase III clinical trials diseases such as cancer, inflammatory conditions, infection, where a significant clinical benefit could not be demon and other conditions. strated; Such cases have also been the cause of great frustra 0004 Since the “Waron Cancer” began in the early 1970's tion and disappointment. Finally, a number of compounds by the United States National Cancer Institute (NCI) of the have reached commercialization but their ultimate clinical National Institutes of Health (NIH), a wide variety of strate utility has been limited by poor efficacy as monotherapy gies and programs have been created and implemented to (<25% response rates) and untoward dose-limiting side-ef prevent, diagnose, treat and cure cancer. One of the oldestand fects (Grade III and IV) (e.g., myelosuppression, neurotoxic arguably most Successful programs has been the synthesis ity, cardiotoxicity, gastrointestinal toxicities, or other signifi and screening of small chemical entities (<1500 MW) for cant side effects). biological activity against cancer. This program was orga 0007. In many cases, after the great time and expense of nized to improve and streamline the progression of events developing and moving an investigational compound into from chemical synthesis and biological Screening to preclini human clinical trials and where clinical failure has occurred, cal studies for the logical progression into human clinical the tendency has been to return to the laboratory to create a trials with the hope of finding cures for the many types of better analog, look for agents with different structures but life-threatening malignant tumors. The synthesis and screen potentially related mechanisms of action, or try other modi ing of hundreds of thousands of chemical compounds from fications of the drug. In some cases, efforts have been made to academic and industrial sources, in addition to the screening try additional Phase I or II clinical trials in an attempt to make of natural products and extracts from prokaryotes, inverte some improvement with the side-effect profile or therapeutic brate animals, plant collections, and other sources from all effect in selected patients or cancer indications. In many of over the world has been and continues to be a major approach those cases, the results did not realize a significant enough for the identification of novel lead structures as potential new improvement to warrant further clinical development toward and useful medicines. This is in addition to other programs product registration. Even for commercialized products, their including biotherapeutics designed to stimulate the human ultimate use is still limited by suboptimal performance. immune system with vaccines, therapeutic antibodies, cytok 0008. With so few therapeutics approved for cancer ines, lymphokines, inhibitors of tumor blood vessel develop patients and the realization that cancer is a collection of US 2016/0243077 A1 Aug. 25, 2016

diseases with a multitude of etiologies and that a patients 0012. In the inventive compositions and methods, the term response and Survival from therapeutic intervention is com Suboptimal therapy includes agents where Phase I toxicity plex with many factors playing a role in the Success or failure precluded further human clinical evaluation. It also includes of treatment including disease indication, stage of invasion those agents from Phase II trials where limited (<25% and metastatic spread, patient gender, age, health conditions, response rates) or no significant tumor responses were iden previous therapies or other illnesses, genetic markers that can tified. Also, Suboptimal therapy includes those agents, the either promote or retard therapeutic efficacy, and other fac subject of Phase III clinical trials the outcome of which was tors, the opportunity for cures in the near term remains elu either medically or statistically not significant to warrant sive. Moreover, the incidence of cancer continues to rise with regulatory Submission or approval by government agencies an approximate 4% increase predicted for 2003 in the United for commercialization or commercialized agents whose clini States by the American Cancer Society such that over 1.3 cal performance (i.e. response rates) as a monotherapy are million new cancer cases are estimated. In addition, with less than 25%, or whose side-effects are severe enough to advances in diagnosis such as mammography for breast can limit wide utility. Agents with suboptimal clinical activity cer and PSA tests for prostate cancer, more patients are being include but are not limited to the following: indirubin and diagnosed at a younger age. For difficult to treat cancers, a analogs thereof, including meisoindigo. patient’s treatment options are often exhausted quickly 0013. One aspect of the invention is a method to improve resulting in a desperate need for additional treatment regi the efficacy and/or reduce the side effects of suboptimally mens. Even for the most limited of patient populations, any administered drug therapy comprising the steps of additional treatment opportunities would be of considerable 0014 (i) identifying at least one factor or parameter asso value. This invention focuses on inventive compositions and ciated with the efficacy and/or occurrence of side effects of methods for improving the therapeutic benefit of subopti the drug therapy; and mally administered chemical compounds including indirubin 00.15 (ii) modifying the factor or parameter to improve the and analogs thereof. efficacy and/or reduce the side effects of the drug therapy. 0009 Relevant literature includes Foye, W. O., “Cancer 0016. In this method, the factor or parameter can be Chemotherapeutic Agents. American Chemical Society, selected from the group consisting of: 1995, and Dorr, R.T., and Von Hoff, D.D., “Cancer Chemo 0017 (i) dose modification; therapy Handbook. Appleton and Lange, 1994. 0018 (ii) route of administration; 0010. Therefore, there is a need for compositions and 0019 (iii) schedule of administration; methods that improve the therapeutic benefit of suboptimally 0020 (iv) indications for use: administered chemical compounds and therapeutic composi 0021 (v) selection of disease stage; tions, particularly with respect to the inhibition of the kinase 0022 (vi) other indications; NEK9. 0023 (vii) patient selection; 0024 (viii) patient/disease phenotype: SUMMARY OF THE INVENTION 0025 (ix) patient/disease genotype: 0026 (x) pre?post-treatment preparation 0011. This invention meets the needs described above for 0027 (xi) toxicity management; compositions and methods that improve the therapeutic ben 0028 (xii) pharmacokinetic/pharmacodynamic monitor efit of Suboptimally administered chemical compounds and 1ng therapeutic compositions. Specifically, this invention relates 0029 (xiii) drug combinations: to novel compositions and methods to improve the utility of 0030 (xiv) chemosensitization; chemical agents with Suboptimal performance in patients 0031 (XV) chemopotentiation: suffering with cancer. The invention describes novel 0032 (xvi) post-treatment patient management; improvements, pharmaceutical ingredients, dosage forms, 0033 (xvii) alternative medicine/therapeutic support; excipients, solvents, diluents, drug delivery systems, preser 0034 (xviii) bulk drug product improvements: Vatives, more accurate drug administrations, improved dose determination and schedules, toxicity monitoring and ame 0035 (xix) diluent systems; liorization, techniques or agents to circumvent or reduce tox 0036 (XX) solvent systems; icity, techniques and tools to identify/predict those patients 0037 (XXi) excipients: who might have a better outcome with a therapeutic agent by 0038 (XXii) dosage forms; the use of phenotype or genotype determination through the 0039 (XXiii) dosage kits and packaging; use of diagnostic kits or pharmacokinetic or metabolism 0040 (xxiv) drug delivery systems; monitoring approaches. The invention also relates to the use 0041 (XXV) drug conjugate forms; of drug delivery systems, novel prodrugs, polymer conju 0042 (XXVi) compound analogs; gates, novel routes of administration, other agents to poten 0043 (XXvii) prodrugs; tiate the activity of the compounds or inhibit the repair of 0044 (XXvii) multiple drug systems: suboptimal cellular effects or sublethal damage or to “push' 0045 (XXviii) biotherapeutic enhancement; the cell into more destructive cellular phases Such as apopto 0046 (xxix) biotherapeutic resistance modulation; sis. In some case, the use of these Suboptimal therapeutics in 0047 (XXX) radiation therapy enhancement; conjunction with radiation or other conventional chemothera 0048 (XXXi) novel mechanisms of action; and peutic agents or biotherapeutic agents such as antibodies, 0049 (XXXii) selective target cell population therapeutics. vaccines, cytokines, lymphokines, gene and antisense thera 0050. The drug therapy can be administered to treat a pies, or other chemotherapeutic or biotherapeutic agents, hyperproliferative disease, Such as cancer. would provide novel approaches and significant improve 0051 Typically, the suboptimally administered drug ment. In particular, the present invention provides novel therapy comprises administration of a therapeutically active approaches for the inhibition of the kinase Nek9. agent selected from the group consisting of: (i) indirubin; (ii) US 2016/0243077 A1 Aug. 25, 2016

an analog of indirubin; (iii) a derivative of indirubin or of an unmodified therapeutic agent is indirubin or an analog analog of indirubin; and (iv) a pharmaceutical composition thereof, particularly meisoindigo. comprising indirubin, an analog of indirubin, or a derivative 0061. In one alternative, the invention provides methods of indirubin or of an analog of indirubin, including meisoin and compositions for the treatment of malignancies by the digo. inhibition of the kinase Nek9. 0052 Another aspect of the present invention is a compo sition to improve the efficacy and/or reduce the side effects of BRIEF DESCRIPTION OF THE DRAWINGS Suboptimally administered drug therapy comprising an alter native selected from the group consisting of 0062. The following invention will become better under 0053 (1) a therapeutically effective quantity of a modified stood with reference to the specification, appended claims, therapeutic agent or a derivative, analog, or prodrug of a and accompanying drawings, where: therapeutic agent or modified therapeutic agent, wherein the 0063 FIG. 1 shows the viability of the AML cell line MV modified therapeutic agent or the derivative, analog or pro 4-11 (FLT-3 ITD) after meisoindigo treatments. Viability at drug of the therapeutic agent or modified therapeutic agent 48 hours is shown by (); viability at 24 hours is shown by (). Results are shown for a control, 1% DMSO without possesses increased therapeutic efficacy or reduced side meisoindigo, 0.1 LM meisoindigo, 1 LM meisoindigo, 10uM effects as compared with an unmodified therapeutic agent; meisoindigo, and 100 uM meisoindigo. 0054 (2) a composition comprising: 0064 FIG. 2 shows the viability of a number of myeloid 0055 (a) a therapeutically effective quantity of a thera cell lines in terms of the percentage of viable cells after 24 peutic agent, a modified therapeutic agent or a deriva hours of treatment with meisoindigo. Results are shown for a tive, analog, or prodrug of a therapeutic agent or modi control, 1% DMSO without meisoindigo, 0.1 uM meisoin fied therapeutic agent; and digo, 1 LM meisoindigo, 10 LM meisoindigo, and 100 LM 0056 (b) at least one additional therapeutic agent, meisoindigo. therapeutic agent Subject to chemosensitization, thera peutic agent Subject to chemopotentiation, diluent, 0065 FIG.3 shows the viability of a number of additional excipient, solvent system, or drug delivery system, myeloid cell lines in terms of the percentage of viable cells wherein the composition possesses increased therapeu after 24 hours of treatment with meisoindigo. Results are tic efficacy or reduced side effects as compared with an shown for a control, 1% DMSO without meiso indigo, 0.1 uM unmodified therapeutic agent; meisoindigo, 1 uM meisoindigo, 10 LM meisoindigo, and 0057 (3) a therapeutically effective quantity of a thera 100 uM meiso indigo. peutic agent, a modified therapeutic agent, or a derivative, 0066 FIG. 4 shows the targeting of kinases by meisoin analog, or prodrug of a therapeutic agent or modified thera digo. peutic agent that is incorporated into a dosage form, wherein 0067 FIG. 5 shows the overall survival with meiso indigo, the therapeutic agent, the modified therapeutic agent, or the alone and with hydroxyurea, in chronic myelogenous leuke derivative, analog, or prodrug of a therapeutic agent or modi mia, together with the results for busulfan. fied therapeutic agent incorporated into the dosage form pos 0068 FIG. 6, shown as FIG. 6A, FIG. 6B, FIG. 6C, FIG. sesses increased therapeutic efficacy or reduced side effects 6D, FIG. 6E, FIG. 6F, FIG. 6G, FIG. 6H, FIG. 6I, and FIG. as compared with an unmodified therapeutic agent; 6A, is a table showing the results of a screen for kinase 0058 (4) a therapeutically effective quantity of a thera inhibition by meisoindigo, wherein meisoindigo shows sig peutic agent, a modified therapeutic agent, or a derivative, nificant inhibition of the kinase Nek9. FIG. 6A shows results analog, or prodrug of a therapeutic agent or modified thera for AAK1 to CDK7. FIG. 6B shows results for ADCK3 to peutic agent that is incorporated into a dosage kit and pack DADK 1. FIG. 6C shows results for DAPK2 to GAK. FIG. 6D aging, wherein the therapeutic agent, the modified therapeu shows results for EGFR (L858R, T790M) to JAK1 tic agent, or the derivative, analog, or prodrug. ofatherapeutic (JH2DOMAIN-PSEUDOKINASE). FIG. 6E shows results agent or modified therapeutic agent incorporated into the for JAK2 (JH1 DOMAIN-CATALYTIC) to MYLK2. FIG. 6F dosage kit and packaging possesses increased therapeutic shows results for LYN to PAK6. FIG. 6G shows results for efficacy or reduced side effects as compared with an unmodi PAK7 to RIPK5. FIG. 6H shows results from PIK3A fied therapeutic agent; and (M1043I) to SRPK1. FIG. 6I shows results for SRPK2 to 0059 (5) a therapeutically effective quantity of a thera TNK1. FIG. 6J shows results for TNK2 to ZAK. peutic agent, a modified therapeutic agent, or a derivative, analog, or prodrug of a therapeutic agent or modified thera DETAILED DESCRIPTION OF THE INVENTION peutic agent that is Subjected to a bulk drug product improve 0069. This invention relates to novel compositions and ment, wherein the therapeutic agent, the modified therapeutic methods to improve the utility of chemical agents including agent, or the derivative, analog, or prodrug of a therapeutic indirubin and analogs and derivatives thereof with subopti agent or modified therapeutic agent Subject to the bulk drug mal performance for patients with cancer, as well as addi product improvement possesses increased therapeutic effi tional therapeutic agents or agents capable of therapeutic cacy or reduced side effects as compared with an unmodified application. The invention describes the novel development therapeutic agent; of improved pharmaceutical ingredients, dosage forms, wherein the therapeutic agent, the modified therapeutic agent excipients, solvents, diluents, drug delivery systems, preser or the derivative, analog, or prodrug of a therapeutic agent or Vatives, more accurate drug administrations, improved dose modified therapeutic agent is selected from the group con determination and Schedules, toxicity monitoring and ame sisting of: (i) indirubin; (ii) an analog of indirubin; and (iii) a liorization, techniques or agents to circumvent or reduce tox derivative of indirubin or of an analog of indirubin. icity, techniques and tools to identify/predict those patients 0060 Typically, the composition possesses increased effi who might have a better outcome with a therapeutic agent by cacy or reduced side effects for cancer therapy. Typically, the the use of phenotype or genotype determination through the US 2016/0243077 A1 Aug. 25, 2016 use of diagnostic kits or pharmacokinetic or metabolism monitoring approaches, the use of drug delivery systems, (I) novel prodrugs, polymer conjugates, novel routes of admin istration, other agents to potentiate the activity of the com pounds or inhibit the repair of suboptimal cellular effects or sub-lethal damage or to “push the cell into more destructive cellular phases such as apoptosis. In some cases, the inventive O examples include the use of these Sub-optimal therapeutics in NH conjunction with radiation or other conventional chemothera peutic agents or biotherapeutic agents such as antibodies, vaccines, cytokines, lymphokines, gene and antisense thera pies, or other chemotherapeutic or biotherapeutic agents. 0070. By definition, the term “suboptimal therapy” includes agents where Phase I toxicity precluded further 0076 Also within the scope of the invention are deriva human clinical evaluation. It also includes those agents from tives of indirubin that, for example, have the hydrogen of the Phase II trials where limited or no significant tumor responses benzene rings replaced with halo or lower alkyl or have the were identified. In addition, it also includes those agents, the hydrogen attached to the nitrogen of the five-membered rings subject of Phase III clinical trials, whose outcome was either replaced with lower alkyl. medically or statistically not significant to warrant Submis 0077 Analogs of indirubin include, but are not limited to, sion or approval by regulatory agencies for commercializa the compounds described below and derivatives thereof. tion or commercialized agents whose response rates as a 0078. One particular analog of indirubin is a compound monotherapy are less than 25% or whose side-effects are named meisoindigo, whose systemic name is 3-(1,2-dihydro severe enough to limit wider utility. Agents with Suboptimal 2-oxo-3H-indol-3-ylidene)-1,3-dihydro-1-methyl-2H-indol activity include but are not limited to the following: indirubin 2-one. Meisoindigo has the structure shown as Formula (II): and analogs and derivatives thereof. More specifically, the inventive methods and compositions also focus on improve ments for indirubin and analogs and derivatives thereof, (II) including meisoindigo. 0071 (I) Suboptimal Therapeutics 0072. In general, examples of compounds with subopti mal therapeutic activity include, but are not limited to, com pounds of the following classes: DNA/nucleic acid binding/ reactive agents, topoisomerase inhibitors, anti-tubulin agents, signal transduction inhibitors, protein synthesis inhibitors, inhibitors of DNA transcribing enzymes, DNA/RNA interca lating agents, DNA minor groove binders, drugs that block steroid hormone action, photochemically active agents, (0079 Meiso indigo is a derivative of the Chinese herb, immune modifying agents, hypoxia selective cytotoxins, Qing dai, Indigo naturalis. It is a second-generation synthetic chemical radiation sensitizers and protectors, antisense derivative of indirubin and is a water-soluble small molecule. nucleic acids, oligonucleotides and polynucleotides thera Meisoindigo is active against both cell lines and primary peutic agents, immune modifying agents, antitumor antibiot cancer cells. Its ICso is in the high-uM concentration range. It ics, biotherapeutics, biologic agents such as cancer vaccines, has shown activity against AML (acute myelogenous leuke antibody therapies, cytokines, lyphokines, gene therapies, mia) (NB4, HL-60, U937), CML (chronic myelogenous leu nucleic acid therapies, vascular disrupting agents, anti-angio kemia (K562), colorectal cancer (HT29), HUVEC, ECV304, genic agents, and cellular therapies. In some cases, a com as well as against primary AML. Meisoindigo is metabolized pound may fall within more than one of these classes; Such by cytochrome P450 1A2 and 2C19 by oxidation. Meisoin compounds are also within the scope of the invention. digo is also metabolized by uridine glucuronyltransferases (UGT) via direct conjugation (i.e., without initial cytochrome 0073. In some cases, compounds or compositions may be P450 oxidation) to produce more polar metabolites. Meisoin in current clinical use for one or more indications, but yet be digo also inhibits cytochrome P450 1A2 and 2C8 at low considered Suboptimal for another indication, such as a dif micromolar concentrations. ferent type of malignancy, either in terms of the cell type 0080 FIG. 1 shows the viability of the AML cell line MV involved in the malignancy or in terms of the stage of the 4-11 (FLT-3 ITD) after meisoindigo treatments. Viability at malignancy. Such compounds or compositions are within the 48 hours is shown by (0); viability at 24 hours is shown by Scope of the invention. (). Results are shown for a control, 1% DMSO without 0074 Specific examples include indirubin analogs and meisoindigo, 0.1 LM meisoindigo, 1 LM meisoindigo, 10uM derivatives thereof as described below, including, but not meisoindigo, and 100 uM meisoindigo. limited to, meisoindigo. I0081 FIG. 2 shows the viability of a number of myeloid 0075) The structure of indirubin is shown in Formula (I), cell lines in terms of the percentage of viable cells after 24 below. hours of treatment with meisoindigo. Results are shown for a US 2016/0243077 A1 Aug. 25, 2016

control, 1% DMSO without meiso indigo, 0.1 uM meisoin 3.9 g/kg, an extremely high dose. In rats, up to 400 mg/kg/d digo, 1 LM meisoindigo, 10 LM meisoindigo, and 100 LM p.o. for 30 days is well tolerated. In dogs, 10 mg/kg/dp.o. for meisoindigo. 6 months is well tolerated. The target organs in rat and dog are I0082 FIG.3 shows the viability of a number of additional liver and the gastrointestinal tract. Meisoindigo is not geno myeloid cell lines in terms of the percentage of viable cells toxic in the Ames and Chromabs tests. after 24 hours of treatment with meisoindigo. Results are I0089. Several decades of clinical work on meisoindigo shown for a control, 1% DMSO without meiso indigo, 0.1 uM Supports its safety, with clinical trials as a single agent or in meisoindigo, 1 uM meisoindigo, 10 LM meisoindigo, and combination with chemotherapy. It has been well tolerated in 100 uM meisoindigo. several patient populations. Common adverse effects include 0083 Meiso indigo has shown to be effective in murine mild nausea and Vomiting (30%), which spontaneously dis cancer models. Meisoindigo has also been shown to be effec tive in the AML NOD/SCID model, showing dose-dependent appeared in two weeks, with no cardiac, hepatic, or renal spleen size reduction, in HT-29 colorectal xenografts, Lewis adverse effects. However, anticipated potential side effects lung sarcoma, and Walker 256 carcino-sarcoma. include marrow Suppression, joint pain, skin itch, gastrointes 0084 Meiso indigo is active via several, possibly novel tinal symptoms including nausea, vomiting, anorexia, mechanisms of action. It has cell cycle specific effects, abdominal pain, abdominal distention, and diarrhea, edema, including arrest in G(O)/G1 for AML cell lines and G2/M pigmentation of face and extremities, headache, head disten arrest for HT-29 colorectal cell lines. It also stimulates apo tion, or alterations in liver function, with slight increase in ptosis through a number of mechanisms, including the alanine transaminase (ALT). upregulation of p21 and p27 and the downregulation of Bcl-2 0090 Previous experience with meiso indigo includes sev in primary AML cells, as well as upregulation of Bak and Bax eral trials in chronic myelogenous leukemia (CML). The in AML cells (DKO insensitive to chemotherapy), and a novel caspase-dependent pathway in K562 cells. Meisoindigo also regimen was 50-200 mg/dp.o. for several days; it was used as has effects on mitochondria, but with no change in Bcl-2. a single agent and in combinations with other chemothera Bax, and Bid protein expression. Meisoindigo also stimulates peutic agents, including busulfan and hydroxyurea. There the cleavage of pro-caspase 3, 8, 9 and PARP in HL-60 was a reduction of splenomegaly (median 25 d) followed by myeloid cells. leukocyte count (median 31 d). There was a complete hema 0085 Meiso indigo also is directed to multiple cellular tologic response in 33% of patients and a partial hematologic targets, which are possibly synergistic and complementary. response in 58% of patients. A synergistic effect with hydrox For example, it promotes differentiation of human myelo yurea was seen. blastic leukemic cells, accompanied by downregulation of 0091. The overall survival with meiso indigo, alone and c-myb gene expression. It also promotes inhibition of DNA with hydroxyurea, in chronic myelogenous leukemia is and RNA synthesis in W256 cells, microtubule assembly, shown in FIG. 5. FIG. 5 also shows the results with busulfan. glycogen synthase kinase-3? (GSK-3?) (at 5-50 nM), CDK1/ 0092. One possible additional use for meiso indigo is in cyclin B, and CDK5/p25 (tau microtubule protein phospho patients with acute myelogenous leukemia, acute promyelo rylation). Additionally, meisoindigo decreases B-catenin and cytic leukemia (APL), acute lymphoblastic leukemia (ALL), c-myc (HL-60 cells, but not in K562), affects the Wnt path or high-risk myelodysplastic syndrome (MDS) who are way through inhibiting GSK-3? and downregulating B-cate unsuitable for treatment with standard chemotherapy regi nin and c-myc protein expression. Meisoindigo also promotes mens (i.e., elderly, considered to be poor risk, or chemore upregulation of CD11b, promoting myeloid differentiation, fractory). The meisoindigo can be administered as a single and upregulation of Ahi-1 in Jurkat cells (inducing phospho agent orally in a softgel capsule; dosing would start at 100 or 150 mg and increase. If no dose limiting toxicity (DLT) is rylation of c-Myb. Furthermore, meiso indigo exhibits antian seen after 4 weeks of treatment, the highest dose level could giogenic effects, including decreased VEGF protection, be used. Other dosing alternatives are described below, VCAM-1, tubule formulation in HUVEC, and ECV304 apo including use with additional agents in addition to meisoin ptosis. digo. I0086. As shown in FIGS. 4 and 6, meisoindigo targets a number of kinases. The strongest binding is with NEK9, but 0093. Other indirubin analogs and derivatives are within significant binding is also shown with STK25, PAK1, the scope of the present invention. These include, but are not TSSK1B, and ERBB3. This suggests that meiso indigo can limited to, the following: affect a Substantial number of pathways regulated by kinase (0094 U.S. Pat. No. 6,566,341 to Wang et al. discloses activity or by proteins phosphorylated by kinase activity. indirubin (Formula I)), isoindigo (Formula (III)), indigo (For 0087 Meiso indigo is rapidly absorbed and widely distrib mula (IV)) and derivatives thereof, shown in Formulas (V), uted in the tissues. It is metabolized by hepatic cytochrome (VI), and (VII) along with 1-(B-D-O-triacetyl-xylopyrano P450 (CYP)-mediated N-demethylation and mono ring syl)-isoindigo (Formula (VIII). In these indirubin, isoindigo, hydroxylation. There are no major interspecies differences or indirubin derivatives, R. R. R. R. R. R. R. Rs. Ro, between the metabolism of meisoindigo in rats, pigs, and and Ro are independently a hydrogen, a monosaccharide, a humans, and no sex differences in metabolism. The human disaccharide, a halogen, a hydrocarbyl group, or a functional hepatic internal CL is ~83 (Cl mL/mg/kg). Excretion of hydrocarbyl group unsubstituted or substituted with one or meisoindigo is through renal and biliary routes, with renal more hydroxyl moieties, carboxyl moieties, nitroxyl moi excretion strongly predominating. eties, monosaccharides, disaccharides, amines, amides, thi 0088 Rodent and non-rodent toxicology studies indicate a ols, Sulfate groups, Sulfonate groups, Sulfonamide groups, or safe profile for meisoindigo. The single dose LDso in rats is halogens, wherein a hydrocarbyl has 1 to 12 carbon atoms. US 2016/0243077 A1 Aug. 25, 2016

which: (i) when X is an oxygenatom, R is a hydrogenatom, (III) a halogen atom, a —NO group, a methyl group, a Sulfona mide group, or SO. NH CHCH, OH, or (ii) when X is NOH. R' is a hydrogen atom or an iodine atom.

(IX)

N (IV)

NH NH

N H

(V) 0096 U.S. Pat. No. 6,987,092 to Eisenbrand et al. dis closes additional indigoid bisindigo derivatives of Formula (X) in which: X and Y are the same or different and are an oxygen atom; a Sulfur atom; a selenium atom; a tellurium atom; a group N-A-B R'' in which A is a single bond or an oxygenatom, NH or NH CO. : B is a single bond or a group (CD)Z, wherein D has the same meaning as R'' and Z is an oxygen atom or -NH-, n is 0 or an integer and m is an integer; and the group R'' is a hydrogen atom, a straight-chain or branched-chain alkyl group having 1 to 18 carbon atoms which can carry one or more hydroxy and/or amino groups and can be substituted by one or more carboxyl (VI) groups and/or phosphoryl groups, a Substituted or unsubsti tuted aryl group which can comprise one or more heteroat oms, an aralkyl group, an acyl group, a gycoside selected from monosaccharides, disaccharides or oligosaccharides, or a group selected from the group consisting of Sugars, amino acids, peptides or steroid hormones; or a hydrazone group N NR'R'', wherein Rand R' can be the same or differ ent and represent a hydrogen atom, a straight-chain or branched-chain alkyl group having 1 to 18 carbon atoms (VII) which can be substituted by one or more carboxyl groups and/or phosphoryl groups, a Substituted or unsubstituted aryl group which can comprise one or more heteroatoms, an aralkyl group, an acyl group, or a glycoside selected from monosaccharides, disaccharides or oligosaccharides, or a group selected from the group consisting of Sugars, amino acids, peptides or steroid hormones: R,R,R,R,R,R,R and R' can be the same or different and representahydrogen atom; a halogen atom; a hydroxy group; a nitroso group; a nitro group; an aryloxy group: an alkoxy group; a straight (VIII) chain or branched-chain alkyl group having 1 to 18 carbon atoms which can additionally carry one or more hydroxy and/or amino groups; a Substituted or unsubstituted aryl group which can comprise one or more heteroatoms; a cycloalkyl group having 3 to 7 carbonatoms which can com prise one or more heteroatoms; an aralkyl group; a trifluo romethyl group; a —COM group; a —COOM group; a —CHCOOM group, wherein M is hydrogen, a straight chain or branched-chain alkyl group having 1 to 18 carbon OAc atoms which can additionally carry one or more hydroxy and/or amino groups, or an aryl group which can comprise AcO AcO one or more heteroatoms and can be substituted with one or more halogenatoms, one or more alkyl groups or one or more alkoxy groups; a NR'R'' group, wherein R'' and R' can 0095 U.S. Pat. No. 6,664,285 to Eisenbrand et al. dis be the same or different and represent a hydrogen atom, a closes indigoid bisindigo derivatives of Formula (IX) in straight-chain or branched-chain alkyl group having 1 to 18 US 2016/0243077 A1 Aug. 25, 2016 carbon atoms which can additionally carry one or more meanings as for Formula (XI) and R'' and R', which may be hydroxy and/or amino groups, a Substituted or unsubstituted the same or different, have the same meanings as R for aryl group which can comprise one or more heteroatoms, or Formula (X). an acyl group, or R'' and R' form together a ring having 2 to 6, optionally substituted, CH groups; a benzyl group, wherein the benzene nucleus can comprise one or more het (X) eroatoms; a hydroxylamino group; a phosphate group; a phosphonate group; a sulfate group; a sulfonamide group, wherein the nitrogen atom can be independently substituted by a hydrogenatom, a straight-chain or branched-chain alkyl group having 1 to 18 carbon atoms which can additionally carry one or more hydroxy and/oramino groups, a Substituted or unsubstituted aryl group or wherein the nitrogen atom is part of a cycloalkyl group having 3 to 7 carbon atoms which can comprise one or more heteroatoms; an azo group N=N R', in which R' represents an aromatic system (XI) which can be substituted by one or more carboxyl groups and/orphosphoryl groups; or a O-glycoside or a N-glycoside, wherein the glycoside is selected from monosaccharides, dis accharides or oligosaccharides; or a group selected from the group consisting of Sugars, amino acids, peptides or steroid hormones; or R' and R, and Rand R', respectively, form independently from each other a ring together having 1 to 4. optionally substituted, CH groups; the groups R' and Rare the same or different and represent a hydrogen atom; a halo gen atom; a hydroxy group; a methylenehydroxy group; a straight-chain or branched-chain alkyl group having 1 to 18 (XII) carbonatoms; a cycloalkyl group having 3 to 7 carbon atoms which can comprise one or more heteroatoms; a Substituted or unsubstituted aryl group which can comprise one or more heteroatoms; a mono-, di- or trialkylsilyl group having 1 to 6 carbon atoms independently of each other in each instance in the straight-chain or branched-chain alkyl group; a mono-, di or triarylsilyl group with substituted or unsubstituted aryl groups independently of each other in each instance; an aralkyl group; a trifluoromethyl group; a —COM group; a (0097 U.S. Pat. No. 7,582,670 to Wang et al. discloses —COOM group; a —CHCOOM group, wherein M is derivatives and analogs of indirubin of Formulas (V), (VI), hydrogen, a straight-chain or branched-chain alkyl group and (VII) wherein: R. R. R. R. R. R. R. and Ro are the having 1 to 18 carbonatoms which can additionally carry one same or different and represent a hydrogen atom; a hydroxy or more hydroxy and/or amino groups, oran aryl group which group; a nitroso group; a nitro group; a monosaccharide; a disaccharide; a halogenatom; a hydrocarbyl group, or a func can comprise one or more heteroatoms and can be substituted tional hydrocarbyl group unsubstituted or substituted with with one or more halogenatoms, one or more alkyl groups or one or more hydroxy moieties, carboxy moieties, nitroxy one or more alkoxy groups; a NR'R' group, wherein R' moieties, monosaccharides, disaccharides, amines, amides, and R' can be the same or different and representahydrogen thiols, Sulfates, Sulfonates, Sulfonamides or halogens, atom, a straight-chain or branched-chain alkyl group having 1 wherein the hydrocarbyl has 1 to 8 carbon atoms; a —RR to 18 carbon atoms which can additionally carry one or more group, wherein R and R2 can be the same or different and hydroxy and/or amino groups, a Substituted or unsubstituted represent a hydrogen atom, a straight-chain or branched aryl group which can comprise one or more heteroatoms, or chain alkyl group having 1 to 18 carbon atoms which can an acyl group; a methyleneamino group —CH2 NR'R''. additionally carry one or more hydroxy and/or amino groups, wherein R'' and R' have the above definitions; a benzyl a Substituted or unsubstituted aryl group which can comprise group, wherein the benzene nucleus can comprise one or one or more heteroatoms, or an acyl group, or R and R. more heteroatoms; a methylenecycloalkyl group having 3 to form togethera ring having 2 to 6, optionally Substituted, CH 7 carbonatoms which can comprise one or more heteroatoms; groups; an azo group —N=N-R, wherein R represents a physiological amino acid residue bound to the nitrogenas an an aromatic system which can be substituted by one or more amide; an O-glycoside or a N-glycoside, wherein the glyco carboxyl groups and/or phosphoryl groups, or a group side is selected from monosaccharides, disaccharides or oli selected from the group consisting of Sugars, amino acids, gosaccharides; or a group selected from the group consisting peptides or steroid hormones; or R and R, and R2 and R-7, of Sugars, amino acids, peptides or steroid hormones; or a respectively, form independently from each other a ring methylene Sulfonate group. The indigo derivatives have the together having 1 to 4, optionally Substituted, CH groups; general formula Formula (XI) wherein R' to R' and XandY and R and R2 are the same or different and represent a have the same meanings as for Formula (X). Also included are hydrogen atom; a halogen atom; a hydroxy group; a hydro compounds of Formula (XII) wherein R' to R' have the same carbyl group, or a functional hydrocarbyl group unsubstituted US 2016/0243077 A1 Aug. 25, 2016

or substituted with one or more hydroxy moieties, carboxy more places with halogen, C-C alkyl, hydroxy, amino and/ moieties, nitroxy moieties, monosaccharides, disaccharides, or C-C alkoxy; or a —COM group, —COOM group, amines, amides, thiols, Sulfates, Sulfonates, Sulfonamides or CHCOOM group, CONR'R'' group, NR R'R'' halogens, wherein the hydrocarbyl has 1 to 8 carbon atoms; a group, SONR R'R'' group, N=N R group or an mono-, di- or trialkylsilyl group having 1 to 6 carbon atoms —S(O), R group; or an O-glycoside or N-glycoside, independently of each other in each instance in the straight whereby the glycosides are selected from the group of mono chain or branched-chain alkyl group; a mono-, di- or triaryl or disaccharides, R stands for oxygen, sulfur, selenium, tel silyl group with Substituted or unsubstituted aryl groups inde lurium or the group —NOR7 or —NR, R and R stand for pendently of each other in each instance; a —NR,Rs group. hydrogen, halogen, hydroxy, nitroso, nitro, C-C alkoxy wherein R, and Rs can be the same or different and repre sent a hydrogen atom, a straight-chain or branched-chain optionally interrupted by one or more oxygen atoms: C-Cls alkyl group having 1 to 18 carbon atoms which can addition alkyl optionally substituted in one or more places with halo ally carry one or more hydroxy and/or amino groups, a Sub gen, hydroxy and/or amino: aryl, benzyl, benzyloxy, or het stituted or unsubstituted aryl group which can comprise one eroaryl optionally Substituted in one or more places with or more heteroatoms, or an acyl group; a methyleneamino halogen, C-C alkyl, hydroxy, amino and/or C-C alkoxy; group —CH2—NR,Rs, wherein R, and Rs have the or aralkyl, aryloxy, methylenaryloxy, C-Cs cycloalkyl, above definitions; a physiological amino acid residue bound C-Cs cycloalkenyl or C-C methylenecycloalkyl optionally to the nitrogen as an amide, Substituted or unsubstituted Substituted in one or more places with halogen, C-C alkyl, monosaccharide, disaccharides or oligosaccharides; or a hydroxy, amino and/or C-C alkoxy and optionally contain Sugar, amino acid, peptide or steroid hormone. Preferably, at ing one or more heteroatoms; or hydroxylamino, phosphate, least R or R is a monosaccharide, a disaccharide unsubsti phosphonate, Sulfate, Sulfonate, Sulfonamide optionally Sub tuted or substituted with one or more hydroxy moieties or stituted in one or more places with halogen, C-C alkyl, carboxy moieties; a halogen; a hydrocarbyl group, or a func hydroxy, amino and/or C-C alkoxy; or a -COM group, tional hydrocarbyl group unsubstituted or substituted with - COOM group, —CHCOOM group, —CONR'R'' one or more hydroxy moieties, carboxy moieties, nitroxy group, NR R'R'' group, -SONR R'R'' group, moieties, monosaccharides, disaccharides, amines, amides, N=N R group or an S(O)R group; or an O-glyco thiols, Sulfates, Sulfonates, Sulfonamides or halogens, side or N-glycoside, whereby the glycosides are selected wherein the hydrocarbyl has 1 to 8 carbon atoms. In another from the group of mono- or disaccharides, and R stands for embodiment at least R or R is an acetylated monosaccha hydrogen, C-C alkyl that is Substituted in one or more ride. Preferably, at least R or R can be a methyl group. U.S. places with halogen, hydroxy and/or amino: aryl, heteroaryl Pat. No. 7,582,670 to Wang et al. also discloses a compound or C-C cycloalkyl that is optionally substituted in one or that is a derivative of Formula (VIII) above in which the more places with halogen, hydroxy, amino, C-C alkyl and/ hydrogen bound to the nitrogen of the five-membered ring is or C-C alkoxy; or R or R, or R or Rs, independently of replaced with a methyl group. This compound is shown as one another, form a ring with 1 to 4-CH groups, which, independently of one another, optionally are Substituted in Formula (XIII): one or two places with halogen, hydroxy, nitroso, nitro, C-Co alkoxy; C-C alkyl that is optionally substituted in (XIII) one or more places with halogen, hydroxy and/or amino: aryl CH or heteroaryl that is optionally substituted in one or more O I places with halogen, C-C alkyl, hydroxy, amino and/or C-C alkoxy; or aralkyl, aryloxy, methylenaryloxy, C-Cs C N cycloalkyl, C-Cs cycloalkenyl or C-C methylenecy S. O cloalkyl that is optionally Substituted in one or more places O N with halogen, C-C alkyl, hydroxy, amino and/or C-C, OAc O alkoxy and that optionally contains one or more heteroatoms; or hydroxylamino, phosphate, phosphonate, Sulfate, Sul AcO fonate, Sulfonamide that is optionally Substituted in one or AcO more places with halogen, C-C alkyl, hydroxy, amino and/ or C-C alkoxy; or a —COM group, —COOM group, CHCOOM group, CONR'R'' group, NR R'R'' 0098 United States Patent Application Publication No. group -SO2NR R'R'' group, N=N R group or an 2002/01074.04 by Prien et al. discloses sulfur-containing —S(O), R group; or an O-glycoside or N-glycoside, indirubin derivatives of Formula (XIV), wherein R' and R' whereby the glycosides are selected from the group of mono stand for hydrogen, halogen, hydroxy, nitroso, nitro, C-Co or disaccharides, R is hydrogen, C-Cls alkyl, C-Cls alk alkoxy optionally interrupted by one or more oxygenatoms; enyl, C-C cycloalkyl, or C-C cycloalkenyl that is option C-C alkyl optionally Substituted in one or more places with ally substituted by one or more oxygenatoms, R is aryl that halogen, hydroxy and/or amino: aryl or heteroaryl optionally is optionally Substituted by one or more carboxyl, phospho Substituted in one or more places with halogen, C-C alkyl, ryl, or sulfonate groups, R is hydrogen, C-Cls alkyl that is hydroxy, amino and/or C-C alkoxy; or aralkyl, aryloxy, optionally Substituted in one or more places with carboxy, methylenaryloxy, C-C cycloalkyl, C-C cycloalkenyl or phosphoryl or Sulfonate groups, or an aryl group with one or C-C, methylenecycloalkyl optionally substituted in one or more heteroatoms that is optionally substituted with aralkyl more places with halogen, C-C alkyl, hydroxy, amino and/ or sulfonate, R'' and R'' are the same or different and mean or C-C alkoxy and optionally containing one or more het hydrogen or C-C alkyl, aryl, heteroaryl or acyl that is eroatoms; or hydroxylamino, phosphate, phosphonate, Sul optionally Substituted with hydroxy and/or amino; or C-Cls fate, Sulfonate, Sulfonamide optionally substituted in one or alkyl, C-Cs cycloalkyl or C-C cycloalkenyl that is option US 2016/0243077 A1 Aug. 25, 2016 ally interrupted by one or more oxygenatoms, or R'' and R' together with the nitrogen atom of the amino group forms a (XV) C-Cs cycloalkyl, which can contain one or more additional heteroatoms, M is hydrogen, C-Cs alkyl that is optionally Substituted by one or more hydroxy groups and/or amino groups, or aryl or heteroaryl that is optionally substituted in one or more places with halogen, C-C alkyl or C-C, alkoxy, n is 0, 1, or 2, and at least one of R. R. R. or R is substituted with an S(O),R group, and optical isomers and salts thereof. Examples include 5-methylsulfanylindirubin, 5-methylsulfinylindirubin, 5-methylsulfinyl-3'-hydroxyimi noindirubin, and 5-methylsulfanyl-5'-N-acetylindirubin. 0100 United States Patent Application Publication No. 2005/008.0020 by Eisenbrand et al. discloses hydroxylated (XIV) indirubin derivatives of Formula (XVI) wherein R is a straight-chain or branched chain alkyl group or a straight chain or branched chain alkoxy group each having 1 to 18 carbon atoms, preferably 1 to 4 carbon atoms, and R' repre sents H, a straight-chain or branched chain alkyl group having 1 to 4 carbonatoms or a glucuronide group, and X is Hor OH, including 6-hydroxy-5-methylindirubin and 6,7-dihydroxy 5-methylindirubin and their glucuronide derivatives, i.e. 34. 5-trihydroxy-6-(5-methyl-1H, 1H-2',3 bis-indolyliden-2,3'- dion-6-yl)-tetrahydropyran-2-carboxylic acid and 3,4,5- trihydroxy-6-(7'-hydroxy-5-methyl-1H1'H-2',3 0099 United States Patent Application Publication No. bisindolyliden-2,3'-dion-6-yl)-tetrahydropyran-2-carboxylic 2002/0132792 by Prien et al. discloses aryl-substituted acid. 5-methylindirubin is also recited as having anticancer indirubin derivatives of Formula (XV) in which: R. R. R. properties. and Rare hydrogen, hydroxy, C-C alkoxy, aryloxy, trifluo romethyl, nitro, halogen, cyano or the group —SOR, —SONRR, or -COR: or is aryl or heteroaryl that is (XVI) R OR. optionally substituted by the same or a different component in one or more places with hydroxy, halogen, C-C alkyl, amino, nitro, trifluoromethyl, —O—Si(C-C alkyl), cyano, COOC-C alkyl, CO alkyl, CO aryl, SO(C-C alkyl). SO, aryland/or C-C alkoxy, R is hydrogen or NOR, or R" and R or RandR optionally form another C-C-mem bered ring, R is hydrogen, C-Cls alkyl that is optionally Substituted in one or more places with halogen, hydroxy and/or amino: aryl, heteroaryl or C-Cs cycloalkyl that is optionally Substituted in one or more places with halogen, hydroxy, amino, C-C alkyl and/or C-C alkoxy, RandR 0101 United States Patent Application Publication No. are the same or different and are hydrogen or for C-C alkyl, 2008/0194021 by Mays discloses a number of indirubin ana aryl, heteroaryl or acyl that is optionally Substituted in one or logs and derivatives, including compounds of Formula more places with hydroxy, halogen and/or amino; or C-Cls (XVII) wherein: each X is independently O, S, N OR', alkyl, C-Cs cycloalkyl or C-C cycloalkenyl that is option N(Z), or two groups independently selected from hydrogen, ally interrupted by one or more oxygen atoms, or R or R fluoro, chloro, bromo, iodo, NO, phenyl, and C-C alkyl, together with the nitrogen atom of the amino group form a wherein R' is hydrogen, C-C alkyl, or (C-C alkyl)C C-Cs cycloalkyl which can contain one or more additional (O)—; each Y is independently hydrogen, C-C alkyl, (C- heteroatoms, R is hydrogen or C-C alkyl that is optionally C alkyl)C(O)—, (C-C alkyl)C(O)O phenyl, N(Z)(Z), Substituted in one or more places with hydroxy, halogen, Sulfonyl, phosphonyl, fluoro, chloro, bromo, or iodo; each Z C-C alkoxy, amino, nitro, trifluoromethyl, cyano, COOC is independently hydrogen, C-C alkyl, phenyl, benzyl, or alkyl, CO alkyl, CO, aryl, SO(C-C alkyl) and/or SO both Zgroups together with the nitrogen to which they are aryl; or for aryl or heteroaryl that is optionally substituted in attached form 5-, 6-, or 7-membered heterocycloalkyl; each n one or more places with halogen, C-C alkyl, hydroxy, is independently 0, 1, 2, 3, or 4: each R is independently amino, nitro, trifluoromethyl, cyano, COOC-C alkyl, CO hydrogen, C-C alkyl, (C-C alkyl)-C(O)—, phenyl, ben alkyl, CO aryl, SO(C- alkyl), SO arylamino and/or Zyl, or benzoyl; wherein alkyl is branched or straight-chain, C-C alkoxy, and p is 0, 1, or 2, and whereby at least of one optionally substituted with 1, 2, 3, 4, or 5 hydroxyl, N(Z)(Z), of the two radicals R' and R or the two radicals RandR is C-C alkyl, phenyl, benzyl, fluoro, chloro, bromo, or iodo; an aryl or heteroaryl radical, as well as isomers and salts where any phenyl, benzyl, or benzoyl is optionally substituted thereof, with the following examples: 5-(4-methoxyphenyl)- with 1, 2, 3, 4, or 5 hydroxyl, N(Z)(Z), C-C alkyl, fluoro, indirubin. chloro, bromo, or iodo; or a salt thereof. US 2016/0243077 A1 Aug. 25, 2016 10

coupled derivatives linked by a double bond of the general (XVII) schematic formula Y=Z, in which Y is an azaindole group represented by Formula (XIX) or (XX) and Z is an indole group represented by Formula (XXI) or (XXII), wherein the double bond in “Y—Z” is located between the 3-position of the azaindole group (Y) and the 2- or 3'-position of the indole group (Z), and in which: RandR' independently are hydro gen or the following groups which may be unsubstituted or substituted by 1 to 3 substituents: C-C alkyl, aryl, aralkyl, acyl, aroyl, glycosyl or biosyl protected by acyl, glycosyl or 0102 United States Patent Application Publication No. biosyl; wherein said Substituents are selected from: halogen, 2008/207594 by Mussmann et al. discloses indirubin deriva hydroxyl, C-C alkyl, nitro or amino: R. R. R. R. R. R' tives of Formula (XVIII) wherein: R and Rare indepen and R independently represent H, halogen, hydroxyl, sulf dently hydrogen, C-C alkyl, optionally Substituted, or hydryl, or the following groups which may be unsubstituted —CO C-C alkyl, optionally substituted, wherein the sub or substituted by 1 to 3 substituents: C-C alkyl, nitro, amino, stituents are independently selected from one or more of halo, amido, amide, C-C alkoxy, methylthio, phenyl, phenoxy, —CN, hydroxyl. —O—C-C alkyl, -COOH, -COOC aryl, aralkyl, trifluoromethyl, acyl, aroyl, Sulfonic group, Sul C alkyl, —CONH, —CONHCC-C alkyl), —CON(C-C, famoyl, isocyanate, or alkyl isocyanate; wherein said Sub alkyl), aryl, heteroaryl, or combinations thereof; each Rand stituents are selected from: halogen, hydroxyl, C-C alkyl, Risindependently selected from C-C alkyl, C-C alkenyl, nitro or amino: R. R. R. R. R. RandR independently C-C alkynyl, C-C cycloalkyl, Cs-Coheterocyclyl, aryl represent H, halogen, hydroxyl, Sulfhydryl, or the following with 6 to 10 carbonatoms, heteroaryl with 5 to 10 ring atoms, each optionally substituted with halo, NO. —CN, OR', groups which may be unsubstituted or substituted by 1 to 3 —COOR', or NR'R', where R' and R are independently Substituents: C-C alkyl, nitro, amino, amido, amide, C-C, selected from one or more of halo. —CN, hydroxyl, alkoxy, methylthio, phenyl, phenoxy, aryl, aralkyl, trifluo —O C-C alkyl, -COOH, -COOC-C alkyl, romethyl, acyl, aroyl, Sulfonic group, Sulfamoyl, isocyanate, —CONH2, —CONHCC-C alkyl), —CON(C-C alkyl). or alkyl isocyanate; wherein said Substituents are selected aryl, heteroaryl, or combinations thereof; wherein alkyl, alk from: halogen, hydroxyl, C-C alkyl, nitro or amino; R rep enyl, or alkynyl is optionally substituted with one or more of resents oxygen, sulfur, or selenium, or a NR or NOR group, oxo, halo, NO, CN, OR, COOR', OCOR', wherein R is Hor the following groups which may be unsub NR'R'', NRCOR, NROCOR, NRCONR'R'', stituted or substituted by 1 to 3 substituents: C-C straight SR', SOR', SOR', SONR'R', SONR'R'', chain or branched chain alkyl, aryl, aralkyl, C-C alicyclic -NR'SONR'R'', or combinations thereof, wherein R' and group, acyl, aroyl, Sulfonyl, or phosphoryl, wherein the Sub Rare as defined above; wherein cycloalkyl, heterocyclyl, stituents are selected from halogen, hydroxyl, C-C alkyl, aryl, or heteroaryl is optionally substituted with C-C alkyl, nitro, or amino; (2) compounds of Formulas (XXIII), oxo, halo, NO. —CN, OR, COOR', OCOR', (XXIV), (XXV), and (XXVI) in which: R,R,R,R,R,R, NR'R2, NRCOR2, NROCOR2, NRCONR'R2, R. R. R. and Rare defined as above in compounds of the SR', SOR', SOR', SONR'R', SONR'R'', general schematic formulaY=Z; and (3) compounds of For -NR'SONR'R'', or combinations thereof, wherein R' and mulas (XXIII), (XXIV), (XXV), and (XXVI) in which: R' Rareas defined above; or two R' or two Ricantogetherform and R' independently are hydrogen, C-C alkyl, aryl, a ring; wherein n is 0, 1, 2, or 3, and wherein m is 0, 1, 2, or 3: aralkyl, acyl, aroyl, glycosyl protected by acyl, or glycosyl; or an optical isomer or salt thereof. R. R. R. R. R. R. and R are hydrogen, halogen, hydroxyl, Sulfhydryl, C-C alkyl, amino, amide, C-C, alkoxy, methylthio, phenyl, phenyloxy, aryl, aralkyl, trifluo

(XVIII) romethyl, acyl, aroyl, Sulfonic group, or isocyanate; and R represents oxygen, sulfur, or selenium, or a NR or NOR group, wherein R is Hor the following groups which may be unsubstituted or substituted by 1 to 3 substituents: C-C, straight-chain or branched chain alkyl, aryl, aralkyl, C-C, alicyclic group, acyl, aroyl, Sulfonyl, or phosphoryl, wherein the substituents are selected from halogen, hydroxyl, C-C, alkyl, nitro, or amino. In these compounds, compounds of Formula (XXIII) are 5-azaindirubin derivatives, compounds of Formula (XXIV) are 5-azaindigo derivatives, compounds (0103 United States Patent Application Publication No. of Formula (XXV) are 7-azaindirubin derivatives, and com 2010/0016252 by Keana et al. discloses Mannich base N-ox pounds of Formula (XXVI) are 7-azaindigo derivatives. Spe ides or N-oxide rearrangement prodrugs of a number of cific compounds include: 3'-oximido-7-azaindirubin; 7-aza indirubin derivatives, including indirubin, indirubin-3'- indirubin-3'-oxime ether; 1-methyl-5-azaindirubin; oxime, indirubin-5-sulfonic acid, and 5-chloroindirubin. 1-benzyl-5'-chloro-5-azaindirubin; 1-butyl-5-azaindirubin 0104 United States Patent Application Publication No. 3'-oxime; 1-butyl-5-azaindirubin-3'-oxime O-methyl ether; 2010/0137356 by Cheng et al. discloses N-1-methylisoindigo 1-isopropyl-5-azaisoindigo; 1-methyl-7-azaindirubin; as well as azaindole-indole coupled derivatives. The com 1-benzyl-5'-bromo-7-azaindirubin; 1-butyl-7-azaindirubin pounds of United States Patent Application Publication No. 3'-oxime; 1-butyl-7-azaindirubin-3'-oxime O-methyl ether; 2010/0137356 by Cheng et al. include: (1) azaindole-indole 1-isopropyl-7-azaisoindigo. US 2016/0243077 A1 Aug. 25, 2016

-continued

(XIX) (XXV)

(XXVI)

(XX)

0105 United States Patent Application Publication No. (XXI) 2010/0160318 by Tang et al. discloses 3-pyrrolobicyclo hexylene-2-dihydroindolinone derivatives of Formula (XX VII), wherein: R is selected from the group consisting of hydrogen, halo, alkyl, cycloalkyl, aryl, heteroaryl, heteroali cyclic, hydroxy, alkoxy, —C(O)R’, NRR,-(CH),R', and C(O)NR'R'': R is selected from the group consisting of hydrogen, halo, alkyl, haloalkyl, hydroxyl, alkoxy, haloalkoxy, nitro, cyano, NRR, NRC(O)R. C(O) R", aryl, heteroaryl, -C(O)NR'R'', -S(O).R.R. and (XXII) —S(O).R'; R is selected from the group consisting of hydrogen, alkyl, haloalkyl, hydroxy, alkoxy, —C(O)R’. NRR, aryl, heteroaryl, S(O).RR, NRC(O)R’, NRC(O)OR, and NRC(O)R’: R is selected from the group consisting of hydrogen, halo, alkyl, hydroxy, alkoxy, and NRR: R is selected from the group consisting of hydrogen, alkyl, and C(O)R'': R is selected from the group consisting of hydroxy, alkoxy, aryloxy, N(R')(CH),R', NRR, and N(R7) CH(R') CR'(OH) CH(R) (XXIII) Z; R is selected from the group consisting of hydrogen, hydroxy, alkoxy, and aryloxy; R and Rare each indepen dently selected from the group consisting of hydrogen, alkyl, cycloalkyl, aryl, and heteroaryl, or R and R together with the atoms to which they are attached may form a heteroali cyclic ring: R" is selected from the group consisting of hydroxy, C(O)R7, NRR, and C(O)NR'R'': R'' and R" are each independently selected from the group consist ing of hydrogen, alkyl, and aryl, or R'' and R'' together with the nitrogen atom to which they are attached may form a (XXIV) heteroalicyclic ring; R' is selected from the group consisting of alkyl, aryl, aralkyl, heteroaryl, and heteroaralkyl; R'' is selected from the group consisting of hydroxy, alkoxy, ary loxy, and NRR: R' is selected from the group consisting of hydrogen and alkyl; R' is selected from the group con sisting of hydroxy, NR'R''. —C(O)R’, aryl, heteroaryl, N(O)RR, N(OH)R, and NHC(O)R", wherein R is selected from the group consisting of unsubstituted alkyl, R. R. R', and Rare each independently selected from the group consisting of hydrogen and alkyl, Z is selected from the group consisting of aryl, heteroaryl, and NR'R'; and in and rare each independently an integer from 1 to 4. US 2016/0243077 A1 Aug. 25, 2016 12

Formula (XXXII) wherein: R' and Rare the same or differ (XVII) ent, and are hydrogen, halogen, hydroxy, methylenehydroxy, a straight-chain or branched-chain C-Cs alkyl group, a straight-chain or branched-chain C-C alkyloxy group, a straight-chain or branched-chain C-C methylenealkyloxy group, a C-C-7 cycloalkyl group which can comprise one or more heteroatoms, a Substituted or unsubstituted aryl group which can comprise one or more heteroatoms, a Substituted or unsubstituted aralkyl group which can comprise one or more heteroatoms, a Substituted or unsubstituted aryloxy group which can comprise one or more heteroatoms, a mono-, di-, or trialkylsilyl group having 1 to 6 carbon atoms independently of each other in each instance in the straight-chain or branched-chain alkyl group, a mono-, di-, or triarylsilyl group 0106 United States Patent Application Publication No. with substituted or unsubstituted aryl groups independently 2010/0331327 by Meijeret al. discloses 7-substituted indiru of each other in each instance, a trifluoromethyl group; a bin-3'-oximes of Formula (XXVIII) wherein R is N-OH, —COM group, a —COOM group, or a —CHCOOM group, N-O-alkyl, or N O CO-alkyl, NO—(R)-Het, or wherein M is hydrogen, a straight-chain or branched-chain N—O—(Y), NRR, N O CO. N(R. R.) radical C-Cls alkyl group which can additionally carry one or more with Het representing an aliphatic nitrogeneous heterocycle, hydroxyl and/or amino groups, or an aryl group which can Y is an optionally substituted—CH radical, n1 is 1 to 3, X comprise one or more heteroatoms and can be substituted is a halogenatom selected from the group consisting of F, Cl, with one or more halogenatoms, one or more alkyl groups, or Br, or I, and Z is H or methyl, and the salts thereof. one or more alkoxy groups; a NR'R'' group, wherein R' and R'' are the same or different and are hydrogen, a straight chain or branched-chain C-C alkyl group which can addi

(XVIII) tionally carry one or more hydroxyl and/or amino groups, a Substituted or unsubstituted aryl group which can comprise one or more heteroatoms, or an acyl group; —CHNR'R'', wherein R'' and R'' are as defined above; a benzyl group, wherein the benzene nucleus can comprise one or more het eroatoms; a C-C methylenecycloalkyl group which can comprise one or more heteroatoms; a physiological amino acid residue bound to the nitrogen as an amide; an O-glyco side oran N-glycoside, wherein the glycoside is selected from monosaccharides and disaccharides; or a methylenesulfonate group; R. R. R. R. R. R. Rand R' can be the same or different and are hydrogen, halogen, hydroxy, nitroso, nitro, 0107 United States Patent Application Publication No. alkoxy, a straight-chain or branched-chain C-Cs alkyl group 2011/0136808 by Meijer et al. discloses 3',6-substituted which can additionally carry one or more hydroxyl and/or indirubins of Formula (XXIX) wherein R is -(A)-R' or amino groups, a Substituted or unsubstituted aryl group which —CO N (R,R) wherein A is a C-C alkylene group, can comprise one or more heteroatoms; a Substituted or optionally substituted by one or several A' radicals, wherein unsubstituted aralkyl group which can comprise one or more A' is Br, OH, OR, or NH, wherein R is C-C alkyl, -R' heteroatoms; a substituted or unsubstituted aryloxy group is halogen, OH, or N(R,R) wherein R and Rare identical which can comprise one or more heteroatoms; a Substituted or or different and are C-C alkyl optionally substituted by A' unsubstituted methylenearyloxy group which can comprise as defined above, or R and Rare part of a cyclic structure one or more heteroatoms; a C-C, cycloalkyl group which can with 5 or 6 members optionally comprising another heteroa comprise one or more heteroatoms; a C-C methylenecy tom such as 0 or N, and n is 1 to 5. cloalkyl group which can comprise one or more heteroatoms; a trifluoromethyl group; a COM group, a —COOM group, or a —CHCOOM group, wherein M is hydrogen, a straight (XXIX) chain or branched-chain C-C alkyl group which can addi tionally carry one or more hydroxyl and/or amino groups, or an aryl group which can comprise one or more heteroatoms and which can be substituted with one or more halogenatoms, one or more alkyl groups, or one or more alkoxy groups; a - NR'R'' group, wherein R'' and R'' are the same or dif ferent and are hydrogen, a straight-chain or branched-chain C-C alkyl group which can additionally carry one or more hydroxyl and/or amino groups, a Substituted or unsubstituted aryl group which can comprise one or more heteroatoms, or an acyl group, or wherein the nitrogenatom is part of a C-C, 0108 PCT Patent Application Publication No. WO 1999/ cycloalkyl group which can comprise one or more heteroat 062503 by Eisenbrand et al. discloses indigoid bisindole oms; a CONR'R'' group, wherein R'' and R'' are as derivatives of either Formula (XXX). Formula (XXXI), or defined above; a hydroxylamino group; a phosphate group; a US 2016/0243077 A1 Aug. 25, 2016

phosphonate group; a Sulfate group; a Sulfonate group; a reference, discloses 5-nitroindirubin-3-oxime, 5'-bromo-5- sulfonamide group; a SONR'R' group, wherein R'' and nitroindirubin-3'-oxime, 5'-hydroxy-5-nitroindirubin-3'- R" are as defined above; an azo group - N=NR', wherein oxime, 5'-hydroxy-5-chloroindirubin-3-oxime, 5'-hydroxy R" is an aromatic system which can be substituted by one or 5-fluoroindirubin-3'-oxime, 5'-chloro-5-nitroindirubin-3'- more carboxyl groups, phosphoryl groups, or Sulfonate oxime, and 5'-methyl-5-nitroindirubin-3'-oxime. Additional groups; or an O-glycoside or an N-glycoside, wherein the compounds of Formula (XXXIII) are disclosed: glycoside is selected from the group consisting of monosac charides and disaccharides; or RandR, and RandR' can, respectively, form independently from each other a ring hav (XXXIII) ing 1 to 4, optionally Substituted, CH2 groups; and X and Y can be the same or different and can be an oxygen atom, a sulfur atom, a selenium atom, a tellurium atom, a NR'' group in which R' represents a hydrogen atom, a C-Cls straight-chain or branched-chain alkyl group which can be Substituted by one or more carboxyl groups, phosphoryl groups, or Sulfonate groups, a Substituted or unsubstituted aryl group which can comprise one or more heteroatoms, an aralkyl group, or a sulfonate group; or an —NR'' group in which R'' is defined above. The compounds include: 5-iodo indirubin, 5-bromoindirubin, 5-chloroindirubin, 5-fluoroin dirubin, 5-methylindirubin, 5-nitroindirubin, 5'-bromoin wherein: R is selected from the group consisting of F, Cl, dirubin, 5,5'-dibromoindirubin, indirubin-3'-oxime, OH, and CH, and R is selected from the group consisting of indirubin-5-Sulfonic acid, and isoindigo. The compounds can F, Cl, and NO. be covalently bound to a polyethylene glycol ester or a poly ethylene glycol ether. 0111 United States Patent Application Publication No. 2012/0101042 by Duffield et al., incorporated herein by this reference, discloses indirubin-5-sulfonic acid (2-hydroxy (XXX) ethyl)-amide. 0112 United States Patent Application Publication No. 2012/0053208 by Liet al., incorporated herein by this refer ence, discloses (3-3-(3,4-dihydroxybutoxyamino)-1H-in dol-2-ylindol-2-one). This compound is compound E804 from a publication, S. Nam et al., “Indirubin Derivatives Inhibit Stat Signaling and Induce Apoptosis in Human Cancer Cells.” Proc. Natl. Acad. Sci. 102: 5998-6003 (2005), incor porated herein by this reference, which discloses compounds of Formula (XXXIV): (XXXI)

(XXXIV)

(XXXII)

1N1 'N-1)son

0113 with the following alternatives for Ro and R: (i) 0109 U.S. Pat. No. 8,349,822 to Yu, incorporated herein Ro is R is H by this reference, discloses several indirubin derivatives as GSK-3 antagonists and Wnt canonical pathway agonists, including 6-bromoindirubin-3'-oxime and 1-methyl-6-bro moindirubin-3'-oxime. 0110 United States Patent Application Publication No. -x" 2012/0295948 by Kim et al., incorporated herein by this

US 2016/0243077 A1 Aug. 25, 2016 20 salts, Solvates, analogues, congeners, bioisosteres, hydrolysis molecule has the characteristics of aromaticity, including the products, metabolites, precursors, and prodrugs thereof (re delocalized electron distribution that is characteristic of aro ferred to herein as “Alternatives (1)–(486)). maticity. Typically the ring systems contain 5 to 12 ring 0121. As described above, and as detailed more generally member atoms and up to four heteroatoms, wherein the het below, derivatives and analogs of meisoindigo can be option eroatoms are selected from the group consisting of N, O, and ally Substituted with one or more groups that do not Substan S. Frequently, the monocyclic heteroaryls contain 5 to 6 ring tially affect the pharmacological activity of the derivative or members and up to three heteroatoms selected from the group analog. These groups are generally known in the art. Defini consisting of N, O, and S; frequently, the bicyclic heteroaryls tions for a number of common groups that can be used as contain 8 to 10 ring members and up to four heteroatoms optional substituents are provided below; however, the omis selected from the group consisting of N, O, and S. The num sion of any group from these definitions cannot be taken to ber and placement of heteroatoms in heteroaryl ring struc mean that such a group cannot be used as an optional Sub tures is in accordance with the well-known limitations of stituent as long as the chemical and pharmacological require aromaticity and stability, where stability requires the het ments for an optional Substituent are satisfied. eroaromatic group to be stable enough to be exposed to water at physiological temperatures without rapid degradation. As 0122. As used herein, the term “alkyl refers to an used herein, the term “hydroxyheteroaryl” refers to a het unbranched, branched, or cyclic Saturated hydrocarbyl resi eroaryl group including one or more hydroxyl groups as due, or a combination thereof, of from 1 to 12 carbon atoms that can be optionally substituted; the alkyl residues contain substituents; as further detailed below, further substituents only C and H when unsubstituted. Typically, the unbranched can be optionally included. As used herein, the terms or branched saturated hydrocarbyl residue is from 1 to 6 “haloaryl' and “haloheteroaryl refer to aryl and heteroaryl carbon atoms, which is referred to herein as “lower alkyl.” groups, respectively, Substituted with at least one halo group, When the alkyl residue is cyclic and includes a ring, it is where “halo' refers to a halogen selected from the group understood that the hydrocarbyl residue includes at least three consisting of fluorine, chlorine, bromine, and iodine, typi carbon atoms, which is the minimum number to form a ring. cally, the halogen is selected from the group consisting of As used herein, the term “alkenyl refers to an unbranched, chlorine, bromine, and iodine; as detailed below, further sub branched or cyclic hydrocarbyl residue having one or more stituents can be optionally included. As used herein, the terms carbon-carbon double bonds. As used herein, the term “alky “haloalkyl,” “haloalkenyl, and “haloalkynyl refer to alkyl, nyl refers to an unbranched, branched, or cyclic hydrocarbyl alkenyl, and alkynyl groups, respectively, Substituted with at residue having one or more carbon-carbon triple bonds; the least one halo group, where "halo” refers to a halogen residue can also include one or more double bonds. With selected from the group consisting of fluorine, chlorine, bro respect to the use of “alkenyl' or “alkynyl.” the presence of mine, and iodine, typically, the halogen is selected from the multiple double bonds cannot produce an aromatic ring. As group consisting of chlorine, bromine, and iodine; as detailed used herein, the terms “hydroxyalkyl,” “hydroxyalkenyl. below, further substituents can be optionally included. When and “hydroxyalkynyl respectively, refer to an alkyl, alkenyl, a range of values is listed, such as for the number of carbon or alkynyl group including one or more hydroxyl groups as atoms in an alkyl group, it is intended to encompass each substituents; as detailed below, further substituents can be value and Subrange within the range. For example, "C-C, optionally included. As used herein, the term “aryl' refers to alkyl includes alkyl groups with 1, 2, 3, 4, 5, or 6 carbon a monocyclic or fused bicyclic moiety having the well-known atoms and all possible Subranges. characteristics of aromaticity; examples include phenyl and I0123. As used herein, the term "heterocycle.” “heterocy naphthyl, which can be optionally Substituted. As used herein, clyl.” “heterocyclic ring or "heterocyclic group' is intended the term "hydroxyaryl” refers to an aryl group including one to mean a stable 4-, 5-, 6-, or 7-membered monocyclic or 7 or more hydroxyl groups as Substituents; as further detailed 8-, 9-, 10-, 11-, 12-, 13-, or 14-membered bicyclic heterocy below, further substituents can be optionally included. As clic ring which is Saturated, partially unsaturated or fully used herein, the term "heteroaryl refers to monocyclic or unsaturated or aromatic, and which consists of carbon atoms fused bicylic ring systems that have the characteristics of and 1, 2, 3 or 4 heteroatoms independently selected from N. aromaticity and include one or more heteroatoms selected O, and S; and including any bicyclic group in which any of the from O, S, and N. The inclusion of a heteroatom permits above-defined heterocyclic rings is fused to a benzene ring. aromaticity in 5-membered rings as well as in 6-membered The nitrogen and Sulfur heteroatoms may optionally be oxi rings. Typical heteroaromatic systems include monocyclic dized. The nitrogenatom may be substituted or unsubstituted Cs-C heteroaromatic groups such as pyridyl, pyrimidyl, (i.e., N or NR wherein R is H or another substituent, if pyrazinyl, thienyl, furanyl, pyrrolyl pyrazolyl, thiazolyl, defined). The heterocyclic ring may be attached to its pendant oxazolyl, triazolyl, triazinyl, tetrazolyl, tetrazinyl, and imida group at any heteroatom or carbonatom that results in a stable Zolyl, as well as the fused bicyclic moieties formed by fusing structure. The heterocyclic rings described herein may be one of these monocyclic heteroaromatic groups with a phenyl Substituted on carbon or on a nitrogen atom if the resulting ring or with any of the heteroaromatic monocyclic groups to compound is stable. A nitrogen in the heterocycle may form a Cs-Co. bicyclic group Such as indolyl, benzimida optionally be quaternized. It is preferred that when the total Zolyl, indazolyl, benzotriazolyl, isoquinolyl, quinolyl, ben number of S and O atoms in the heterocycle exceeds 1, then Zothiazolyl, benzofuranyl, pyrazolylpyridyl, quinazolinyl, these heteroatoms are not adjacent to one another. When the quinoxalinyl, cinnolinyl, and other ring systems known in the term "heterocycle.” “heterocyclyl. “heterocyclic ring or art. Any monocyclic or fused ring bicyclic system that has the "heterocyclic group' is used, it is intended to include het characteristics of aromaticity interms of delocalized electron eroaryl unless heteroaryl is excluded. Examples of hetero distribution throughout the ring system is included in this cycles include, but are not limited to, acridinyl, azocinyl, definition. This definition also includes bicyclic groups where benzimidazolyl, benzofuranyl, benzothiofuranyl, benzox at least the ring that is directly attached to the remainder of the azolyl, benzoxazolinyl, benzthiazolyl, benztriazolyl, benztet US 2016/0243077 A1 Aug. 25, 2016 razolyl, benzisoxazolyl, benzisothiazolyl, benzimidazolinyl, NZ°C(O)O, NZ'C(O)OZ’, NZ'C(S)OZ, NZ'C carbazolyl, carbolinyl, chromanyl, chromenyl, cinnolinyl, (O)NZZ, NZC(NZ)Z, NZPC(NZ)NZZ, wherein decahydroquinolinyl, 2H,6H-1.5.2-dithiazinyl, dihydrofuro Z is selected from the group consisting of alkyl, cycloalkyl, 2,3-btetrahydrofuran, furanyl, furazanyl, imidazolidinyl, heteroalkyl, cycloheteroalkyl, aryl, arylalkyl, heteroaryland imidazolinyl, imidazolyl, 1H-indazolyl, indolenyl, indolinyl, heteroarylalkyl; each Z” is independently hydrogen or Z", and indolizinyl, indolyl, 3H-indolyl, isatinoyl, isobenzofuranyl, each Z is independently Zor, alternatively, the two Zi's may isochromanyl, isolindazolyl, isoindolinyl, isoindolyl, iso be taken together with the nitrogen atom to which they are quinolinyl, isothiazolyl, isothiazolopyridinyl, isoxazolyl, bonded to form a 4-, 5-, 6-, or 7-membered cycloheteroalkyl isoxazolopyridinyl, methylenedioxyphenyl, morpholinyl, ring structure which may optionally include from 1 to 4 of the naphthyridinyl, octahydroisoquinolinyl, oxadiazolyl, 1.2.3- same or different heteroatoms selected from the group con oxadiazolyl, 1,2,4-oxadiazolyl, 1,2,5-oxadiazolyl, 1,3,4-oxa sisting of N, O, and S. As specific examples, —NZZ is diazolyl, oxazolidinyl, oxazolyl, OXindolyl pyrimidinyl, meant to include NH, -NH-alkyl, - N-pyrrolidinyl, and phenanthridinyl, phenanthrolinyl, phenazinyl, phenothiazi —N-morpholinyl, but is not limited to those specific alterna nyl, phenoxathinyl, phenoxazinyl, phthalazinyl, piperazinyl, tives and includes other alternatives known in the art. Simi piperidinyl, piperidonyl, 4-piperidonyl, piperonyl, pteridinyl, larly, as another specific example, a Substituted alkyl is meant purinyl, pyranyl, pyrazinyl, pyrazolidinyl, pyrazolinyl, pyra to include -alkylene-O-alkyl, -alkylene-heteroaryl, -alky Zolyl pyridazinyl, pyridooxazole, pyridoimidazole, pyridot lene-cycloheteroaryl, -alkylene-C(O)OZ, -alkylene-C(O) hiazole, pyridinyl, pyrimidinyl, pyrrolidinyl, pyrrolinyl, NZZ”, and —CH, CH, C(O)—CH, but is not limited to 2-pyrrolidonyl, 2H-pyrrolyl pyrrolyl, quinazolinyl, quinoli those specific alternatives and includes other alternatives nyl, 4H-quinolizinyl, quinoxalinyl, quinuclidinyl, tetrahy known in the art. The one or more Substituent groups, together drofuranyl, tetrahydroisoquinolinyl, tetrahydroquinolinyl, with the atoms to which they are bonded, may form a cyclic tetrazolyl, 6H-1,2,5-thiadiazinyl, 1,2,3-thiadiazolyl, 1,2,4- ring, including, but not limited to, cycloalkyl and cyclohet thiadiazolyl, 1,2,5-thiadiazolyl, 1,3,4-thiadiazolyl, thianthre eroalkyl. nyl, thiazolyl, thienyl, thienothiazolyl, thienooxazolyl, 0.126 Similarly, substituent groups useful for substituting thienoimidazolyl, thiophenyl, triazinyl, 1,2,3-triazolyl, 1,2,4- unsaturated carbon atoms in the specified group, moiety, or triazolyl, 1,2,5-triazolyl, 1,3,4-triazolyl, and Xanthenyl. Also radical include, but are not limited to, —Z', halo, —O, included are fused ring and Spiro compounds containing, for —OZ, -SZ, -S, NZZ, trihalomethyl, -CF, CN, example, the above heterocycles. 0124. As used herein, the term “optionally substituted” indicates that the particular group or groups referred to as optionally Substituted may have no non-hydrogen Substitu ents, or the group or groups may have one or more non hydrogen Substituents consistent with the chemistry and pharmacological activity of the resulting molecule and Such that a stable compound is formed thereby, i.e., a compound that does not spontaneously undergo transformation Such as by rearrangement, cyclization, elimination, hydrolysis, or I0127. Similarly, substituent groups useful for substituting other reaction. If not otherwise specified, the total number of nitrogen atoms in heteroalkyl and cycloheteroalkyl groups Such Substituents that may be present is equal to the total include, but are not limited to, -Z", halo, O, OZ’, number of hydrogenatoms present on the unsubstituted form —SZ, -S, NZZ, trihalomethyl, -CF, —CN, of the group being described; fewer than the maximum num OCN, -SCN, NO, NO, S(O)Z, -S(O)O, ber of such substituents may be present. Where an optional —S(O)OZ’, —OS(O)OZ’, —OS(O)O, -P(O)(O), Substituent is attached via a double bond. Such as a carbonyl - P(O)(OZ)(O), P(O)(OZ)(OZ), C(O)Z, C(S) oxygen (C=O), the group takes up two available valences on Z, C(NZ)Z, C(O)OZ’, C(S)OZ, C(O)NZZ, the carbon atom to which the optional substituent is attached, C(NZ)NZZ, OC(O)Z, OC(S)Z, OC(O)OZ’, so the total number of substituents that may be included is OC(S)OZ, NZ'C(O)Z, NZ'C(S)Z, NZ'C(O) reduced according to the number of available valiences. As OZ, NZC(S)OZ, NZPC(O)NZZ, NZ'C(NZ)Z, used herein, the term “substituted, whether used as part of and - NZ'C(NZ)NZZ, wherein Z, Z, and Z are as “optionally substituted” or otherwise, when used to modify a defined above. specific group, moiety, or radical, means that one or more I0128. The compounds described herein may contain one hydrogen atoms are, each, independently of each other, or more chiral centers and/or double bonds and therefore, replaced with the same or different substituent or substitu may exist as stereoisomers, such as double-bond isomers entS. (i.e., geometric isomers such as E and Z), enantiomers or 0.125 Substituent groups useful for substituting saturated diastereomers. The invention includes each of the isolated carbon atoms in the specified group, moiety, or radical Stereoisomeric forms (such as the enantiomerically pure iso include, but are not limited to, -Z", =O, OZ’, SZ, mers, the E and Z isomers, and other stereoisomeric forms) as =S, NZZ, =NZ", =N-OZ’, trihalomethyl, -CF, well as mixtures of stereoisomers in varying degrees of chiral purity or percentage of E and Z, including racemic mixtures, mixtures of diastereomers, and mixtures of E and Z isomers. Accordingly, the chemical structures depicted herein encom pass all possible enantiomers and stereoisomers of the illus trated compounds including the stereoisomerically pure form (e.g., geometrically pure, enantiomerically pure or diastereo merically pure) and enantiomeric and stereoisomeric mix US 2016/0243077 A1 Aug. 25, 2016 22 tures. Enantiomeric and stereoisomeric mixtures can be of the crystalline solid. The term “solvate” encompasses both resolved into their component enantiomers or stereoisomers solution-phase and isolatable solvates. Suitable solvates in using separation techniques or chiral synthesis techniques which the solvent is other than water include, but are not well known to the skilled artisan. Such separation techniques limited to, ethanolates or methanolates. When water is the include, but are not limited to, chiral high pressure liquid Solvent, the corresponding Solvate is a “hydrate.” Examples chromatography (HPLC) and formation and crystallization of of hydrates include, but are not limited to, hemihydrate, chiral salts. The invention includes each of the isolated stere monohydrate, dihydrate, trihydrate, hexahydrate, and other oisomeric forms as well as mixtures of stereoisomers in vary hydrated forms. It should be understood by one of ordinary ing degrees of chiral purity, including racemic mixtures. It skill in the art that the pharmaceutically acceptable salt and/or also encompasses the various diastereomers. Other structures prodrug of the present compound may also exist in a Solvate may appear to depict a specific isomer, but that is merely for form. When the solvate is a hydrate, the hydrate is typically convenience, and is not intended to limit the invention to the formed via hydration which is either part of the preparation of depicted olefin isomer. When the chemical name does not the present compound or through natural absorption of mois specify the isomeric form of the compound, it denotes any ture by the anhydrous compound of the present invention. one of the possible isomeric forms or mixtures of those iso Additionally, compounds may exist as clathrates or other meric forms of the compound. complexes, which are therapeutic agent-host inclusion com 0129. The compounds may also exist in several tautomeric plexes wherein the therapeutic agent and the host are present forms, and the depiction herein of one tautomer is for conve in Stoichiometric or non-stoichiometric amounts. nience only, and is also understood to encompass other tau 0.132. As used herein, the term “ester” means any ester of tomers of the form shown. Accordingly, the chemical struc a present compound in which any of the COON functions tures depicted herein encompass all possible tautomeric of the molecule is replaced by a —COOR function, in which forms of the illustrated compounds. The term “tautomer as the R moiety of the ester is any carbon-containing group used herein includes two or more interconvertible compounds which forms a stable ester moiety, including but not limited to resulting from at least one formal migration of a hydrogen alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkylalkyl, aryl, ary atom and at least one change in Valency (e.g., a single bond to lalkyl, heterocyclyl, heterocyclylalkyl and substituted deriva a double bond, a triple bond to a double bond, a triple bond to tives thereof. The hydrolyzable esters of the present com a single bond, or vice versa). The exact ratio of the tautomers pounds are the compounds whose carboxyls are present in the present can depend on several factors, including pH, solvent, form of hydrolyzable ester groups. That is, these esters are and temperature. Tautomerization reactions can be catalyzed pharmaceutically acceptable and can be hydrolyzed to the by acid or base. Examples of tautomerization include keto/ corresponding carboxyl acid in vivo. enol, amidefimide, lactam/lactim, and enaminefimine. 0133. In addition to the substituents described above, 0130. As used herein, the terms “nucleic acid.” “oligo alkyl, alkenyl and alkynyl groups can alternatively or in addi nucleotide.” “polynucleotide.” and similar terms typically tion be substituted by C-C acyl, C-C heteroacyl, C-Co refer to polymers of deoxyribonucleotides (DNA) or ribo aryl, C-Cs cycloalkyl, C-Cs heterocyclyl, or Cs-Co het nucleotides (RNA) in either single- or double-stranded form, eroaryl, each of which can be optionally substituted. Also, in and complements thereof according to the conventional Wat addition, when two groups capable of forming a ring having son-Crick base-pairing scheme unless the complementary 5 to 8 ring members are present on the same or adjacent sequence is excluded. Unless specifically excluded, the terms atoms, the two groups can optionally be taken together with also refer to nucleic acids containing known nucleotide ana the atom or atoms in the Substituent groups to which they are logs or modified backbone residues or linkages, which are attached to form Such a ring. synthetic, naturally-occurring, or non-naturally-occurring, I0134) “Heteroalkyl,” “heteroalkenyl,” and “heteroalky which have similar binding properties to a corresponding nyl' and the like are defined similarly to the corresponding nucleic acid comprising naturally-occurring conventional hydrocarbyl (alkyl, alkenyl and alkynyl) groups, but the het bases (adenine, guanine, cytosine, and thymine (for DNA) or ero terms refer to groups that contain 1-3 O.S or N heteroa uracil (for RNA), and which typically are metabolized in a toms or combinations thereof within the backbone residue; manner similar to a corresponding nucleic acid comprising thus at least one carbon atom of a corresponding alkyl, alk naturally-occurring conventional bases; however, in some enyl, or alkynyl group is replaced by one of the specified contexts, the requirement for metabolism in a manner similar heteroatoms to form, respectively, a heteroalkyl, heteroalk to a corresponding nucleic acid comprising naturally-occur enyl, or heteroalkynyl group. For reasons of chemical stabil ring conventional bases can be dispensed with. Examples of ity, it is also understood that, unless otherwise specified. Such Such analogs include, but are not necessarily limited to, phos groups do not include more than two contiguous heteroatoms phorothioates, phosphoramidates, methyl phosphonates, except where an oxo group is present on NorSas in a nitro or chiral methyl phosphonates, 2-O-methyl ribonucleotides, Sulfonyl group. and peptide nucleic acids (PNAs). I0135 While “alkyl as used herein includes cycloalkyl 0131. As used herein, the term “solvate” means a com and cycloalkylalkyl groups, the term "cycloalkyl may be pound formed by solvation (the combination of solvent mol used hereinto describe a carbocyclic non-aromatic group that ecules with molecules or ions of the Solute), or an aggregate is connected via a ring carbon atom, and “cycloalkylalkyl that consists of a solute ion or molecule, i.e., a compound of may be used to describe a carbocyclic non-aromatic group the invention, with one or more solvent molecules. The term that is connected to the molecule through an alkyl linker. 'solvate” typically means a physical association of a com 0.136. Similarly, "heterocyclyl may be used to describe a pound involving varying degrees of ionic and/or covalent non-aromatic cyclic group that contains at least one heteroa bonding, including hydrogen bonding. In certain instances tom (typically selected from N, O and S) as a ring member and the solvate will be capable of isolation, for example when one that is connected to the molecule via a ring atom, which may or more solvent atoms are incorporated into the crystal lattice be C (carbon-linked) or N (nitrogen-linked); and "heterocy US 2016/0243077 A1 Aug. 25, 2016 clylalkyl may be used to describe Such a group that is con stituents optionally present on the aryl or heteroaryl portion nected to another molecule through a linker. The heterocyclyl are the same as those described above for aryl groups gener can be fully saturated or partially saturated, but non-aromatic. ally. The sizes and substituents that are suitable for the cycloalkyl, 0141 'Arylalkyl groups as used herein are hydrocarbyl cycloalkylalkyl, heterocyclyl, and heterocyclylalkyl groups groups if they are unsubstituted, and are described by the total are the same as those described above for alkyl groups. The number of carbon atoms in the ring and alkylene or similar heterocyclyl groups typically contain 1, 2 or 3 heteroatoms, linker. Thus a benzyl group is a C-arylalkyl group, and selected from N, O and S as ring members; and the Nor S can phenylethyl is a C-arylalkyl. be substituted with the groups commonly found on these 0.142 “Heteroarylalkyl as described above refers to a atoms in heterocyclic systems. As used herein, these terms moiety comprising an aryl group that is attached through a also include rings that contain a double bond or two, as long linking group, and differs from “arylalkyl in that at least one as the ring that is attached is not aromatic. The Substituted ring atom of the aryl moiety or one atom in the linking group cycloalkyl and heterocyclyl groups also include cycloalkyl or is a heteroatom selected from N, O and S. The heteroarylalkyl heterocyclic rings fused to an aromatic ring or heteroaromatic groups are described herein according to the total number of ring, provided the point of attachment of the group is to the atoms in the ring and linker combined, and they include aryl cycloalkyl or heterocyclyl ring rather than to the aromatic/ groups linked through a heteroalkyl linker, heteroaryl groups heteroaromatic ring. linked through a hydrocarbyl linker Such as an alkylene; and 0.137 As used herein, “acyl encompasses groups com heteroaryl groups linked through a heteroalkyl linker. Thus, prising an alkyl, alkenyl, alkynyl, aryl or arylalkyl radical for example, C7-heteroarylalkyl would include pyridylm attached at one of the two available valence positions of a ethyl, phenoxy, and N-pyrrolylmethoxy. carbonyl carbon atom, and heteroacyl refers to the corre 0.143 “Alkylene' as used herein refers to a divalent hydro sponding groups wherein at least one carbon other than the carbyl group; because it is divalent, it can link two other carbonyl carbon has been replaced by a heteroatom chosen groups together. Typically it refers to —(CH), where n is from N, O and S. 1-8 and preferably n is 1-4, though where specified, an alky 0138 Acyl and heteroacyl groups are bonded to any group lene can also be substituted by other groups, and can be of or molecule to which they are attached through the open other lengths, and the open Valences need not be at opposite valence of the carbonyl carbon atom. Typically, they are ends of a chain. The general term “alkylene' encompasses C-C acyl groups, which include formyl, acetyl, pivaloyl, more specific examples Such as “ethylene wherein n is 2. and benzoyl, and C-Cs heteroacyl groups, which include “propylene,” wherein n is 3, and “butylene,” wherein n is 4. methoxyacetyl, ethoxycarbonyl, and 4-pyridinoyl. The hydrocarbyl groups of the alkylene can be optionally 0139 Similarly, “arylalkyl and “heteroarylalkyl refer to substituted as described above. aromatic and heteroaromatic ring systems which are bonded 0144. In general, any alkyl, alkenyl, alkynyl, acyl, or aryl to their attachment point through a linking group Such as an orarylalkyl group that is contained in a Substituent may itself alkylene, including Substituted or unsubstituted, Saturated or optionally be substituted by additional substituents. The unsaturated, cyclic or acyclic linkers. Typically the linker is nature of these substituents is similar to those recited with C-Cs alkyl. These linkers may also include a carbonyl group, regard to the primary substituents themselves if the substitu thus making them able to provide Substituents as an acyl or ents are not otherwise described. heteroacyl moiety. An aryl or heteroaryl ring in an arylalkyl or 0145 Amino” as used herein refers to —NH, but where heteroarylalkyl group may be substituted with the same sub an amino is described as “substituted' or “optionally substi stituents described above for aryl groups. Preferably, an ary tuted’, the term includes NR'R'" wherein each R' and R" is lalkyl group includes a phenyl ring optionally Substituted independently H, or is an alkyl, alkenyl, alkynyl, acyl, aryl, or with the groups defined above for aryl groups and a C-C, arylalkyl group, and each of the alkyl, alkenyl, alkynyl, acyl, alkylene that is unsubstituted or is substituted with one or two aryl, or arylalkyl groups is optionally substituted with the C-C alkyl groups or heteroalkyl groups, where the alkyl or substituents described herein as suitable for the correspond heteroalkyl groups can optionally cyclize to form a ring Such ing group; the R' and R" groups and the nitrogen atom to as cyclopropane, dioxolane, or oxacyclopentane. Similarly, a which they are attached can optionally form a 3- to 8-mem heteroarylalkyl group preferably includes a C-C monocy bered ring which may be saturated, unsaturated or aromatic clic heteroaryl group that is optionally substituted with the and which contains 1-3 heteroatoms independently selected groups described above as Substituents typical on aryl groups from N, O and S as ring members, and which is optionally and a C-C alkylene that is unsubstituted or is Substituted substituted with the substituents described as suitable for with one or two C-C alkyl groups or heteroalkyl groups, or alkyl groups or, if NR'R" is an aromatic group, it is optionally it includes an optionally substituted phenyl ring or Cs-C substituted with the substituents described as typical for het monocyclic heteroaryl and a C-C heteroalkylene that is eroaryl groups. unsubstituted or is substituted with one or two C-C alkyl or 0146. As used herein, the term “carbocycle “carbocy heteroalkyl groups, where the alkyl or heteroalkyl groups can clyl or “carbocyclic” refers to a cyclic ring containing only optionally cyclize to form a ring Such as cyclopropane, diox carbon atoms in the ring, whereas the term "heterocycle' or olane, or oxacyclopentane. "heterocyclic” refers to a ring comprising a heteroatom. The 0140. Where an arylalkyl or heteroarylalkyl group is carbocyclyl can be fully saturated or partially saturated, but described as optionally substituted, the substituents may be non-aromatic. For example, the general term "carbocyclyl on either the alkyl or heteroalkyl portion or on the aryl or encompasses cycloalkyl. The carbocyclic and heterocyclic heteroaryl portion of the group. The substituents optionally structures encompass compounds having monocyclic, bicy present on the alkyl or heteroalkyl portion are the same as clic or multiple ring systems; and Such systems may mix those described above for alkyl groups generally; the Sub aromatic, heterocyclic, and carbocyclic rings. Mixed ring US 2016/0243077 A1 Aug. 25, 2016 24 systems are described according to the ring that is attached to to the molecule. Similarly, the term “aryloxyalkylcarbonyl the rest of the compound being described. refers to an ester Substituent including an alkyl group wherein 0147 As used herein, the term "heteroatom” refers to any the alkyl group is itself Substituted by an aryloxy group. atom that is not carbon or hydrogen, Such as nitrogen, oxygen 0157. Other combinations of substituents are known in the or sulfur. When it is part of the backbone or skeleton of a chain art and, are described, for example, in U.S. Pat. No. 8.344,162 or ring, a heteroatom must be at least divalent, and will typi to Jung et al., incorporated herein by this reference. For cally be selected from N, O, P, and S, more typically from. example, the term “thiocarbonyl and combinations of sub 0148. As used herein, the term "alkanoyl refers to an stituents including “thiocarbonyl include a carbonyl group alkyl group covalently linked to a carbonyl (C=O) group. in which a double-bonded sulfur replaces the normal double The term “lower alkanoyl refers to an alkanoyl group in bonded oxygen in the group. The term “alkylidene' and simi which the alkyl portion of the alkanoyl group is C-C. The lar terminology refer to an alkyl group, alkenyl group, alkynyl alkyl portion of the alkanoyl group can be optionally Substi group, or cycloalkyl group, as specified, that has two hydro tuted as described above. The term “alkylcarbonyl can alter gen atoms removed from a single carbon atom so that the natively be used. Similarly, the terms “alkenylcarbonyl and group is double-bonded to the remainder of the structure. “alkynylcarbonyl refer to an alkenyl or alkynyl group, 0158 Accordingly, methods and compositions according respectively, linked to a carbonyl group. to the present invention encompass meisoindigo derivatives 0149. As used herein, the term “alkoxy' refers to an alkyl and analogs including one or more optional Substituents as group covalently linked to an oxygen atom; the alkyl group defined above, provided that the optionally substituted can be considered as replacing the hydrogen atom of a meisoindigo derivative or analog possesses Substantially hydroxyl group. The term “lower alkoxy” refers to an alkoxy equivalent pharmacological activity to the meisoindigo group in which the alkyl portion of the alkoxy group is C-C. derivative or analog as defined in terms of cell cycle arrest or The alkyl portion of the alkoxy group can be optionally Sub apoptosis enhancement. Methods for determining cell cycle stituted as described above. As used herein, the term arrest are known in the art and are described in K. Polyak et “haloalkoxy' refers to an alkoxy group in which the alkyl al., “p27Kip1, a Cyclin-Cdk Inhibitor, Links Transforming portion is Substituted with one or more halo groups. Growth Factor-Beta and Contact Inhibition to Cell Cycle 0150. As used herein, the term "sulfo’ refers to a sulfonic Arrest. Genes & Develop. 8: 9-22 (1994), incorporated acid ( -SOH) substituent. herein by this reference. Methods for determining enhance 0151. As used herein, the term "sulfamoyl refers to a ment of apoptosis are known in the art and are described in M. substituent with the structure—S(O)NH2, wherein the nitro Faber et al., “Overexpression of the Rabies Virus Glycopro gen of the NH2 portion of the group can be optionally Substi tein Results in Enhancement of Apoptosis and Antiviral tuted as described above. Immune Response.” J. Virol. 76:3374-3381 (2002), incorpo 0152. As used herein, the term “carboxyl refers to a group rated herein by this reference. of the structure —C(O)H. 0159. A number of indirubin derivatives have the activity 0153. As used herein, the term “carbamyl refers to a of glycogen synthase kinase-3 antagonists or Wntagonists, as group of the structure —C(O)NH2, wherein the nitrogen of described in U.S. Pat. No. 8,349,822 to Yu, incorporated the NH, portion of the group can be optionally substituted as herein by this reference. The Wnt signaling pathway is described above. described in C. Y. Logan & R. Nusse, “The Wnt Signaling 0154 As used herein, the terms “monoalkylaminoalkyl Pathway in Development and Disease.” Annu. Rev. Cell. Dev. and “dialkylaminoalkyl refer to groups of the structure Biol. 20: 781-810 (2004), incorporated herein by this refer -Alk-NH-Alk, and -Alk-N(Alk)(Alk3), wherein Alk, ence. The Wnt pathway involves a diverse family of Wnt Alk, and Alks refer to alkyl groups as described above. glycoproteins that act as ligands to regulate the production of (O155 As used herein, the term “alkylsulfonyl refers to a intracellular signaling molecules to produce a cellular group of the structure —S(O)-Alk wherein Alk refers to an response. These Wnt glycoprotein ligands include several alkyl group as described above. The terms “alkenylsulfonyl different proteins such as WNT1, WNT2, WNT2B, WNT3, and “alkynylsulfonyl refer analogously to Sulfonyl groups WNT3A, WNT4, WNT5A, WNTSB, WNT6, WNT7A, covalently bound to alkenyl and alkynyl groups, respectively. WNT7B, WNT8A, WNT8B, WNT9A, WNT9B, WNT10A, The term “arylsulfonyl refers to a group of the structure WNT10B, WNT11, and WNT16. There are several variations —S(O), Ar wherein Ar refers to an aryl group as described of the Wnt pathway, namely the canonical Wnt pathway, the above. The term “aryloxyalkylsulfonyl refers to a group of noncanonical Planar Cell Polarity pathway, and the nonca the structure —S(O)-Alk-O Ar, where Alk is an alkyl nonical Wnt/calcium pathway. The canonical pathway, the group as described above and Aris an aryl group as described best known, causes an accumulation off-catenin in the cyto above. The term “arylalkylsulfonyl refers to a group of the plasm and the eventual translocation of the B-catenin to the structure —S(O)-AlkAr, where Alk is an alkyl group as nucleus to act as a transcriptional coactivator of transcription described above and Ar is an aryl group as described above. factors that belong to the TCF/LEF family. GSK-3 is a kinase 0156. As used herein, the term “alkyloxycarbonyl refers that phosphorylates serine and threonine amino acid residues to an ester Substituent including an alkyl group wherein the in glycogen synthase and other proteins. GSK-3 phosphory carbonyl carbon is the point of attachment to the molecule. An lates -catenin, thus tying GSK-3 to the Wnt pathway; phos example is ethoxycarbonyl, which is CHCHOC(O)—. phorylated B-catenin is targeted for degradation. The activity Similarly, the terms “alkenyloxycarbonyl.” “alkynyloxycar of GSK-3 is described in R. S. Jope, “The Glamour and bonyl, and “cycloalkylcarbonyl refer to similar ester sub Gloom of Glycogen Synthase Kinase-3, ' Trends Biochem. stituents including an alkenyl group, alkenyl group, or Sci. 29: 95-102 (2004), incorporated herein by this reference. cycloalkyl group respectively. Similarly, the term “aryloxy Indirubin and some indirubin derivatives also have the activ carbonyl refers to an ester Substituent including an aryl ity of inhibiting cyclin-dependent protein kinase-2 (CDK2). group wherein the carbonyl carbon is the point of attachment CDK2 is a member of the cyclin-dependent family of serine/ US 2016/0243077 A1 Aug. 25, 2016

threonine protein kinase and regulates the cell cycle. The (0162 (II) Dose Modification activity of CDK2 is described in J. Du et al., “Critical Role of 0163 Improvements for suboptimal chemotherapeutics CDK2 for Melanoma Growth Linked to Its Melanocyte-Spe including indirubin or analogs and derivatives thereof, cific Transcriptional Regulation by MITF Cancer Cell: 6: including meisoindigo, are made by alterations to the time 565-576 (2004), incorporated herein by this reference. that the compound is administered, the use of dose-modifying Accordingly, within the scope of the present invention are agents that control the rate of metabolism of the compound, indirubin derivatives and analogs that are GSK-3 antagonists. normal tissue protective agents, and other alterations. General Also within the scope of the present invention are indirubin examples include: variations of infusion schedules (e.g., derivatives and analogs that are Wntagonists. Additionally, bolus i.V. Versus continuous infusion), the use of lymphokines within the scope of the present invention are indirubin deriva (e.g., G-CSF, GM-CSF, EPO) to increase leukocyte count for tives and analogs that are CDK2 inhibitors. improved immune response or for preventing anemia caused 0160 Also within the scope of the present invention are by myelosuppressive agents, or the use of rescue agents. pharmaceutically acceptable salts of indirubin analogs and Specific inventive examples for indirubin or analogs and derivatives as described above, including pharmaceutically derivatives thereof, including meisoindigo, include: continu acceptable salts of meisoindigo. "Pharmaceutically accept ous i.v. infusion for hours to days; biweekly administration; able salt” refers to conventional acid-addition or base-addi doses greater than 5 mg/m/day; progressive escalation of tion salts that retain the biological effectiveness and proper dosing from 1 mg/m/day based on patient tolerance; doses ties of the indirubin analogs and derivatives described above less than 1 mg/m for greater than 14 days; use of caffeine to and are formed from Suitable non-toxic organic or inorganic modulate metabolism; use ofisoniazid to modulate metabo acids or organic or inorganic bases. All of these salts may be lism; selected and intermittent boost dose administrations; prepared by conventional means from the corresponding bolus single and multiple doses escalating from 5 mg/m, oral compound of the invention by reacting, for example, the doses below 30 or above 130 mg/m; chronic low dose admin appropriate acid or base with the compound of the invention. istration of from about 10 mg/day to about 25 mg/day; inter When a basic group and an acid group are present in the same mittent administration of from about 50 mg to about 150 mg molecule, a compound of the invention may also form inter twice weekly or three times weekly; administration of from nal salts. Accordingly, when the indirubin analog or deriva about 50 mg/day to about 150 mg/day for 10-14 days per tive is negatively charged, pharmaceutically acceptable salts month; or chronic daily dosing at a dose of equal to or greater can be formed with the following positively-charged ions: than 100 mg/day. Sodium, potassium, aluminum, lithium, calcium, magnesium, (0164 (III) Route of Administration Zinc, ammonium, caffeine, arginine, diethylamine, N-eth 0.165 Improvements for suboptimal chemotherapeutics ylpiperidine, histidine, glucamine, isopropylamine, lysine, including indirubin or analogs and derivatives thereof, morpholine, N-ethylmorpholine, piperazine, piperidine, tri including meisoindigo, are made by alterations in the route by ethylamine, trimethylamine, ethanolamine, diethanolamine, which the compound is administered. General examples N-methylglucamine, and tris(hydroxymethyl)ami include: changing route from oral to intravenous administra nomethane. When the indirubin analog or derivative is posi tion and vice versa; or the use of specialized routes Such as tively charged, pharmaceutically acceptable salts can be Subcutaneous, intramuscular, intraarterial, intraperitoneal, formed with the following negatively charged ions: chloride, intralesional, intralymphatic, intratumoral, intrathecal, intra bromide, iodide, carbonate, nitrate, Sulfate, bisulfate, phos vesicular, intracranial. Specific inventive examples for phate, monohydrogen phosphate, dihydrogen phosphate, indirubin or analogs and derivatives thereof, including metaphosphate, pyrophosphate, formate, acetate, adipate, meisoindigo, include: topical; intravesicular for bladder can butyrate, propionate. Succinate, glycolate, gluconate, lactate, cer, oral administration; slow release oral delivery; intrathe malate, tartrate, citrate, ascorbate, glucuronate, maleate, cal; intraarterial; continuous infusion; or intermittent infu fumarate, pyruvate, aspartate, glutamate, benzoate, anthra S1O. nilate, mesylate, 4'-hydroxybenzoate, phenylacetate, mande 0166 (IV) Schedule of Administration late, embonate (pamoate), methanesulfonate, ethane 0.167 Improvements for suboptimal chemotherapeutics Sulfonate, ethanedisulfonate, benzenesulfonate, including indirubin or analogs and derivatives thereof, pantothenate, 2-hydroxyethanesulfonate, p-toluene including meisoindigo, are made by alterations to the time Sulfonate, Sulfanilate, cyclohexylaminosulfonate, camphor that the compound is administered. General examples ate, camphorsulfonate, digluconate, cyclopentanepropionate, include: changing from a monthly administration to a weekly dodecylsulfonate, glucoheptanoate, glycerophosphonate, or daily dosing or variations of the schedule. Specific inven heptanoate, hexanoate, 2-hydroxyethanesulfonate, nicoti tive examples for indirubin or analogs and derivatives thereof, nate, isonicotinate, 1-naphthalenesulfonate, 2-naphthalene including meisoindigo, include: daily; weekly for three Sulfonate, oxalate, palmoate, pectinate, persulfurate, 2-phe weeks, weekly for two weeks, biweekly; biweekly for three nylpropionate, picrate, pivalate, thiocyanate, mesylate, weeks with a 1-2 week rest period; intermittent boost dose undecanoate, Stearate, algenate, 3-hydroxybutyrate, salicy administration; daily for one week then once per week for late, galactarate, galacturonate, caprylate, isobutyrate, mal multiple weeks. onate, Suberate, sebacate, chlorobenzoate, methylbenzoate, (0168 (V) Indications for Use dinitrobenzoate, phthalate, phenylacetate, isethionate, lacto 0169. Improvements for suboptimal chemotherapeutics bionate, p-aminobenzoate, Sulfamate, diethylacetate, pime including indirubin or analogs and derivatives thereof, late, aminosulfonate, acrylate, Y-hydroxybutyrate, and meth including meisoindigo, are made by alterations in the types of oxybenzoate. disease or the clinical stage of disease that the compound is 0161 Prodrugs of indirubin and analogs and derivatives administered. General examples include: the use of solid thereof, including prodrugs of meisoindigo, are described tumor agents for leukemias and vice versa, the use of antitu below. mor agents for the treatment of benign hyperproliferative US 2016/0243077 A1 Aug. 25, 2016 26 disease such as psoriasis or benign prostate hypertrophy. Spe ornithine decarboxylase; patients with disease conditions cific inventive examples for indirubin or analogs and deriva with low levels of metabolic enzymes, histone deacetylase, tives thereof, including meisoindigo, include: use for the protein kinases, ornithine decarboxylase; patients with low or treatment of leukemias (acute and chronic, ALL, CLL, CML, high Susceptibility to thrombocytopenia, neutropenia; CLL); use for the treatment of acute promyelocytic leukemia patients intolerant of GI toxicities; over- or under-expression (APL); use for the treatment of myelodysplastic syndrome of jun, GPCRs and signal transduction proteins, VEGF, pros (MDS); use for the treatment of T-cell lymphomas; use for the tate specific genes, protein kinases, or telomerase. treatment of B-cell lymphomas; use for the treatment of (0176 (IX) Patient/Disease Phenotype mantle cell lymphomas; use for the Suppression of prolifera 0177 Improvements for suboptimal chemotherapeutics tion of cancer stem cells; use for the treatment of malignant including indirubin or analogs and derivatives thereof, pericardial effusions; use for the treatment of ovarian carci including meisoindigo, are made by more precise identifica noma; use for the treatment of carcinoma of the lung; use for tion of a patient’s ability to tolerate, metabolize and exploit the treatment of angiogenic diseases; use for the treatment of the use of the compound. General examples include: use of benign prostate hypertrophy; use for the treatment of psoria diagnostic tools and kits to better characterize a patients sis; use for the treatment of gout; use for the treatment of ability to process/metabolize a chemotherapeutic agent or autoimmune conditions; use for the treatment of rheumatoid their susceptibility to toxicity caused by potential specialized arthritis; use for treatment of insulin-resistant diabetes; use cellular, metabolic, or organ system phenotypes. Specific for prevention of transplantation rejection; use for restenosis inventive examples for indirubin or analogs and derivatives prevention in cardiovascular disease; use for the treatment of thereof, including meisoindigo, include: diagnostic tools, mycosis fungoides; use in bone marrow transplantation; use techniques, kits and assays to confirm a patient's particular as an antiinfective; use for the treatment of AIDS; or use for phenotype and for the measurement of metabolism enzymes the treatment of Alzheimer's disease, including disease pre and metabolites, histone deacetylase, protein kinases, orni vention, slowing of disease progression, and amelioration or thine decarboxylase, VEGF, prostate specific genes, protein reversal of disease symptoms, due to the effect of meisoin kinases, telomerase, jun GPCRs; or Surrogate compound digo on the phosphorylation of t (tau) protein. dosing or low dose drug pre-testing for enzymatic status. (0170 (VI) Disease Stages (0178 (X) Patient/Disease Genotype 0171 Improvements for suboptimal chemotherapeutics 0179 Improvements for suboptimal chemotherapeutics including indirubin or analogs and derivatives thereof, including indirubin or analogs and derivatives thereof, including meisoindigo, are made by alterations in the stage of including meisoindigo, are made by testing and analyzing a disease at diagnosis/progression that the compound is admin patient’s genotype for unique features that may be of value to istered. General examples include: the use of chemotherapy predict efficacy, toxicity, metabolism, or other factors affect for non-resectable local disease, prophylactic use to prevent ing the therapeutic efficacy of the drug. General examples metastatic spread or inhibit disease progression or conversion include: biopsy samples of tumors or normal tissues (e.g., to more malignant stages. Specific inventive examples for white blood cells) that may also be taken and analyzed to indirubin or analogs and derivatives thereof, including specifically tailor or monitor the use of a particular drug meisoindigo, include: use for the treatment of localized polyp againstagene target; studies of unique tumor gene expression stage colon cancer; leukoplakia in the oral cavity; angiogen patterns; or analysis of SNPs (single nucleotide polymor esis inhibition to prevent or limit metastatic spread; against phisms), to enhance efficacy or to avoid particular drug-sen HIV with AZT, DDI, reverse transcriptase inhibitors. sitive normal tissue toxicities. Specific inventive examples for (0172 (VII) Other Indications indirubin or analogs and derivatives thereof, including 0173 Improvements for suboptimal chemotherapeutics meisoindigo, include: diagnostic tools, techniques, kits and including indirubin or analogs and derivatives thereof, assays to confirm a patient’s particular genotype; gene/pro including meisoindigo, are made by using the compound for tein expression chips and analysis. Single Nucleotide Poly non-malignant diseases and conditions. General examples morphisms (SNPs) assessment; SNPs for histone deacety include: premalignant conditions, benign hyperproliferative lase, ornithine decarboxylase, GPCRs, protein kinases, conditions, treatment of infections, parasites, usage to relieve telomerase, jun: or identification and measurement of pain, control of pleural effusions. Specific inventive examples metabolism enzymes and metabolites. for indirubin or analogs and derivatives thereof, including 0180 (XI) Pre/Post-Treatment Preparation meisoindigo, include: use as antiinfectives, antivirals; anti 0181 Improvements for suboptimal chemotherapeutics bacterials; for pleural effusions; antifungals; antiparasitics; including indirubin or analogs thereof, including meisoin eczema: shingles; condylomata; anti HPV; or anti HSV. digo, are made by specialized preparation of a patient prior to (0174 (VIII) Patient Selection or after the use of a chemotherapeutic agent. General 0175 Improvements for suboptimal chemotherapeutics examples include: induction or inhibition of metabolizing including indirubin or analogs and derivatives thereof, enzymes, specific protection of sensitive normal tissues or including meisoindigo, are made by alterations to the type of organ systems. Specific inventive examples for indirubin or patient that would best tolerate or benefit from the use of the analogs and derivatives thereof, including meisoindigo, compound. General examples include: use of pediatric doses include: the use of colchicine or analogs; use of diuretics Such for elderly patients, altered doses for obese patients; exploi as probenecid; use of uricase; non-oral use of nicotinamide; tation of co-morbid disease conditions such as diabetes, cir sustained release forms of nicotinamide; use of inhibitors of rhosis, or other conditions that may uniquely exploit a feature polyADP ribose polymerase; use of caffeine; leucovorin res of the compound. Specific inventive examples for indirubin or cue; infection control; or antihypertensives. analogs and derivatives thereof, including meisoindigo, 0182 (XII) Toxicity Management include: patients with disease conditions with high levels of 0183 Improvements for suboptimal chemotherapeutics metabolic enzymes, histone deacetylase, protein kinases, including indirubin or analogs and derivatives thereof, US 2016/0243077 A1 Aug. 25, 2016 27 including meisoindigo, are made by use of additional drugs or with dasatinib; use with lonidamine; use with 5-azacytidine: procedures to prevent or reduce potential side-effects or tox use with thalidomide or analogs thereof; use with EGFR icities. General examples include: the use of anti-emetics, inhibitors such as erlotinib, afatinib, lapatinib, or dacomi anti-nausea, hematological Support agents to limit or prevent tinib, use with gold salts such as aurothiomalate or aurothio neutropenia, anemia, thrombocytopenia, Vitamins, antide glucose; use with dibromodulcitol; use with dianhydrogalac pressants, treatments for sexual dysfunction, and other Sup titol; use with decitabine; or use with bortezomib or other portive techniques. Specific inventive examples for indirubin proteasome inhibitors. One particularly significant use is use or analogs and derivatives thereof, including meisoindigo, with Nek9 inhibitors or agents that inhibit the expression of include: the use of colchicine or analogs; use of diuretics Such Nek9, addressed below. as probenecid; use of uricase; non-oral use of nicotinamide; 0188 (XV) Chemosensitization sustained release forms of nicotinamide; use of inhibitors of 0189 Improvements for suboptimal chemotherapeutics poly ADP-ribose polymerase; use of caffeine; leucovorin res including indirubin or analogs and derivatives thereof, cue; use of Sustained release allopurinol; non-oral use of including meisoindigo, are made by exploiting them as allopurinol; bone marrow transplant stimulants, blood, plate chemosensitizers where no measureable activity is observed let infusions, Neupogen, G-CSF; GM-CSF; pain manage when used alone but in combination with other therapeutics a ment; anti-inflammatories; fluids; corticosteroids; insulin more than additive or synergistic improvement in efficacy is control medications; anti-pyretics; anti-nausea treatments; observed. General examples include: misonidazole with anti-diarrhea treatment, N-acetyl cysteine; or antihistamines. alkylating agents, or tirapazamine with cisplatin. Specific 0184 (XIII) Pharmacokinetic/Pharmacodynamic Moni inventive examples for indirubin or analogs and derivatives toring thereof, including meisoindigo, include: as a chemosensitizer 0185. Improvements for suboptimal chemotherapeutics in combination with topoisomerase inhibitors; use with including indirubin or analogs and derivatives thereof are fraudulent nucleosides; use with fraudulent nucleotides; use made by the use of monitoring drug levels after dosing in an with thymidylate synthetase inhibitors; use with signal trans effort to maximize a patient's drug plasma level, to monitor duction inhibitors; use with cisplatin or platinum analogs; use the generation of toxic metabolites, monitoring of ancillary with alkylating agents such as BCNU Gliadel wafers, CCNU, medicines that could be beneficial or harmful in terms of bendamustine (Treanda) Temozoloimide (Temodar); use drug-drug interactions. General examples include: the moni with anti-tubulin agents; use with antimetabolites; use with toring of drug plasma protein binding, and monitoring of berberine; use with apigenin; use with amonafide; use with other pharmacokinetic or pharmacodynamic variables. Spe colchicine or analogs thereof use with genistein; use with cific inventive examples for indirubin or analogs and deriva etoposide; use with cytarabine; use with camptothecins; use tives thereof include: multiple determinations of drug plasma with vinca alkaloids; use with topoisomerase inhibitors; use levels; or multiple determinations of metabolites in the blood with 5-fluorouracil; use with curcumin; use with NF-KB or urine. inhibitors; use with rosmarinic acid; use with mitoguaZone; 0186 (XIV) Drug Combinations or use with tetrandrine. 0187 Improvements for suboptimal chemotherapeutics (0190 (XVI) Chemopotentiation including indirubin or analogs and derivatives thereof, 0191 Improvements for suboptimal chemotherapeutics including meisoindigo, are made by exploiting unique drug including indirubin or analogs and derivatives thereof, combinations that may provide a more than additive or Syn including meisoindigo, are made by exploiting them as ergistic improvement in efficacy or side-effect management. chemopotentiators where minimal therapeutic activity is General examples include: alkylating agents with anti-me observed alone but in combination with other therapeutics tabolites, topoisomerase inhibitors with antitubulin agents. unique drug a more than additive or synergistic improvement Specific inventive examples for indirubin or analogs and in efficacy is observed. General examples include: amonafide derivatives thereof, including meisoindigo, include: use with with cisplatin or 5-FU. Specific inventive examples for topoisomerase inhibitors; use with fraudulent nucleosides; indirubin or analogs and derivatives thereof, including use with fraudulent nucleotides; use with thymidylate syn meisoindigo, include: as a chemopotentiator in combination thetase inhibitors; use with signal transduction inhibitors; use with topoisomerase inhibitors; use with fraudulent nucleo with cisplatin or platinum analogs such as Oxaliplatin, satra sides; use with thymidylate synthetase inhibitors; use with platin, or carboplatin; use with alkylating agents such as the signal transduction inhibitors; use with cisplatin or platinum nitrosoureas (BCNU, Gliadel wafers, CCNU) bendamustine analogs; use with alkylating agents such as BCNU, BCNU (Treanda); use with anti-tubulin agents; use with antimetabo wafers, Gliadel, bendamustine (Treanda); use with anti-tubu lites; use with berberine; use with apigenin; use with amona lin agents; use with antimetabolites; use with berberine; use fide; use with colchicine and analogs; use with genistein; use with apigenin; use with amonafide; use with colchicine or with etoposide; use with cytarabine; use with camptothecins; analogs thereof, use with genistein; use with etoposide; use use with vinca alkaloids; use with 5-fluorouracil; use with with cytarabine; use with camptothecins; use with vinca alka curcumin; use with NF-kB inhibitors; use with rosmarinic loids; use with topoisomerase inhibitors; use with 5-fluorou acid; use with mitoguaZone; use with tetrandrine; use with racil; use with curcumin; use with NF-kB inhibitors; use with antineoplastic agents not metabolized by cytochrome P450 roSmarinic acid; use with mitoguaZone; or use with tetran CYP1A2 or CYP2C19; use in combination with biological drine. therapies such as antibodies Such as Avastin, Rituxan, Her (0192 (XVII) Post-Treatment Patient Management ceptin, or Erbitux; use with tyrosine kinase inhibitors; use 0193 Improvements for suboptimal chemotherapeutics with all-trans-retinoic acid; use with arsenicals such as including indirubin or analogs and derivatives thereof, arsenic trioxide; use with hydroxyurea; use with thioguanine; including meisoindigo, are made by drugs, treatments and use with mercaptopurine; use with homoharringtonine; use diagnostics to allow for the maximum benefit to patients with oridonin; use with uracil mustard; use with nilotinibuse treated with a compound. General examples include: pain US 2016/0243077 A1 Aug. 25, 2016 28 management, nutritional Support, anti-emetics, anti-nausea (0202 (XXII) Excipients therapies, anti-anemia therapy, anti-inflammatories. Specific 0203 Improvements for suboptimal chemotherapeutics inventive examples for indirubin or analogs and derivatives including indirubin or analogs and derivatives thereof, thereof, including meisoindigo, include: use with therapies including meisoindigo, are made by alterations in the mate associated with pain management; nutritional Support; anti rials/excipients, buffering agents, or preservatives required to emetics; anti-nausea therapies; anti-anemia therapy; anti-in stabilize and present a chemical compound for proper admin flammatories: antipyretics; or immune stimulants. istration. General examples include: mannitol, albumin, EDTA, sodium bisulfite, benzyl alcohol. Specific inventive (0194 (XVIII) Alternative Medicine/Therapeutic Support examples for indirubin or analogs and derivatives thereof, 0.195 Improvements for suboptimal chemotherapeutics including meisoindigo, include: the use of mannitol; albumin; including indirubin or analogs and derivatives thereof, EDTA; sodium bisulfite; benzyl alcohol; carbonate buffers: including meisoindigo, are made by the use of unapproved/ or phosphate buffers. non-conventional therapeutics or methods to enhance effec (0204 (XXII) Dosage Forms tiveness or reduce side effects. General examples include: 0205 Improvements for suboptimal chemotherapeutics hypnosis, acupuncture, meditation, herbal medications and including indirubin or analogs and derivatives thereof, extracts, applied kinesiology, prayer. Specific inventive including meisoindigo, are made by alterations in the poten examples for indirubin or analogs and derivatives thereof, tial dosage forms of the compound dependent on the route of including meisoindigo, include: hypnosis; acupuncture; administration, duration of effect, plasma levels required, meditation; herbal medications created either synthetically or exposure to side-effect normal tissues and metabolizing through extraction including NF-kB inhibitors (such as par enzymes. General examples include: tablets, capsules, topi thenolide, curcumin, rosmarinic acid); natural anti-inflam cal gels, creams, patches, Suppositories. Specific inventive matories (including rhein, parthenolide); immunostimulants examples for indirubin or analogs and derivatives thereof, (such as those found in Echinacea); antimicrobials (such as including meisoindigo, include: the use of tablets; capsules; berberine); flavonoids, isoflavones, and flavones (such as api topical gels; topical creams; patches; Suppositories; or lyo genenin, genistein, genistin, 6"-O-malonylgenistin, 6"-O- philized dosage fills. acetylgenistin, daidzein, daidzin, 6"-O-malonyldaidzin, 0206 (XXIV) Dosage Kits and Packaging 6"-O-acetylgenistin, glycitein, glycitin, 6"-O-malonylgly 0207 Improvements for suboptimal chemotherapeutics citin, and 6-O-acetylglycitin); or applied kinesiology. including indirubin or analogs and derivatives thereof, (0196 (XIX) Bulk Drug Product Improvements including meisoindigo, are made by alterations in the dosage forms, container/closure systems, accuracy of mixing and 0197) Improvements for suboptimal chemotherapeutics dosage preparation and presentation. General examples including indirubin or analogs and derivatives thereof, include: amber vials to protect from light, stoppers with spe including meisoindigo, are made by alterations in the phar cialized coatings. Specific inventive examples for indirubin maceutical bulk Substance. General examples include: salt or analogs and derivatives thereof, including meisoindigo, formation, homogeneous crystalline structure, pure isomers. include: the use of amber vials to protect from light; or stop Specific inventive examples for indirubin or analogs and pers with specialized coatings to improve shelf-life stability. derivatives thereof, including meisoindigo, include: salt for (0208 (XXV) Drug Delivery Systems mation; homogeneous crystalline structure; pure isomers; 0209 Improvements for suboptimal chemotherapeutics increased purity; or lower residual solvents and heavy metals. including indirubin or analogs and derivatives thereof, (0198 (XX) Diluent Systems including meisoindigo, are made by the use of delivery sys 0199 Improvements for suboptimal chemotherapeutics tems to improve the potential attributes of a pharmaceutical including indirubin or analogs and derivatives thereof, product Such as convenience, duration of effect, reduction of including meisoindigo, are made by alterations in the diluents toxicities. General examples include: nanocrystals, bioerod used to solubilize and deliver/present the compound for ible polymers, liposomes, slow release injectable gels, micro administration. General examples include: Cremophor-EL, spheres. Specific inventive examples for indirubin or analogs cyclodextrins for poorly water soluble compounds. Specific and derivatives thereof, including meisoindigo, include: the inventive examples for indirubin or analogs and derivatives use of nanocrystals; bioerodible polymers; liposomes; slow thereof, including meisoindigo, include: use of emulsions; release injectable gels; or microspheres. dimethyl sulfoxide (DMSO); N-methylformamide (NMF): 0210 (XXVI) Drug Conjugate Forms dimethylformamide (DMF); dimethylacetamide (DMA); 0211 Improvements for suboptimal chemotherapeutics ethanol; benzyl alcohol; dextrose containing water for injec including indirubin or analogs and derivatives thereof, tion; Cremophor, cyclodextrins; or polyethylene glycol including meisoindigo, are made by alterations to the parent molecule with covalent, ionic, or hydrogen bonded moieties (PEG). to alter the efficacy, toxicity, pharmacokinetics, metabolism, (0200 (XXI) Solvent Systems or route of administration. General examples include: poly 0201 Improvements for suboptimal chemotherapeutics mer systems such as polyethylene glycols, polylactides, including indirubin or analogs and derivatives thereof, polyglycolides, amino acids, peptides, or multivalent linkers. including meisoindigo, are made by alterations in the solvents Specific inventive examples for indirubin or analogs and used or required to solubilize a compound for administration derivatives thereof, including meisoindigo, include: the use or for further dilution. General examples include: ethanol, of polymer systems such as polyethylene glycols; polylac dimethylacetamide (DMA). Specific inventive examples for tides; polyglycolides; amino acids; peptides; or multivalent indirubin or analogs and derivatives thereof, including linkers. meisoindigo, include: the use of emulsions; DMSO; NMF; 0212 (XXVII) Compound Analogs DMF, DMA; ethanol; benzyl alcohol; dextrose containing 0213 Improvements for suboptimal chemotherapeutics water for injection; Cremophor; or PEG. including indirubin or analogs and derivatives thereof, US 2016/0243077 A1 Aug. 25, 2016 29 including meisoindigo, are made by alterations to the parent 0220 (XXXI) Biotherapeutic Resistance Modulation structure of a molecule with additional chemical functional 0221) Improvements for suboptimal chemotherapeutics ities that may alter efficacy, or reduce toxicity, pharmacologi including indirubin or analogs and derivatives thereof, cal performance, route of administration, or another relevant including meisoindigo, are made by exploiting their selective factor for therapeutic efficacy. General examples include: use to overcome developing or complete resistance to the alteration of side chains to increase or decrease lipophilicity, efficient use of biotherapeutics. General examples include: additional chemical functionalities to alter reactivity, electron tumors resistant to the effects of biological response modifi affinity, binding capacity, Salt forms. Specific inventive ers, cytokines, lymphokines, therapeutic antibodies, anti examples for indirubin or analogs and derivatives thereof, sense therapies, gene therapies. Specific inventive examples including meisoindigo, include: alteration of side chains to for indirubin or analogs and derivatives thereof, including increase or decrease lipophilicity; additional chemical func meisoindigo, include: the use against tumors resistant to the tionalities to alter reactivity; electron affinity; binding capac effects of biological response modifiers; cytokines; lymphok ity; or salt forms. The additional chemical functionalities to ines; therapeutic antibodies; antisense therapies; therapies alter reactivity can include, but are not necessarily limited to, Such as Avastin, Rituxan, Herceptin, ErbituX; gene therapies; alkylating functionalities or binding sites for colchicine. ribozymes; or RNA interference. 0214 (XXVIII) Prodrugs 0222 (XXXII) Radiation Therapy Enhancement 0215 Improvements for suboptimal chemotherapeutics including indirubin or analogs and derivatives thereof, 0223 Improvements for suboptimal chemotherapeutics including meisoindigo, are made by alterations to the mol including indirubin or analogs and derivatives thereof, ecule Such that improved pharmaceutical performance is including meisoindigo, are made by exploiting their use in gained with a variant of the active molecule in that after combination with ionizing radiation, phototherapies, heat introduction into the body a portion of the molecule is cleaved therapies, or radio-frequency generated therapies. General to reveal the preferred active molecule. General examples examples include: hypoxic cell sensitizers, radiation sensitiz include: enzyme sensitive esters, dimers, Schiff bases. Spe ers/protectors, photosensitizers, radiation repair inhibitors. cific inventive examples for indirubin or analogs and deriva Specific inventive examples for indirubin or analogs and tives thereof, including meisoindigo, include: the use of derivatives thereof, including meisoindigo, include: the use enzyme sensitive esters; the use of dimers; the use of Schiff with hypoxic cell sensitizers; radiation sensitizers/protectors; bases; the use of pyridoxal complexes; the use of caffeine photosensitizers; radiation repair inhibitors; thiol depletion; complexes; the use of N-substituted carbohydrate derivatives: vaso-targeted agents; use with radioactive seeds; use with the use of Mannich N-oxides; the use of products of reaction radionuclides; use with radiolabeled antibodies; or use with with an acylating or carbamylating agent; the use of hex brachytherapy. anoate conjugates; the use of polymer-agent conjugates; or 0224 (XXXIII) Novel Mechanisms of Action the use of prodrugs that are Subject to redox activation. 0225 Improvements for suboptimal chemotherapeutics 0216 (XXIX) Multiple Drug Systems including indirubin or analogs and derivatives thereof, 0217. Improvements for suboptimal chemotherapeutics including meisoindigo, are made by optimizing their utility including indirubin or analogs and derivatives thereof, by determining the various mechanisms of action, biological including meisoindigo, are made by the use of additional targets of a compound for greater understanding and preci compounds, biological agents that when administered in the sion to better exploit the utility of the molecule. General proper fashion, a unique and beneficial effect can be realized. examples include: Gleevec for chronic myelocytic leukemia General examples include: inhibitors of multi-drug resis (CML), arsenic trioxide for acute promyelocytic leukemia tance, specific drug resistance inhibitors, specific inhibitors (APL), retinoic acid for APL. Specific inventive examples for of selective enzymes, signal transduction inhibitors, repair indirubin or analogs and derivatives thereof, including inhibition. Specific inventive examples for indirubin or ana meisoindigo, include: the use with inhibitors of poly-ADP logs and derivatives thereof, including meisoindigo, include: ribose polymerase; agents that effect vasculature; vasodila the use of inhibitors of multi-drug resistance; specific drug tion; oncogenic targeted agents; signal transduction inhibi resistance inhibitors; specific inhibitors of selective enzymes: tors; EGFR inhibition; Protein Kinase Cinhibition: Phospho signal transduction inhibitors; repair inhibition; or topoi lipase C down-regulation; jun down-regulation; histone Somerase inhibitors with non-overlapping side effects. genes; VEGF; ornithine decarboxylase;jun D; V-jun: GPCRs: protein kinase A. telomerase, prostate specific genes; protein 0218 (XXX) Biotherapeutic Enhancement kinases; or histone deacetylase. 0219. Improvements for suboptimal chemotherapeutics including indirubin or analogs and derivatives thereof, 0226 (XXXIV) Selective Target Cell Population Thera including meisoindigo, are made by their use in combination peutics as sensitizers/potentiators with biological response modifi 0227 Improvements for suboptimal chemotherapeutics ers. General examples include: use in combination as sensi including indirubin or analogs and derivatives thereof, tizers/poteniators with biological response modifiers, cytok including meisoindigo, are made by more precise identifica ines, lymphokines, therapeutic antibodies, antisense tion and exposure of the compound to those select cell popu therapies, gene therapies. Specific inventive examples for lations where the compound's effect can be maximally indirubin or analogs and derivatives thereof, including exploited. General examples include: tirapazamine and mito meisoindigo, include: use in combination as sensitizers/po mycin C for hypoxic cells, vinca alkaloids for cells entering tentiators with biological response modifiers; cytokines; lym mitosis. Specific inventive examples for indirubin or analogs phokines; therapeutic antibodies; antisense therapies such as and derivatives thereof, including meisoindigo, include: use Avastin, Herceptin, Rituxan, and ErbituX; gene therapies; against radiation sensitive cells; radiation resistant cells; ribozymes; or RNA interference. energy depleted cells; or endothelial cells. US 2016/0243077 A1 Aug. 25, 2016 30

0228. Additional alternatives specifically applicable to system, including: (1) cancers of the cervix uteri, including indirubin derivatives and analogs, including meisoindigo, are cervical intraepithelial neoplasia (Grade I), cervical intraepi further described below in the description of detailed alterna thelial neoplasia (Grade II), cervical intraepithelial neoplasia tives for these categories. (Grade III) (squamous cell carcinoma in situ), keratinizing 0229. Accordingly, one aspect of the present invention is a squamous cell carcinoma, nonkeratinizing squamous cell car method to improve the efficacy and/or reduce the side effects cinoma, Verrucous carcinoma, adenocarcinoma in situ, of Suboptimally administered drug therapy employing a adenocarcinoma in situ, endocervical type, endometrioid therapeutic agent selected from the group consisting of adenocarcinoma, clear celladenocarcinoma, adenosquamous indirubin and an analog or derivative thereof comprising the steps of: carcinoma, adenoid cystic carcinoma, Small cell carcinoma, 0230 (1) identifying at least one factor or parameter asso and undifferentiated carcinoma; (2) cancers of the corpus ciated with the efficacy and/or occurrence of side effects of uteri, including endometrioid carcinoma, adenocarcinoma, the drug therapy employing atherapeutic agent selected from adenocanthoma (adenocarcinoma with squamous metapla the group consisting of indirubin and an analog or derivative sia), adenosquamous carcinoma (mixed adenocarcinoma and thereof, and squamous cell carcinoma, mucinous adenocarcinoma, serous 0231 (2) modifying the factor or parameter to improve the adenocarcinoma, clear cell adenocarcinoma, squamous cell efficacy and/or reduce the side effects of the drug therapy. adenocarcinoma, and undifferentiated adenocarcinoma; (3) 0232 Typically, the factor or parameter is selected from cancers of the ovary, including serous cystadenoma. Serous the group consisting of cystadenocarcinoma, mucinous cystadenoma, mucinous cys 0233 (1) dose modification; tadenocarcinoma, endometrioid tumor, endometrioid adeno 0234 (2) route of administration: carcinoma, clear cell tumor, clear cell cystadenocarcinoma, 0235 (3) schedule of administration; and unclassified tumor; (4) cancers of the vagina, including 0236 (4) indications for use: squamous cell carcinoma and adenocarcinoma; and (5) can 0237 (5) selection of disease stage; cers of the Vulva, including Vulvar intraepithelial neoplasia 0238 (6) other indications: (Grade I), Vulvar intraepithelial neoplasia (Grade II), Vulvar 0239 (7) patient selection: intraepithelial neoplasia (Grade III) (squamous cell carci 0240 (8) patient/disease phenotype: noma in situ); squamous cell carcinoma, Verrucous carci 0241 (9) patient/disease genotype: noma, Paget’s disease of the Vulva, adenocarcinoma (NOS), 0242 (10) prefpost-treatment preparation basal cell carcinoma (NOS), and Bartholin's gland carci 0243 (11) toxicity management; noma; (C) cancers of the male reproductive system, includ 0244 (12) pharmacokinetic/pharmacodynamic monitor ing: (1) cancers of the penis, including squamous cell carci 1ng noma; (2) cancers of the prostate, including adenocarcinoma, 0245 (13) drug combinations: sarcoma, and transitional cell carcinoma of the prostate; (3) 0246 (14) chemosensitization; cancers of the testis, including seminomatous tumor, non 0247 (15) chemopotentiation: seminomatous tumor, teratoma, embryonal carcinoma, yolk 0248 (16) post-treatment patient management; sac tumor, and Choriocarcinoma, (D) cancers of the cardiac 0249 (17) alternative medicine/therapeutic support; system, including sarcoma (angiosarcoma, fibrosarcoma, 0250 (18) bulk drug product improvements: rhabdomyosarcoma, liposarcoma), myxoma, rhabdomyoma, 0251 (19) diluent systems: fibroma, lipoma and teratoma; (E) cancers of the respiratory 0252 (20) solvent systems; system, including squamous cell carcinoma of the larynx, 0253 (21) excipients: primary pleural mesothelioma, and squamous cell carcinoma 0254 (22) dosage forms: of the pharynx, (F) cancers of the lung, including squamous 0255 (23) dosage kits and packaging; cell carcinoma (epidermoid carcinoma), variants of squa mous cell carcinoma, spindle cell carcinoma, Small cell car 0256 (24) drug delivery systems: cinoma, carcinoma of other cells, carcinoma of intermediate 0257 (25) drug conjugate forms: cell type, combined oat cell carcinoma, adenocarcinoma, aci 0258 (26) compound analogs; nar adenocarcinoma, papillary adenocarcinoma, bronchiolo 0259 (27) prodrugs: alveolar carcinoma, Solid carcinoma with mucus formation, 0260 (28) multiple drug systems; large cell carcinoma, giant cell carcinoma, clear cell carci 0261 (29) biotherapeutic enhancement; noma, and sarcoma; (G) cancers of the gastrointestinal tract, 0262 (30) biotherapeutic resistance modulation; including: (1) cancers of the ampulla of Vater, including pri 0263 (31) radiation therapy enhancement; mary adenocarcinoma, carcinoid tumor, and lymphoma; (2) 0264 (32) novel mechanisms of action; and cancers of the anal canal, including adenocarcinoma, squa 0265 (33) selective target cell population therapeutics. mous cell carcinoma, and melanoma; (3) cancers of the extra 0266 Typically, the hyperproliferative disease is cancer. hepatic bile ducts, including carcinoma in situ, adenocarci Methods according to the present invention are applicable to noma, papillary adenocarcinoma, adenocarcinoma, intestinal many forms of cancer, including, but not limited to: (A) breast type, mucinous adenocarcinoma, clear cell adenocarcinom, cancer, including: (1) ductal carcinoma, including ductal car Segnet-ring cell carcinoma, adenosquamous carcinoma, cinoma in situ (DCIS) (comedocarcinoma, cribriform, papil squamous cell carcinoma, Small cell (oat) carcinoma, undif lary, micropapillary), infiltrating ductal carcinoma (IDC), ferentiated carcinoma, carcinoma (NOS), sarcoma, and car tubular carcinoma, mucinous (colloid) carcinoma, papillary cinoid tumor; (4) cancers of the colon and rectum, including carcinoma, metaplastic carcinoma, and inflammatory carci adenocarcinoma in situ, adenocarcinoma, mucinous adeno noma; (2) lobular carcinoma, including lobular carcinoma in carcinoma (colloid type; greater than 50% mucinous carci situ (LCIS) and invasive lobular carcinoma; and (3) Paget’s noma), signet ring cell carcinoma (greater than 50% signet disease of the nipple; (B) cancers of the female reproductive ring cell), squamous cell (epidermoid) carcinoma, adenos US 2016/0243077 A1 Aug. 25, 2016

quamous carcinoma, Small cell (oat cell) carcinoma, undif domyosarcoma, synovial sarcoma, and sarcoma (NOS); (3) ferentiated carcinoma, carcinoma (NOS), sarcoma, lym cancers of the nervous system, including cancers of the skull phoma, and carcinoid tumor; (5) cancers of the esophagus, (osteoma, hemangioma, granuloma, Xanthoma, osteitis including squamous cell carcinoma, adenocarcinoma, lei deformans), cancers of the meninges (meningioma, menin omyosarcoma, and lymphoma; (6) cancers of the gallbladder, giosarcoma, gliomatosis), cancers of the brain (astrocytoma, including adenocarcinoma, adenocarcinoma, intestinal type, medulloblastoma, glioma, ependymoma, germinoma adenosquamous carcinoma, carcinoma in situ, carcinoma (pilealoma), glioblastoma multiform, oligodendroglioma, (NOS), clear cell adenocarcinoma, mucinous adenocarci Schwannoma, retinoblastoma, congenital tumors), and can noma, papillary adenocarcinoma, signet-ring cell carcinoma, cers of the spinal cord neurofibroma, meningioma, glioma, Small cell (oat cell) carcinoma, squamous cell carcinoma, and sarcoma); (4) hematologic cancers, including myeloid leuke undifferentiated carcinoma; (7) cancers of the lip and oral mia (acute and chronic), acute lymphloblastic leukemia, cavity, including squamous cell carcinoma; (8) cancers of the chronic lymphocytic leukemia, myeloproliferative diseases, liver, including hepatoma (hepatocellular carcinoma), cho multiple myeloma; myelodysplastic syndrome), Hodgkin’s langiocarcinoma, hepatoblastoma, angiosarcoma, hepatocel disease, and non-Hodgkin’s lymphoma (malignant lym lular adenoma, and hemangioma; (9) cancers of the exocrine phoma); (5) cancers of the endocrine system, including: (a) pancreas, including duct cell carcinoma, pleomorphic giant cancers of the thyroid gland, including papillary carcinoma cell carcinoma, giant cell carcinoma, osteoclastoid type, (including those with follicular foci), follicular carcinoma, adenocarcinoma, adenosquamous carcinoma, mucinous (col medullary carcinoma, and undifferentiated (anaplastic) car loid) carcinoma, cystadenocarcinoma, acinar cell carcinoma, cinoma, and (b) neuroblastomas, including sympathicoblas papillary carcinoma, Small cell (oat cell) carcinoma, mixed toma, sympathicogonioma, malignant ganglioneuroma, gan cell typed, carcinoma (NOS), undifferentiated carcinoma, gliosympathicoblastoma, and ganglioneuroma; (6) cancers of endocrine cell tumors arising in the islets of langerhans, and carcinoid; (10) cancers of the salivary glands, including the skin, including squamous cell carcinoma, spindle cell acinic (acinar) cell carcinoma, adenoid cystic carcinoma (cy variant of squamous cell carcinoma, basal cell carcinoma, lindroma), adenocarcinoma, squamous cell carcinoma, car adenocarcinoma developing from Sweat or sebaceous gland, cinoma in pleomorphic adenoma (malignant mixed tumor), and malignant melanoma; (7) cancers of the eye, including: mucoepidermoid carcinoma (well differentiated or low (a) cancers of the conjunctiva, including carcinoma of the grade), and mucoepidermoid carcinoma (poorly differenti conjunctiva; (b) cancers of the eyelid, including basal cell ated or high grade); (11) cancers of the stomach, including carcinoma, squamous cell carcinoma, melanoma of the eye adenocarcinoma, papillary adenocarcinoma, tubular adeno lid, and sebaceous cell carcinoma; (c) cancers of the lacrimal carcinoma, mucinous adenocarcinoma, signet ring cell carci gland, including adenocarcinoma, adenoid cystic carcinoma, noma, adenosquamous carcinoma, squamous cell carcinoma, carcinoma in pleomorphic adenoma, mucoepidermoid carci Small cell carcinoma, undifferentiated carcinoma, lym noma, and squamous cell carcinoma; (d) cancers of the uvea, phoma, sarcoma, and carcinoid tumor, and (12) cancers of the including spindle cell melanoma, mixed cell melanoma, and Small intestine, including adenocarcinoma, lymphoma, car epithelioid cell melanoma; (e) cancers of the orbit, including cinoid tumors, Kaposi's sarcoma, leiomyoma, hemangioma, sarcoma of the orbit, Soft tissue tumor, and sarcoma of bone; lipoma, neurofibroma, and fibroma; (H) cancers of the uri and (f) retinoblastoma; and (1) cancers of unknown primary nary system, including: (1) cancers of the kidney, including origin. In particular, methods according to the present inven renal cell carcinoma, carcinoma of Bellini's collecting ducts, tion and compositions suitable for use according to those adenocarcinoma, papillary carcinoma, tubular carcinoma, methods are applicable to lower grade astrocytomas and other granular cell carcinoma, clear cell carcinoma (hyperneph primary central nervous system tumors besides glioblastoma roma), sarcoma of the kidney, and nephroblastoma; (2) can multiforme (GBM), as well as to central nervous system cers of the renal pelvis and ureter, including transitional cell metastases of other tumors including solid tumors and hema carcinoma, papillary transitional cell carcinoma, squamous tologic tumors (e.g., breast, lung, bladder, and bowel tumors, cell carcinoma, and adenocarcinoma; (3) cancers of the ure leukemias, and lymphomas), in addition to squamous cell thra, including transitional cell carcinoma, squamous cell non-Small cell lung cancer. In addition, methods according to carcinoma, and adenocarcinoma; and (4) cancers of the uri the present invention and compositions Suitable for use nary bladder, including carcinoma in situ, transitional urothe according to those methods are applicable to melanoma, lial cell carcinoma, papillary transitional cell carcinoma, breast lymphoma (both Hodgkins and non-Hodgkins), col squamous cell carcinoma, adenocarcinoma, undifferentiated; orectal cancer, acute lymphoblastic leukemia, and for lower (1) cancers of muscle, bone, and Soft tissue, including: (1) ing the incidence of central nervous system leukemia, non cancers of bone, including: (a) bone-forming: osteosarcoma; Small cell lung cancer, cervical carcinoma, bladder (b) cartilage-forming: chondrosarcoma and mesenchymal carcinoma, and metastatic hemangiopericytoma. In particu chondrosarcoma; (c) diant cell tumor, malignant; (d) Ewings lar, when the therapeutically active compound is meisoin sarcoma; (e) vascular tumors: hemangioendothelioma, digo, methods according to the present invention and compo hemangiopericytoma, and angiosarcoma, (f) connective tis sitions suitable for use according to those methods are Sue tumors: fibrosarcoma, liposarcoma, malignant mesen applicable to acute myelogenous leukemia, chronic myelog chymoma, and undifferentiated sarcoma; and (g) other enous leukemia, and colon cancer. tumors: chordoma and adamantinoma of long bones; (2) can 0267 In one alternative, the therapeutic agent is indirubin, cers of Soft tissues, including: alveolar soft-part sarcoma, an analog thereof, or a derivative of indirubin or an analog angiosarcoma, epithelioid sarcoma, extraskeletal chondrosa thereof. Analogs of indirubin and derivatives of indirubin and rcoma, fibrosarcoma, leiomyosarcoma, liposarcoma, malig analogs of indirubin are described above as Alternatives (1)- nant fibrous histiocytoma, malignant hemangiopericytoma, (486). In particular, methods and compositions according to malignant mesenchymoma, malignant Schwannoma, rhab the present invention are applicable to meisoindigo. US 2016/0243077 A1 Aug. 25, 2016 32

0268. The following improvements all apply either to 0304 (c) use for treatment of T-cell lymphomas; indirubin, analogs of indirubin, or derivatives of indirubin or 0305 (d) use for treatment of B-cell lymphomas; analogs of indirubin, including, but not limited to. Alterna 0306 (e) use for treatment of mantle cell lymphoma; tives (1)–(486), as indicated with respect to the specific 0307 (f) use for suppression of proliferation of cancer improvement described below. Methods according to the stem cells; present invention are particularly applicable to meisoindigo, 0308 (g) use for treatment of ovarian carcinoma; and all methods encompass the use of meisoindigo or a 0309 (h) use for treatment of carcinoma of the lung; derivative thereofas specifically described. 0310 (i) use for treatment of angiogenic diseases: 0269. When the improvement is dose modification, the 0311 () use for treatment of benign prostatic hyperpla dose modification can be, but is not limited to, at least one S1a, dose modification selected from the group consisting of 0312 (k) use for treatment of psoriasis: 0270 (a) continuous i.v. infusion for hours to days; 0313 (1) use for treatment of gout: 0271 (b) biweekly administration; 0314 (m) use for treatment of autoimmune conditions: (0272 (c) doses greater than 5 mg/m/day; 0315 (n) use for treatment of treatment of insulin-resis 0273 (d) progressive escalation of dosing from 1 tant diabetes; mg/m/day based on patient tolerance; 0316 (o) use for treatment of transplantation rejection; 0274 (e) use of caffeine to modulate metabolism; 0317 (p) use for prevention of restenosis in cardiovas 0275 (f) use of isonazid to modulate metabolism; cular disease; 0276 (g) selected and intermittent boosting of dosage 0318 (q) use for treatment of mycosis fungoides: administration; 0319 (r) use in bone marrow transplantation; 0277 (h) administration of single and multiple doses 0320 (s) use as an anti-infective agent; escalating from 5 mg/m/day via bolus; 0321 (t) use for treatment of AIDS; and (0278 (i) oral dosages of below 30 mg/m: 0322 (u) use for treatment of Alzheimer's disease, (0279 (j) oral dosages of above 130 mg/m. including disease prevention, slowing of disease pro 0280 (k) chronic low dose administration of from about gression and amelioration or reversal or disease symp 10 mg/day to about 25 mg/day; toms, based on the effect of meisoindigo on phosphory 0281 (1) intermittent administration of from about 50 lation of t (tau) protein. mg to about 150 mg twice weekly or three times weekly; 0323. The leukemia to be treated can be, but is not limited 0282 (m) administration of from about 50 mg/day to to, acute myelocytic leukemia (AML), chronic myelocytic about 150 mg/day for 10-14 days per month; and leukemia (CML), acute lymphocytic leukemia (ALL), 0283 (n) chronic daily dosing at a dose of equal to or chronic lymphocytic leukemia (CLL), or acute promyelo greater than 100 mg/day. cytic leukemia (APL). 0284. When the improvement is made by route of admin 0324. In one alternative, the indication for use is for treat istration, the route of administration can be, but is not limited ment of AML with or without administration of a therapeu to, at least one route of administration selected from the group tically effective quantity of an antifungal agent, wherein the consisting of: antifungal agent is metabolized by a different cytochrome 0285 (a) topical administration; P450 enzyme than the cytochrome P450 enzyme metaboliz 0286 (b) intravesicular administration for bladder can ing the indirubin derivative or analog. Antifungal agents cer, include, but are not limited to, amphotericin B. Voriconazole, 0287 (c) oral administration: itraconazole, fluconazole, caspofungin, flucytosine, keto 0288 (d) slow release oral delivery: conazole, posaconazole, clotrimazole, miconazole, econa 0289 (e) intrathecal administration; Zole, butoZonazole, oxiconazole, Sertaconazole, Sulconazole, 0290 (f) intraarterial administration; and terconazole. 0291 (g) continuous infusion; and 0325 In another alternative, the indication for use is for 0292 (h) intermittent infusion. treatment of APL with or without the administration of a 0293 When the improvement is made by schedule of therapeutically effective quantity of retinoic acids Such as administration, the schedule of administration can be, but is all-trans-retinoic acid (tretinoin) and arsenicals such as not limited to, at least one schedule of administration selected arsenic trioxide. from the group consisting of 0326 In still another alternative, the indication for use is 0294 (a) daily administration; for treatment of CML. In one aspect of this alternative, the 0295) (b) weekly administration: treatment of CML is of CML subsequent to or simultaneously 0296 (c) weekly administration for three weeks; with failure of treatment by tyrosine kinase inhibitors (TKIs). 0297 (d) biweekly administration: In another aspect of this alternative, the treatment of CML is 0298 (e) biweekly administration for three weeks with of CML that is transitioning or has transitioned to blast crisis. a 1-2 week rest period; In still another aspect of this alternative, the treatment of 0299 (f) intermittent boost dose administration; and CML is in combination with administration of a therapeuti 0300 (g) daily administration for one week for multiple cally effective quantity of a tyrosine kinase inhibitor (TKI). weeks. Tyrosine kinase inhibitors used for treatment of chronic 0301 When the improvement is made by indication for myelocytic leukemia (CML) include, but are not limited to, use, the indication for use can be, but is not limited to, at least imatinib, bosutinib, nilotinib, dasatinib, erlotinib, afatinib, one indication for use selected from the group consisting of and dacomitinib. Additional tyrosine kinase inhibitors are 0302) (a) use for treatment of leukemias; known in the art. For example, the use of tyrosine kinase 0303 (b) use for treatment of myelodysplastic syn inhibitors is described in United States Patent Application drome; Publication No. 2011/0206661 by Zhang et al., which is US 2016/0243077 A1 Aug. 25, 2016

directed to trimethoxyphenyl inhibitors of tyrosine kinase, elmustine, Erbamont FCE-24517, estramustine phosphate and in United States Patent Application Publication No. 2011/ sodium, fotemustine, Unimed G-6-M, Chinoin GYKI-17230, 0.195066, which is directed to quinoline inhibitors of tyrosine hepsul-fam, ifosfamide, iproplatin, lomustine, mafosfamide, kinase, both of which are incorporated herein by this refer melphalan, mitolactol, Nippon Kayaku NK-121, NCI NSC ence. The use of tyrosine kinase inhibitors is also described in 264395, NCINSC-342215, oxaliplatin, Upjohn PCNU, pred United States Patent Application Publication No. 2011/ nimustine, Proter PTT-119, ranimustine, semustine, Smith 053968 by Zhang et al., incorporated herein by this reference, Kline SK&F-101772, Yakult Honsha SN-22, spiromustine, which is directed to aminopyridine inhibitors of tyrosine Tanabe Seiyaku TA-077, tauromustine, temozolomide, terox kinase. The use of tyrosine kinase inhibitors is also described irone, tetraplatin, trimelamol, and uramustine, as described in in United States Patent Application Publication No. 2010/ U.S. Pat. No. 7,446,122 by Chao et al., incorporated herein by 0291025, incorporated herein by this reference, which is this reference. In still another aspect of this alternative, the directed to indazole inhibitors of tyrosine kinase. The use of treatment of CML is in combination with administration of a tyrosine kinase inhibitors is also described in United States therapeutically effective quantity of hydroxyurea. Patent Application Publication No. 2010/0190749 by Ren et 0328. When the improvement is made by selection of dis al., incorporated herein by this reference; these tyrosine ease stage, the selection of disease stage can be, but is not kinase inhibitors are benzoxazole compounds; compounds of limited to, at least one selection of disease stage selected from this class can also inhibit mTOR and lipid kinases such as the group consisting of phosphoinositide 3-kinases. The use of tyrosine kinase 0329 (a) use for the treatment of localized polyp stage inhibitors is also described in U.S. Pat. No. 8,242.270 by colon cancer, Lajeunesse et al., incorporated herein by this reference; these 0330 (b) use for leukoplakia in the oral cavity; tyrosine kinase inhibitors are 2-aminothiazole-5-aromatic 0331 (c) use for angiogenesis inhibition to prevent or carboxamides. Still other tyrosine kinase inhibitors are limit metastatic spread of a malignancy; and known in the art or are under development, and are described 0332 (d) use for treatment of HIV with a therapy in B. J. Druker & N. B. Lydon, “Lessons Learned from the Selected from the group consisting of azidothymidine Development of an Abl Tyrosine Kinase Inhibitor for Chronic (AZT), dideoxyadenosine (DDI), and reverse tran Myelogenous Leukemia. J. Clin. Invest. 105: 3-7 (2000), Scriptase inhibitors. incorporated herein by this reference. Instill another aspect of 0333 When the improvement is made by other indica this alternative, the treatment of CML is in combination with tions, the other indications can be, but are not limited, to at administration of a therapeutically effective quantity of least one other indication selected from the group consisting homoharringtonine. Homoharringtonine (omacetaxine of: mepesuccinate) has the structure shown below: 0334 (a) use as an anti-infective agent; 0335 (b) use as an antiviral agent;

0336 (c) use as an antibacterial agent; 0337 (d) use as an agent to treat pleural effusion; 0338 (e) use as an antifungal agent; 0339 (f) use as an anti-parasitic agent; 0340 (g) use as an agent to treat eczema: 0341 (h) use as an agent to treat herpes Zoster (shingles); 0342 (i) use as an agent to treat condylomata; 0343 () use as an agent to treat human papilloma virus (HPV); and 0344 (k) use as an agent to treat herpes simplex virus (HSV). 0345 When the improvement is made by patient selection, the patient selection can be, but is not limited to, a patient 0327 and is a protein translation inhibitor. Homohar selection carried out by a criterion selected from the group ringtonine inhibits protein translation by preventing the ini consisting of: tial elongation step of protein synthesis. It interacts with the 0346 (a) selecting patients with a disease condition ribosomal A-site and prevents the correct positioning of characterized by a high level of a metabolic enzyme amino acid side chains of incoming aminoacyl-tRNAS. In still Selected from the group consisting of histone deacety another aspect of this alternative, the treatment of CML is in lase and ornithine decarboxylase; combination with administration of a therapeutically effec 0347 (b) selecting patients with a low or high suscep tive quantity of an alkylating agent. Alkylating agents tibility to a condition selected from the group consisting include, but are not limited to, Shionogi 254-S, aldo-phos of thrombocytopenia and neutropenia; phamide analogues, altretamine, anaxirone, Boehringer 0348 (c) selecting patients intolerant of GI toxicities: Mannheim BBR-2207, bendamustine, bestrabucil, budoti and tane, buSulfan, Wakunaga CA-102, carboplatin, carmustine, 0349 (d) selecting patients characterized by over- or Chinoin-139, Chinoin-153, chlorambucil, cisplatin, cyclo under-expression of a gene selected from the group con phosphamide, American Cyanamid CL-286558, Sanofi sisting of c-Jun, a GPCR, a signal transduction protein, CY-233, cyplatate, Degussa D-19-384, Sumimoto DACHP VEGF, a prostate-specific gene, and a protein kinase. (Myr), dianhydrogalactitol, dibromodulcitol, other substi 0350. The cellular proto-oncogene c-Jun encodes a pro tuted hexitols, diphenylspiromustine, diplatinum cytostatic, tein that, in combination with c-Fos, forms the AP-1 early Erba distamycin derivatives, Chugai DWA-2114R, ITI E09, response transcription factor. This proto-oncogene plays a US 2016/0243077 A1 Aug. 25, 2016 34 key role in transcription and interacts with a large number of 0361 (e) measurement of the level of a metabolite or a proteins affecting transcription and gene expression. It is also metabolic enzyme. involved in proliferation and apoptosis of cells that form part 0362. The use of gene chips is described in A. J. Lee & S. of a number of tissues, including cells of the endometrium Ramaswamy, “DNA Microarrays in Biological Discovery and glandular epithelial cells. G-protein coupled receptors and Patient Care' in Essentials of Genomic and Personalized (GPCRs) are important signal transducing receptors. The Medicine (G. S. Ginsburg & H. F. Willard, eds., Academic Superfamily of G protein coupled receptors includes a large Press, Amsterdam, 2010), ch. 7, pp. 73-88, incorporated number of receptors. These receptors are integral membrane herein by this reference. proteins characterized by amino acid sequences that contain 0363. When the method is the use of single nucleotide seven hydrophobic domains, predicted to represent the trans polymorphism (SNP) analysis, the SNP analysis can be car membrane spanning regions of the proteins. They are found in ried out on a gene selected from the group consisting of a wide range of organisms and are involved in the transmis histone deacetylase, ornithine decarboxylase, VEGF, a pros sion of signals to the interior of cells as a result of their tate specific gene, c-Jun, and a protein kinase. The use of SNP interaction with heterotrimeric G proteins. They respond to a analysis is described in S. Levy and Y.-H. Rogers, “DNA diverse range of agents including lipid analogues, amino acid Sequencing for the Detection of Human Genome Variation” derivatives, Small molecules Such as epinephrine and dopam in Essentials of Genomic and Personalized Medicine (G. S. ine, and various sensory stimuli. The properties of many Ginsburg & H. F. Willard, eds., Academic Press, Amsterdam, known GPCR are summarized in S. Watson & S. Arkinstall, 2010), ch. 3, pp. 27-37, incorporated herein by this reference. “The G-Protein Linked Receptor Facts Book” (Academic Press, London, 1994), incorporated herein by this reference. 0364 Still other genomic techniques such as copy number GPCR receptors include, but are not limited to, acetylcholine variation analysis and analysis of DNA methylation can be receptors, B-adrenergic receptors, B-adrenergic receptors, employed. Copy number variation analysis is described in C. serotonin (5-hydroxytryptamine) receptors, dopamine recep Lee et al., “Copy Number Variation and Human Health' in tors, adenosine receptors, angiotensin Type II receptors, Essentials of Genomic and Personalized Medicine (G. S. bradykinin receptors, calcitonin receptors, calcitonin gene Ginsburg & H. F. Willard, eds., Academic Press, Amsterdam, related receptors, cannabinoid receptors, cholecystokinin 2010), ch.5, pp. 46-59, incorporated herein by this reference. receptors, chemokine receptors, cytokine receptors, gastrin DNA methylation analysis is described in S. Cottrell et al., receptors, endothelin receptors, y-aminobutyric acid (GABA) “DNA Methylation Analysis: Providing New Insight into receptors, galanin receptors, glucagon receptors, glutamate Human Disease' in Essentials of Genomic and Personalized receptors, luteinizing hormone receptors, choriogonadotro Medicine (G. S. Ginsburg & H. F. Willard, eds., Academic phin receptors, follicle-stimulating hormone receptors, thy Press, Amsterdam, 2010), ch. 6, pp. 60-72, incorporated roid-stimulating hormone receptors, gonadotrophin-releas herein by this reference. ing hormone receptors, leukotriene receptors, Neuropeptide 0365. When the improvement is made by prefpost-treat Y receptors, opioid receptors, parathyroid hormone recep ment preparation, the prefpost-treatment preparation can be, tors, platelet activating factor receptors, prostanoid (prostag but is not limited to, a method of prefpost treatment prepara landin) receptors, somatostatin receptors, thyrotropin-releas tion selected from the group consisting of: ing hormone receptors, vasopressin and oxytocin receptors. 0366 (a) the use of colchicine or an analog thereof; 0351 When the improvement is made by analysis of 0367 (b) the use of a uricosuric; patient or disease phenotype, the analysis of patient or disease 0368 (c) the use of uricase: phenotype can be, but is not limited to, a method of analysis 0369 (d) the non-oral use of nicotinamide: of patient or disease phenotype carried out by a method 0370 (e) the use of a sustained-release form of nicoti selected from the group consisting of: namide: 0352 (a) use of a diagnostic tool, a diagnostic tech nique, a diagnostic kit, or a diagnostic assay to confirm 0371 (f) the use of an inhibitor of poly-ADP ribose a patient’s particular phenotype; polymerase; 0353 (b) use of a method for measurement of a marker 0372 (g) the use of caffeine: Selected from the group consisting of histone deacety 0373 (h) the use of leucovorin rescue: lase, ornithine decarboxylase, VEGF, a protein that is a 0374 (i) infection control; and gene product of a prostate specific gene, a protein that is 0375 () the use of an anti-hypertensive agent. a gene product of jun, and a protein kinase; 0376 Uricosurics include, but are not limited to, 0354 (c) Surrogate compound dosing; and probenecid, benzbromarone, and Sulfinpyrazone. A particu 0355 (d) low dose pre-testing for enzymatic status. larly preferred uricosuric is probenecid. Uricosurics, includ 0356. When the improvement is made by analysis of ing probenecid, may also have diuretic activity. patient or disease genotype, the analysis of patient or disease 0377 Poly-ADP ribose polymerase inhibitors are genotype can be, but is not limited to, a method of analysis of described in G. J. Southan & C. Szabó, “Poly(ADP-Ribose) patient or disease genotype carried out by a method selected Inhibitors.” Curr. Med. Chem. 10:321-240 (2003), incorpo from the group consisting of rated herein by this reference, and include nicotinamide, 0357 (a) use of a diagnostic tool, a diagnostic tech 3-aminobenzamide, Substituted 3,4-dihydroisoquinolin-1 nique, a diagnostic kit, or a diagnostic assay to confirm (2H)-ones and isoquinolin-1 (2H)-ones, benzimidazoles, a patient’s particular genotype; indoles, phthalazin-1 (2H)-ones, quinazolinones, isoindoli 0358 (b) use of a gene chip; nones, phenanthridinones, and other compounds. 0359 (c) use of gene expression analysis: 0378 Leucovorin rescue comprises administration of foli 0360 (d) use of single nucleotide polymorphism (SNP) nic acid (leucovorin) to patients in which methotrexate has analysis; and been administered. Leucovorin is a reduced form of folic acid US 2016/0243077 A1 Aug. 25, 2016 that bypasses dihydrofolate reductase and restores hemato fen, ketoprofen, fenoprofin, oxaprozin, mefenamic acid, poietic function. Leucovorin can be administered either intra meclofenamic acid, piroXicam, meloxicam, nabumetone, venously or orally. rofecoxib, celecoxib, etodolac, nimeSulide, aceclofenac, 0379. In one alternative, wherein the prefpost treatment is alclofenac, alminoprofen, amfenac, ampiroXicam, apaZone, the use of a uricoSuric, the uricoSuric is probenecid or an araprofen, azapropaZone, bendazac, benoxaprofen, benzy analog thereof. damine, bermoprofen, benzpiperylon, bromfenac, bucloxic 0380 When the improvement is made by toxicity manage acid, bumadizone, butibufen, carprofen, cimicoxib, cinmeta ment, the toxicity management can be, but is not limited to, a cin, cinnoxicam, clidanac, clofeZone, clonixin, clopirac, dar method of toxicity management selected from the group con bufelone, deracoxib, droxicam, eltenac, enfenamic acid, epi sisting of: rizole, esflurbiprofen, ethenzamide, etofenamate, etoricoxib, 0381 (a) the use of colchicine or an analog thereof; felbinac, fenbufen, fenclofenac, fenclozic acid, fenclozine, 0382 (b) the use of a uricosuric; fendosal, fentiazac, feprazone, filenadol, flobufen, florife 0383 (c) the use of uricase; nine, flosulide, flubichin methanesulfonate, flufenamic acid, 0384 (d) the non-oral use of nicotinamide: flufenisal, flunixin, flunoxaprofen, fluprofen, fluproduaZone, 0385 (e) the use of a sustained-release form of nicoti furofenac, ibufenac, imrecoxib, indoprofen, isofeZolac, isox namide: epac, isoxicam, licofelone, lobuprofen, lomoxicam, lona (0386 (f) the use of an inhibitor of poly-ADP ribose Zolac, loxaprofen, lumaricoxib, mabuprofen, miroprofen, polymerase; mofebutaZone, mofeZolac, moraZone, nepafanac, niflumic (0387 (g) the use of caffeine; acid, nitrofenac, nitroflurbiprofen, nitronaproxen, orpanoxin, 0388 (h) the use of leucovorin rescue; oxaceprol, oxindanac, oXpinac, oxyphenbutaZone, pam 0389 (i) the use of sustained-release allopurinol: icogrel, parcetasal, parecoxib, parsalmide, pelubiprofen, 0390 () the non-oral use of allopurinol: pemedolac, phenylbutaZone, piraZolac, pirprofen, pranopro 0391 (k) the use of bone marrow transplants; fen, Salicin, Salicylamide, salicylsalicylic acid, satigrel, 0392 (1) the use of a blood cell stimulant; Sudoxicam, Suprofen, talmetacin, talniflumate, tazofelone, 0393 (m) the use of blood or platelet infusions: tebufelone, tenidap, tenoxicam, tepoxalin, tiaprofenic acid, 0394 (n) the administration of an agent selected from tiaramide, tilmacoxib, tinoridine,tiopinac, tioxaprofen, tolfe the group consisting offilgrastim (Neupogen.(R), G-CSF. namic acid, triflusal, tropesin, ursolic acid, Valdecoxib, Ximo and GM-CSF: profen, Zaltoprofen, Zidometacin, and Zomepirac, and the 0395 (o) the application of a pain management tech salts, Solvates, analogues, congeners, bioisosteres, hydrolysis nique; products, metabolites, precursors, and prodrugs thereof. 0396 (p) the administration of an anti-inflammatory 0407. The clinical use of corticosteroids is described in B. agent, P. Schimmer & K. L. Parker, Adrenocorticotropic Hormone: 0397 (d) the administration of fluids; Adrenocortical Steroids and Their Synthetic Analogs; Inhibi 0398 (r) the administration of a corticosteroid; tors of the Synthesis and Actions of Adrenocortical Hor 0399 (s) the administration of an insulin control medi mones' in Goodman & Gilman's The Pharmacological Basis cation; of Therapeutics (L. L. Brunton, ed., 11" ed., McGraw-Hill, 04.00 (t) the administration of an antipyretic: New York, 2006), ch.59, pp. 1587-1612, incorporated herein 0401 (u) the administration of an anti-nausea treat by this reference. ment; 0408 Anti-nausea treatments include, but are not limited 0402 (v) the administration of an anti-diarrheal treat to, ondansetron, metoclopramide, promethazine, cyclizine, ment; hyoscine, dronabinol, dimenhydrinate, diphenhydramine, 0403 (w) the administration of N-acetylcysteine; and hydroxy Zine, medizine, dolasetron, granisetron, pal 0404 (x) the administration of an antihistamine. onosetron, ramosetron, domperidone, haloperidol, chlorpro 04.05 Filgrastim is a granulocytic colony-stimulating fac mazine, fluiphenazine, perphenazine, prochlorperazine, tor (G-CSF) analog produced by recombinant DNA technol betamethasone, dexamethasone, lorazepam, and thiethylp ogy that is used to stimulate the proliferation and differentia erazine. tion of granulocytes and is used to treat neutropenia; G-CSF 04.09 Anti-diarrheal treatments include, but are not lim can be used in a similar manner. GM-CSF is granulocyte ited to, diphenoxylate, difenoxin, loperamide, codeine, macrophage colony-stimulating factor and stimulates stem racecadotril, octreoside, and berberine. cells to produce granulocytes (eosinophils, neutrophils, and 0410 N-acetylcysteine is an antioxidant and mucolytic basophils) and monocytes; its administration is useful to pre that also provides biologically accessible sulfur. vent or treat infection. 0411 When the improvement is made by pharmacoki 0406 Anti-inflammatory agents are well known in the art netic/pharmacodynamic monitoring, the pharmacokinetic/ and include corticosteroids and non-steroidal anti-inflamma pharmacodynamic monitoring can be, but is not limited to a tory agents (NSAIDs). Corticosteroids with anti-inflamma method selected from the group consisting of: tory activity include, but are not limited to, hydrocortisone, 0412 (a) multiple determinations of blood plasma lev cortisone, beclomethasone dipropionate, betamethasone, els; and dexamethasone, prednisone, methylprednisolone, triamcino 0413 (b) multiple determinations of at least one lone, fluocinolone acetonide, and fludrocortisone. Non-Ste metabolite in blood or urine. roidal anti-inflammatory agents include, but are not limited 0414 Typically, determination of blood plasma levels or to, acetylsalicylic acid (aspirin), sodium salicylate, choline determination of at least one metabolite in blood or urine is magnesium trisalicylate, Salsalate, diflunisal, Sulfasalazine, carried out by immunoassays. Methods for performing olsalazine, acetaminophen, indomethacin, Sulindac, tol immunoassays are well known in the art, and include radio metin, diclofenac, ketorolac, ibuprofen, naproxen, flurbipro immunoassay, ELISA (enzyme-linked immunosorbent US 2016/0243077 A1 Aug. 25, 2016 36 assay), competitive immunoassay, immunoassay employing 0463 Topoisomerase inhibitors include, but are not lim lateral flow test strips, and other assay methods. Immunoas ited to, irinotecan, topotecan, camptothecin, lamellarin D, says can be sandwich or competitive immunoassays. In amsacrine, etoposide, etoposide phosphate, teniposide, doxo another alternative, the determination of plasma levels of rubicin, and ICRF-193. meisoindigo can be determined by liquid chromatograpy 0464 Fraudulent nucleosides include, but are not limited mass spectrometry (LC-MS). The use of LC-MS is well to, cytosine arabinoside, gemcitabine, and fludarabine; other known in the art and is described in M. S. Lee & E. H. Kerns, fraudulent nucleosides are known in the art. “LC/MS Applications in Drug Development.” Mass Spec 0465 Fraudulent nucleotides include, but are not limited trometry Rev. 18:187-279 (1999), incorporated herein by this to, tenofovir disoproxil fumarate and adefovir dipivoxil; other reference. fraudulent nucleotides are known in the art. 0415. When the improvement is made by drug combina tion, the drug combination can be, but is not limited to, a drug 0466 Thymidylate synthetase inhibitors include, but are combination selected from the group consisting of not limited to, raltitrexed, pemetrexed, nolatrexed, ZD9331, 0416 (a) use with topoisomerase inhibitors; GS7094L, fluorouracil, and BGC 945. 0417 (b) use with fraudulent nucleosides: 0467 Signal transduction inhibitors are described in A. V. 0418 (c) use with fraudulent nucleotides; Lee et al., “New Mechanisms of Signal Transduction Inhibi 0419 (d) use with thymidylate synthetase inhibitors; tor Action: Receptor Tyrosine Kinase Down-Regulation and 0420 (e) use with signal transduction inhibitors; Blockade of Signal Transactivation.” Clin. Cancer Res. 9: 0421 (f) use with cisplatin or platinum analogs; 516s (2003), incorporated herein in its entirety by this refer 0422 (g) use with alkylating agents; CCC. 0423 (h) use with anti-tubulin agents: 0468 Cisplatin or platinum analogs include, but are not 0424 (i) use with antimetabolites; limited to, cisplatin, oxaliplatin, satraplatin, and carboplatin. 0425 (i) use with berberine; 0469 Alkylating agents include, but are not limited to, 0426 (k) use with apigenin: Shionogi 254-S, aldo-phosphamide analogues, altretamine, 0427 (1) use with amonafide: anaxirone, Boehringer Mannheim BBR-2207, bendamus 0428 (m) use with colchicine or an analog thereof; tine, bestrabucil, budotitane, Wakunaga CA-102, carboplatin, 0429 (n) use with genistein; carmustine, Chinoin-139, Chinoin-153, chlorambucil, cispl 0430 (o) use with etoposide; atin, cyclophosphamide, American Cyanamid CL-286558, 0431 (p) use with cytarabine; Sanofi CY-233, cyplatate, Degussa D-19-384. Sumimoto 0432 (q) use with a camptothecin: DACHP(Myr), diphenylspiromustine, diplatinum cyto 0433 (r) use with a vinca alkaloid; static, Erba distamycin derivatives, Chugai DWA-2114R, ITI 0434 (s) use with 5-fluorouracil; E09, elmustine, Erbamont FCE-24517, estramustine phos 0435 (t) use with curcumin; phate sodium, fotemustine, Unimed G-6-M, Chinoin GYKI 0436 (u) use with an NF-kB inhibitor; 17230, hepsul-fam, ifosfamide, iproplatin, lomustine, mafos 0437 (v) use with rosmarinic acid; famide, melphalan, mitolactol, Nippon Kayaku NK-121, NCI 0438 (w) use with mitoguaZone; NSC-264395, NCINSC-342215, oxaliplatin, Upjohn PCNU, 0439 (x) use with tetandrine; prednimustine, Proter PTT-119, ranimustine, semustine, 0440 (y) use with an antineoplastic agent not metabo SmithKline SK&F-101772, Yakult Honsha SN-22, spiromus lized by cytochrome P450 CYP1A2 or CYP2C19; tine, Tanabe Seiyaku TA-077, tauromustine, temozolomide, 0441 (Z) use with a biological therapy: teroxirone, tetraplatin and trimelamol, as described in U.S. 0442 (aa) use with a tyrosine kinase inhibitor; Pat. No. 7,446,122 by Chao et al., incorporated herein by this 0443 (ab) use with all-trans-retinoic acid; reference. 0444 (ac) use with an arsenical; 0445 (ad) use with hydroxyurea; 0470 Anti-tubulin agents include, but are not limited to, 0446 (ae) use with thioguanine: vinca alkaloids, taxanes, podophyllotoxin, halichondrin B, 0447 (af) use with mercaptopurine; and homohalichondrin B. 0448 (ag) use with homoharringtonine; 0471 Antimetabolites include, but are not limited to: methotrexate, pemetrexed, 5-fluorouracil, capecitabine, cyt 0449 (ah) use with oridonin; arabine, gemcitabine, 6-mercaptopurine, and pentostatin, 0450 (a) use with uracil mustard; alanosine, AG2037 (Pfizer), 5-FU-fibrinogen, acanthifolic 0451 (ak) use with nilotinib; acid, aminothiadiazole, brequinar Sodium, carmofur, Ciba 0452 (al) use with dasatinib; Geigy CGP-30694, cyclopentyl cytosine, cytarabine phos 0453 (am) use with lonidamine; phate stearate, cytarabine conjugates, Lilly DATHF, Merrill 0454 (an) use with 5-azacytidine; Dow DDFC, deazaguanine, dideoxycytidine, 0455 (ao) use with thalidomide or an analog thereof; dideoxyguanosine, didox, Yoshitomi DMDC, doxifluridine, 0456 (ap) use with an EGFR inhibitor such as erlotinib, Wellcome EHNA, Merck & Co. EX-015, fazarabine, floxu afatinib, lapatinib, or dacomitinib; ridine, fludarabine phosphate, N-(2-furanidyl)-5-fluorou 0457 (aq) use with a gold salt such as aurothiomalate or racil, Daiichi Seiyaku FO-152, isopropyl pyrrolizine, Lilly aurothioglucose: LY-188011, Lilly LY-264618, methobenzaprim, methotrex 0458 (ar) use with dibromodulcitol; ate, Wellcome MZPES, norspermidine, NCI NSC-127716, 0459 (as) use with dianhydrogalactitol; NCI NSC-264880, NCI NSC-39661, NCI NSC-612567, 0460 (at) use with decitabine; Warner-Lambert PALA, piritrexim, plicamycin, Asahi 0461) (au) use with a proteasome inhibitor; and Chemical PL-AC, Takeda TAC-788, thioguanine, tiazofurin, 0462 (av) use with a Nek9 inhibitor or an agent inhib Erbamont TIF, trimetrexate, tyrosine kinase inhibitors, iting the expression of the NEK9 gene. tyrosine protein kinase inhibitors, Taiho UFT and uricytin. US 2016/0243077 A1 Aug. 25, 2016 37

0472 Berberine has antibiotic activity and prevents and 0488 5-AZacytidine inhibits DNA methyltransferase and Suppresses the expression of pro-inflammatory cytokines and can be incorporated directly into RNA and DNA, causing cell E-selectin, as well as increasing adiponectin expression. death. 0473 Apigenin is a flavone that can reverse the adverse 0489. Thalidomide, and its analogs lenalidomide and effects of cyclosporine and has chemoprotective activity, pomalidomide, inhibit myeloma cells by a number of mecha either alone or derivatized with a Sugar. nisms, including inhibition of angiogenesis, inhibition of pro 0474 Amonafide is a topoisomerase inhibitor and DNA duction of interleukin-6, activation of apoptotic pathways intercalator that has anti-neoplastic activity. through caspase-8-mediated cell death, and other mecha 0475. Curcumin is believed to have anti-neoplastic, anti nisms. The antineoplastic activity of thalidomide is described inflammatory, antioxidant, anti-ischemic, anti-arthritic, and in W. D. Figg et al., “A Randomized Phase II Trial of Thali anti-amyloid properties and also has hepatoprotective activ domide, an Angiogenesis Inhibitor, in Patients with Andro ity. gen-Independent Prostate Cancer. Clin. Cancer Res. 7: 0476 NF-KB inhibitors include, but are not limited to, 1888-1893 (2001), incorporated herein by this reference. bortezomib. 0490 Epidermal growth factor receptor (EGFR) inhibi 0477 Rosmarinic acid is a naturally-occurring phenolic tors include, but are not limited to, erlotinib, afatinib, lapa antioxidant that also has anti-inflammatory activity. tinib, and dacomitinib. Other EGFR inhibitors are known in 0478 MitoguaZone is an inhibitor of polyamine biosyn the art. thesis through competitive inhibition of S-adenosylmethion 0491 Hydroxyurea is an antineoplastic agent that may act ine decarboxylase. by inhibiting the enzyme ribonucleotide reductase and also 0479. Tetrandrine has the chemical structure 6,6'.7.12-tet may induce double-stranded DNA breaks. ramethoxy-2,2'-dimethyl-1 B-berbaman and is a calcium 0492. Thioguanine is a purine analog of guanine and is channel blocker that has anti-inflammatory, immunologic, incorporated into DNA, as well as inhibition of the GTP and antiallergenic effects, as well as an anti-arrhythmic effect binding protein Rac1. similar to that of quinidine. It has been isolated from 0493 Mercaptopurine inhibits purine nucleotide synthe Stephania tetranda and other Asian herbs. sis and metabolism by inhibiting the enzyme phosphoribosyl 0480 Etoposide is an antineoplastic agent that forms a pyrophosphate amidotransferase. ternary complex with DNA and the enzyme topoisomerase II 0494 Gold salts, including, but not limited to, aurothi and thereby prevents religation of the DNA strands and omalate or aurothioglucose, act as inhibitors of lymphocyte causes DNA strand breakage. This causes errors in DNA proliferation. synthesis and promotes apoptosis of the cancer cells. 0495 Decitabine (5-aza-2'-deoxycytidine) is an agent that 0481 Cytarabine (cytosine arabinoside) is an antimetabo inhibits DNA methyltransferase and acts in a similar manner lite that interferes with DNA synthesis in cancer cells when to 5-azacytidine. converted to its triphosphate, cytosine arabinoside triphos 0496 Proteasome inhibitors include, but are not limited phate. to, bortezomib. Other proteasome inhibitors are known in the 0482 Camptothecins include, but are not limited to, art, including disulfiram, epigallocatechin-3-gallate, salino camptothecin, topotecan, and irinotecan. Other camptoth sporamide A, carfilzomib, oproZomib, delanzomib, ecins include DB 67, BNP 1350, exatecan, lurtotecan, ST MLN9708 (4-(carboxymethyl)-2-((R)-1-(2-(2,5-dichlo 1481, and CKD 602. The activity of camptothecins is robenzamido)acetamido)-3-methylbutyl)-6-oxo-1,3,2-diox described in H. Ulukan & P. W. Swaan, “Camptothecins, a aborinane-4-carboxylic acid), epoxomicin, and MG132 (N- Review of Their Chemotherapeutical Potential. Drugs 62: (benzyloxycarbonyl)leucinyleucinylleucinal-Z-Leu-Leu 2039-2057 (2002), incorporated herein by this reference. Leu-al). 0483 Vinca alkaloids include, but are not limited to, Vin 0497 As detailed below, a significant application of blastine, Vincristine, Vinorelbine, and vindesine. indirubin analogs and derivatives, particularly meisoindigo, 0484 5-Fluorouracil is a pyrimidine analog that acts by is for inhibiting Nek9 kinase. Therefore, indirubin analogs inhibiting the enzyme thymidylate synthase and thus prevents and derivatives, particularly meisoindigo, can be used in a the synthesis of thymidine. drug combination with Nek9 inhibitors or agents for inhibit 0485 Oridonin is a diterpenoid purified from Rabdosia ing the expression of the Nek9 gene. Other Nek9 inhibitors rubescens and inhibits proliferation of cells from lymphoid and agents for inhibiting the expression of the NEK9 gene are malignancies; its primary action appears to be blockage of the detailed below. NF-kB signal pathway; its activity is described in T. Ikezoe et 0498 United States Patent Application Publication No. al., “Oridonin, a Diterpenoid Purified from Rabdosia rubes 2010/0069458 by Atada et al., incorporated herein by this cens, Inhibits the Proliferation of Cells from Lymphoid reference discloses the use of the following additional thera Malignancies in Association with Blockade of the NF-kB peutic agents, which can be used together with indirubin Signal Pathways.” Mol. Cancer Ther: 4: 578-586 (2005), analogs and derivatives: incorporated herein by this reference. 0499 (1) ACE inhibitors, including, but not limited to, 0486 Nilotinib and dasatinib are tyrosine kinase inhibi benazepril, enazepril, captopril, enalapril, fosinopril, lisino tOrS. pril, moexipril, quinapril, ramipril, perindopril and trandola 0487 Lonidamine (1-(2,4-dichlorobenzyl)-1H-indazole pril; 3-carboxylic acid) is an aerobic glycolysis inhibitor in cancer 0500 (2) adenosine kinase inhibitors, including, but not cells; its activity is described in H. Pelicano et al., “Glycolysis limited to, 5-iodotubericidin; Inhibition for Anticancer Treatment. Oncogene 25: 4633 0501 (3) adrenal cortex antagonists, including, but not 4646 (2006), incorporated herein by this reference. limited to, mitotane; US 2016/0243077 A1 Aug. 25, 2016

0502 (4) AKT pathway inhibitors (protein kinase B 0508 (10) apoptosis inducers, including, but not limited inhibitors) including, but not limited to, deguelin and 1.5- to, 2-3-(2,3-dichlorophenoxy)propylamino-ethanol, dihydro-5-methyl-1-B-D-ribofuranosyl-1,4,5,6,8-pen gambogic acid, embellin, and arsenic trioxide; taaZaacenaphthylen-3-amine; 0509 (11) ATI receptor antagonists, including, but not 0503 (5) angiogenesis inhibitors, including, but not lim limited to, Valsartan; ited to, fumagillin, Shikonin, Tranilast, ursolic acid; Suramin; 0510 (12) aurora kinase inhibitors, including, but not lim thalidomide, lenalidomide; phthalazines, including, but not ited to, binucleine 2: limited to, 1-(4-chloroanilino)-4-(4-pyridylmethyl)phthala 0511 (13) aromatase inhibitors, including, but not limited Zine, 1-(4-methylanilino)-4-(4-pyridylmethyl)phthalazine, to: (a) steroids, including, but not limited to, atamestane, 1-(3-chloroanilino)-4-(4-pyridylmethyl)phthalazine, exemestane, and formestane; and (b) non-steroids, including, 1-anilino-4-(4-pyridylmethyl)phthalazine, 1-benzylamino but not limited to, aminoglutethimide, roglethimide, pyrido 4-(4-pyridylmethyl)phthalazine, 1-(4-methoxyanilino)-4-(4- glutethimide, triloStane, testolactone, ketokonazole, Voro pyridylmethyl)phthalazine, 1-(3-benzyloxyanilino)-4-(4-py Zole, fadrozole, anastroZole, and letrozole; ridylmethyl)phthalazine, 1-(3-methoxyanilino)-4-(4- 0512 (14) bisphosphonates, including, but not limited to, pyridylmethyl)phthalazine, 1-(2-methoxyanilino)-4-(4- etidronic acid, clodronic acid, tiludronic acid, alendronic pyridylmethyl)phthalazine, 1-(4-trifluoromethylanilino)-4- acid, ibandronic acid, risedronic acid, and Zoledronic acid; (4-pyridylmethyl)phthalazine, 1-(4-fluoroanilino)-4-(4- 0513 (15) Bruton's tyrosine kinase inhibitors, including, pyridylmethyl)phthalazine, 1-(3-hydroxyanilino)-4-(4- but not limited to, terreic acid; pyridylmethyl)phthalazine, 1-(4-hydroxyanilino)-4-(4- 0514 (16) calcineurin inhibitors, including, but not lim pyridylmethyl)phthalazine, 1-(3-aminoanilino)-4-(4- ited to, cypermethrin, deltamethrin, fenvalerate, and tyrphos pyridylmethyl)phthalazine, 1-(3,4-dichloroanilino)-4-(4- tin 8: pyridylmethyl)phthalazine, 1-(4-bromoanilino)-4-(4- 0515 (17) CaM kinase II inhibitors, including, but not pyridylmethyl)phthalazine, 1-(3-chloro-4-methoxyanilino)- limited to, the 5-isoquinolinesulfonic acid 4-(2S)-2-(5-iso 4-(4-pyridylmethyl)phthalazine, 1-(4-cyanoanilino)-4-(4- quinolinylsulfonyl)methylamino-3-oxo-3-(4-phenyl-1-pip pyridylmethyl)phthalazine, 1-(3-chloro-4-fluoroanilino)-4- erazinyl)propylphenyl ester, and N-2-3-(4-chlorophe (4-pyridylmethyl)phthalazine, 1-(3-methylanilino)-4-(4- nyl)-2-propenylmethylaminomethylphenyl-N-(2- pyridylmethyl)phthalazine, and other phthalazines disclosed hydroxyethyl)-4-methoxy-benzenesulfonamide: in PCT Patent Application Publication No. WO98/035958 by Bold et al., incorporated herein in its entirety by this refer 0516 (18) CD45 tyrosine phosphatase inhibitors, includ ence, isoquinolines disclosed in PCT Patent Application Pub ing, but not limited to, 2-(4-bromophenoxy)-5-nitrophenyl lication No. WO 00/09495 by Altmann et al., incorporated hydroxymethyl-phosphonic acid; herein in its entirety by this reference, including 1-(3,5-dim 0517 (19) CDC25 phosphatase inhibitors, including, but ethylanilino)-4-(pyridin-4-ylmethyl)-isoquinoline; phthala not limited to, 2,3-bis(2-hydroyethyl)thiol-1,4-naphthalene zines disclosed in PCT Patent Application Publication No. dione; WO 00/59509 by Bold et al., incorporated herein in its 0518 (20) CHK kinase inhibitors, including, but not lim entirety by this reference, including E-1-(3-methylanilino)- ited to, debromohymenialdisine; 4-(2-(pyridin-3-yl) vinylphthalazine, Z-1-(3-methyla 0519 (21) compounds targeting/decreasing a protein or nilino)-4-(2-(pyridin-3-yl) vinylphthalazine, 1-(3-methyla lipid kinase activity; or a protein or lipid phosphatase activity; nilino)-4-(2-(pyridin-3-yl)ethylphthalazine, 1-(3- or furtheranti-angiogenic compounds, including, but not lim methylanilino)-4-(2-(pyridin-4-yl) vinylphthalazine, 1-(4- ited to, protein tyrosine kinase and/or serine and/or threonine chloro-3-trifluoromethylanilino)-4-(2-(pyridin-3-yl)ethyl kinase inhibitors or lipid kinase inhibitors, including, but not phthalazine, 1-(4-chloroanilino)-4-(2-(pyridin-3-yl)ethyl limited to: phthalazine, 1-(3-chlorobenzylamino)-4-(2-(pyridin-3-yl) 0520 (a) compounds targeting, decreasing or inhibiting ethylphthalazine, 1-(4-chloro-3-trifluoromethylanilino)-4- the activity of the vascular endothelial growth factor 3-(pyridin-3-yl)propylphthalazine, 1-(4-chloroanilino)-4- receptors (VEGFR) or of vascular endothelial growth 3-(pyridin-3-yl)propylphthalazine, 1-(3-chloro-5- factor (VEGF), including, but not limited to, 7H-pyrrolo trifluoromethylanilino)-4-3-(pyridin-3-yl)propyl 2,3-dipyrimidine derivatives, including: 6-4-(4- phthalazine, and 1-(4-tert-butylanilino)-4-3-(pyridin-3-yl) ethyl-piperazine-1-ylmethyl)-phenyl-7H-pyrrolo2,3- propylphthalazine; and monoclonal antibodies; dpyrimidinpyrimidin-4-yl)-(R)-1-phenyl-ethyl)-amine 0504 (6) angiostatic steroids, including, but not limited to, (known as AEE788), BAY 43-9006; and isoquinoline anecortave, triamcinolone, hydrocortisone, 11 C-epihydroco compounds disclosed in PCT Patent Application Publi tisol, cortexolone, 17O-hydroxyprogesterone, corticosterone, cation No. WO 00/09495, such as (4-tert-butyl-phenyl)- desoxycorticosterone, testosterone, estrone, and dexametha 94-pyridin-4-ylmethyl-isoquinolin-1-yl)-amine; 0521 (b) compounds targeting, decreasing or inhibiting Sone; the activity of the platelet-derived growth factor-recep 0505 (7) anti-androgens, including, but not limited to, tor (PDGFR), including, but not limited to: N-phenyl-2- nilutamide and bicalutamide; pyrimidine-amine derivatives, e.g., imatinib, SU101, 0506 (8) anti-estrogens, including, but not limited to, SU6668 and GFB-111; toremifene, letrozole, testolactone, anastroZole, bicaluta 0522 (c) compounds targeting, decreasing or inhibiting mide, flutamide, exemestane, tamoxifen, fulvestrant, and ral the activity of the fibroblast growth factor-receptor oxifene; (FGFR); 0507 (9) anti-hypercalcemia agents, including, but not 0523 (d) compounds targeting, decreasing or inhibiting limited to, gallium (III) nitrate hydrate and pamidronate diso the activity of the insulin-like growth factor receptor 1 dium; (IGF-1R), including, but not limited to: the compounds US 2016/0243077 A1 Aug. 25, 2016 39

disclosed in WO 02/092599 and derivatives thereof of able salt thereof, especially a compound selected from 4-amino-5-phenyl-7-cyclobutyl-pyrrolo2,3-dipyrimi the benzylidenemalonitrile class or the 5-arylbenzen dine derivatives; emalonirile or bisubstrate quinoline class of com 0524 (e) compounds targeting, decreasing or inhibiting pounds, more especially any compound selected from the activity of the Trk receptor tyrosine kinase family; the group consisting of Tyrphostin A23/RG-50810, Tyr 0525 (f) compounds targeting, decreasing or inhibiting phostin AG 99, Tyrphostin AG 213, Tyrphostin AG the activity of the Axl receptor tyrosine kinase family; 1748, Tyrphostin AG 490, Tyrphostin B44, Tyrphostin 0526 (g) compounds targeting, decreasing or inhibiting B44 (+) enantiomer, Tyrphostin AG 555, AG 494, Tyr the activity of the c-Met receptor; phostin AG 556;Tyrphostin AG957, and adaphostin (4- 0527 (h) compounds targeting, decreasing or inhibiting (2,5-dihydroxyphenyl)methylamino-benzoic acid the activity of the Ret receptor tyrosine kinase; adamantyl ester or NSC 680410): 0528 (i) compounds targeting, decreasing or inhibiting 0533 (n) compounds targeting, decreasing or inhibiting the activity of the Kit/SCFR receptor tyrosine kinase; the activity of the epidermal growth factor family of 0529 () compounds targeting, decreasing or inhibiting receptor tyrosine kinases (EGFR, ErbB2, ErbB3, ErbB4 the activity of the C-kit receptor tyrosine kinases, as homodimers or heterodimers). Such as, but not limited including, but not limited to, imatinib; to, those compounds, proteins or monoclonal antibodies 0530 (k) compounds targeting, decreasing or inhibiting generically and specifically disclosed in PCT Patent the activity of members of the c-Abl family and their Application Publication No. WO97/02266 by Traxleret gene-fusion products, e.g., BCR-Abl kinase, such as al. Such as (R)-6-(4-hydroxyphenyl)-4-(1-phenyl N-phenyl-2-pyrimidine-amine derivatives, including, ethyl)-amino-7H-pyrrolo-2,3-dipyrimidine, or in but not limited to: imatinib, 6-(2,6-dichlorophenyl)-2- European Patent Application Publication No. EP (4-fluoro-3-methylphenyl)amino-8-methyl-pyrido2, 0564409 by Zimmermann, PCT Patent Application Pub 3-dipyrimidin-7(8H)-one (PD180970), methyl-4-N- lication No. WO99/03854 by Zimmermannet al., Euro (2',5'-dihydroxybenzyl)aminobenzoate (Tyrphostin pean Patent Application Publication No. EP 0520722 by AG957), 4-(2,5-dihydroxyphenyl)methylaminoben Barker et al., European Patent Application Publication Zoic acid tricyclo[3.3.1.13.7 dec-1-yl ester (adaphostin No. EP 0566226 by Barker et al., European Patent or NSC 680410), 6-(2,6-dichlorophenyl)-8-methyl-2- Application Publication EP 0787722 by Wissner et al., (3-methylsulfanylanilino)pyrido2,3-dipyrimidin-7- European Patent Application Publication EP 0837063 one (PD173955), and desatinib; by Arnoldet al., U.S. Pat. No. 5,747,498 by Schnuretal. 0531 (1) compounds targeting, decreasing or inhibiting PCT Patent Application Publication WO 98/10767 by the activity of members of the protein kinase C (PKC) McMahon et al., PCT Patent Application Publication and Raffamily of serine/threonine kinases, members of WO97/30034 by Barker, PCT Patent Application Pub the MEK, SRC, JAK, FAK, PDKand Ras/MAPK family lication WO 97/49688 by Schnur, PCT Patent Applica members, or P1(3) kinase family, or of the PI(3)-kinase tion Publication WO 97/38983 by Bridges et al., PCT related kinase family, and/or members of the cyclin Patent Application Publication WO96/30347 by Schnur dependent kinase family (CDK) and are especially those et al., including, but not limited to, N-(3-ethylnylphe staurosporine derivatives disclosed in U.S. Pat. No. nyl)-6,7-bis(2-methoxyethoxy)-4-quinazolinamine (CP 5,093,330, such as, but not limited to, midostaurin; 358774 or erlotinib), PCT Patent Application Publica examples of further compounds include, e.g., UCN-01; tion WO 96/33980 by Gibson et al., including, but not safingol, Sorafenib, Bryostatin 1: Perifosine; Ilmofos limited to, N-(3-chloro-4-fluoro-phenyl)-7-methoxy-6- ine: 3-3-2,5-Dihydro-4-(1-methyl-1H-indol-3-yl)-2, (3-morpholin-4-ylpropoxy)guinazolin-4-amine (gefi 5-dioxo-1H-pyrrol-3-yl)-1H-indol-1-ylpropyl carbam tinib); and PCT Patent Application Publication WO imidothioic acid ester (RO 318220), 3-(8S)-8- 95/03283 by Barker et al., including, but not limited to, (dimethylamino)methyl-6,7,8,9-tetrahydropyrido1, compound 6-amino-4-(3-methylphenyl-amino)- 2-alindol-10-yl)-4-(1-methyl-1H-indol-3-yl)-1H quinazoline (ZM105180); monoclonal antibodies, pyrrole-2,5-dione (RO 320432), 12-(2-cyanoethyl)-6.7, including, but not limited to trastuzumab and cetuximab; 12,13-tetrahydro-13-methyl-5-oxo-5H-indolo2,3-a and other Small molecule inhibitors, including, but not pyrrolo[3,4-ccarbazole (GO 6976); Isis 3521; (S)-13 limited to: canertinib, pelitinib, lapatinib, and 7H-pyr (dimethylamino)methyl-10,11,14, 15-tetrahydro-4.9: rolo-2,3-dipyrimidine derivatives which are disclosed 16, 21-dimetheno H, 13H-dibenzoekpyrrolo 3,4-h in PCT Patent Application Publication WO 03/013541 1.4.13oxadiazacy clohexadecene-1,3(2H)-dione by Bold et al.: (LY333531), LY379196; isoquinoline compounds, such 0534 (22) compounds which target, decrease or inhibit as those disclosed in PCT Patent Application Publica the activity of a protein or lipid phosphatase, including, but tion No. WO 00/09495; farnesyltransferase inhibitors, not limited to, inhibitors of phosphatase 1, phosphatase 2A, including, but not limited to, tipifarnib and lonafarnib; PTEN or CDC25, such as, but not limited to okadaic acid or 2-(2-chloro-4-iodo-phenylamino)-N-cyclopropyl a derivative thereof; methoxy-3,4-difluoro-benzamide (PD184352); and 0535 (23) compounds which induce cell differentiation QAN697, a PI3K inhibitor; processes, including, but not limited to, retinoic acid, C-to 0532 (m) compounds targeting, decreasing or inhibit copherol, Y-tocopherol, Ö-tocopherol, C-tocotrienol, Y-tocot ing the activity of protein-tyrosine kinase, such as, but rienol, and 8-tocotrienol; not limited to, imatinib mesylate, a tyrphostin, 0536 (24) cRAF kinase inhibitors, including, but not lim pyrymidylaminobenzamide and derivatives thereof a ited to, 3-(3,5-dibromo-4-hydroxybenzylidene)-5-iodo-1,3- tyrphostin is preferably a low molecular weight dihydroindol-2-one and 3-(dimethylamino)-N-3-(4-hy (M.<1500) compound, or a pharmaceutically accept droxybenzoyl)amino-4-methylphenyl-benzamide; US 2016/0243077 A1 Aug. 25, 2016 40

0537 (25) cyclindependent kinase inhibitors, including, cines; and epothilones and derivatives thereof. Such as but not limited to, N9-isopropyl-olomoucine; olomoucine; epothilone B or a derivative thereof; purvalanol B, roascovitine, kenpaullone, and purvalanol A. 0555 (43) mitogen-activated protein (MAP) kinase 0538 (26) cysteine protease inhibitors, including, but not inhibitors, including, but not limited to, N-2-3-(4-chlo limited to, N-(1S)-3-fluoro-2-oxo-1-(2-phenylethyl)pro rophenyl)-2-propenylmethylaminomethylphenyl-N-(2- pylamino-2-oxo-1-(phenylmethyl)ethyl-4-morpholin hydroxyethyl)-4-methoxy-benzenesulfonamide: ecarboxamide; 0556 (44) MDM2 inhibitors, including, but not limited to, 0539 (27) DNA intercalators, including, but not limited trans-4-iodo,4'-boranyl-chalcone; to, plicamycin and dactinomycin; 0557 (45) MEK inhibitors, including, but not limited to, 0540 (28) DNA strand breakers, including, but not limited bisamino 2-aminophenyl)thiomethylene-butanedinitrile; to, bleomycin; 0558 (46) methionine aminopeptidase inhibitors, includ 0541 (29) E3 ligase inhibitors, including, but not limited ing, but not limited to, bengamide and derivatives thereof; to, N-((3,3,3-trifluoro-2-trifluoromethyl)propionyl)sulfanil 0559 (47) MMP inhibitors, including, but not limited to: actinonin; epigallocatechin gallate; collagen peptidomimetic amide; and non-peptidomimetic inhibitors; tetracycline derivatives 0542 (30) EDG binders, including, but not limited to, Such as hydroxamate, batimastat, marimastat, primomastat, FTY720; TAA211, N-hydroxy-2(R)-(4-methoxyphenyl)sulfonyl(3- 0543 (31) endocrine hormones, including, but not limited to, leuprolide and megestrol acetate; picolyl)amino-3-methylbutanamide hydrochloride (MMI27OB), and AAJ996; 0544 (32) farnesyltransferase inhibitors, including, but 0560 (48) NGFR tyrosine kinase inhibitors, including, not limited to, C.-hydroxyfarnesylphosphonic acid, 2-(2S)- but not limited to, Tyrphostin AG 879; 2-(2S,3S)-2-(2R)-2-amino-3-mercaptopropylamino-3- 0561 (49) p38 MAP kinase inhibitors, including, but not methylpentylloxy-1-oxo-3-phenylpropylamino-4-(meth limited to, 3-(dimethylamino)-N-3-(4-hydroxybenzoyl) ylsulfonyl)-, 1-methylethyl butanoic acid ester (2S), and amino-4-methylphenyl-benzamide: manumycin A; 0562 (50) p56 tyrosine kinase inhibitors, including, but 0545 (33) Flk-1 kinase inhibitors, including, but not lim not limited to, 9,10-dihydro-3-hydroxy-1-methoxy-9,10-di ited to, 2-cyano-3-4-hydroxy-3,5-bis(1-methylethyl)phe oXo-2-anthracenecarboxaldehyde and Tyrphostin 46: nyl-N-(3-phenylpropyl)-, (2-E)-2-propenamide: 0563 (51) PDGFR tyrosine kinase inhibitors, including, 0546 (34) Flt-3 inhibitors, including, but not limited to, but not limited to, Tyrphostin AG 1296: Tyrphostin 9. N-benzoyl-staurosporine, midostaurin, and N-(2-diethylami 2-amino-4-(1H-indol-5-yl)-1,3-butadiene-1,1,3-tricarboni noethyl)-5-(Z)-(5-fluoro-2-oxo-1H-indol-3-ylidene)me trile, and imatinib; thyl-2,4-dimethyl-1H-pyrrole-3-carboxamide (sunitinib); 0564 (52) phosphatidylinositol 3-kinase inhibitors, 0547 (35) gonadorelinagonists, including, but not limited including, but not limited to, wortmannin and quercetin dihy to, abarelix, goserelin, and goserelin acetate; drate; 0548 (36) heparanase inhibitors, including, but not lim 0565 (53) phosphatase inhibitors, including, but not lim ited to, phosphomannopentaose sulfate (PI-88); ited to, cantharidic acid, cantharidin, and (E)-N-4-(2-car (0549 (37) histone deacetylase (HDAC) inhibitors, includ boxyethenyl)benzoylglycyl-L-O-glutamyl-L-leucinamide; ing, but not limited to, compounds disclosed in PCT Patent 0566 (54) platinum agents, including, but not limited to, Application Publication No. WO 02/22577 by Bair et al., carboplatin, cisplatin, oxaliplatin, satraplatin, and ZD0473; including, but not limited to, N-hydroxy-3-4-(2-hydroxy 0567 (55) protein phosphatase inhibitors, including, but ethyl)2-(1H-indol-3-yl)ethyl-aminomethylphenyl-2E-2- not limited to: propenamide, Suberoylanilide hydroxamic acid, 4-(2-amino 0568 (a) PP1 and PP2A inhibitors, including, but not phenylcarbamoyl)-benzyl-carbamic acid pyridine-3- limited to, cantharidic acid and cantharidin; and ylmethyl ester and derivatives thereof, butyric acid, 0569 (b) tyrosine phosphatase inhibitors, including, pyroxamide, trichostatin A, oxamflatin, apicidin, depsipep but not limited to, L-P-bromotetramisole oxalate, ben tide, depudecin, trapoxin, HC toxin, and Sodium phenylbu Zylphosphonic acid, and (5R)-4-hydroxy-5-(hydroxym tyrate; ethyl)-3-(1-oxohexadecyl)-2(5H)-furanone; 0550 (38) HSP90 inhibitors, including, but not limited to: (0570 (56) PKC inhibitors, including, but not limited to, 17-allylamino,17-demethoxygeldanamycin (17AAG); a —1-3-(dimethylamino)propyl)-1H-indol-3-yl)-4-(1H-in geldanamycin derivative; other geldanamycin-related com dol-3-yl)-1H-pyrrolo-2,5-dione, sphingosine, staurosporine, pounds; radicicol; and 5-(2,4-dihydroxy-5-isopropyl-phe Tyrphostin 51, and hypericin; nyl)-4-(4-morpholin-4-ylmethyl-phenyl)-isoxazole-3-car (0571 (57) PKC delta kinase inhibitors, including, but not boxylic acid ethylamide: limited to, rottlerin; 0551 (39) IKBO. inhibitors (IKKs), including, but not lim 0572 (58) polyamine synthesis inhibitors, including, but ited to, 3-(4-methylphenyl)sulfonyl-(2E)-2-propenenitrile; not limited to, (RS)-2,5-diamino-2-(difluoromethyl)pen 0552 (40) insulin receptor tyrosine kinase inhibitors, tanoic acid (DMFO): including, but not limited to, hydroxy-2-naphthalenylmeth 0573 (59) proteasome inhibitors, including, but not lim ylphosphonic acid; ited to, aclacinomycin A, gliotoxin, and bortezomib; 0553 (41) c-Jun N-terminal kinase inhibitors, including, (0574 (60) PTP1B inhibitors, including, but not limited to, but not limited to, pyrazoleanthrone and epigallocatechin (E)-N-4-(2-carboxyethenyl)benzoylglycyl-L-O-glutamyl gallate; L-leucinamide; 0554 (42) microtubule binding agents, including, but not 0575 (61) protein tyrosine kinase inhibitors, including, limited to: vinblastine sulfate; Vincristine sulfate; Vindesine; but not limited to: Tyrphostin AG 126: Tyrphostin AG 1288: vinorelbine; docetaxel; paclitaxel; discodermolides; colchi Tyrphostin AG 1295; geldanamycin; and genistein; US 2016/0243077 A1 Aug. 25, 2016

0576 (62) SRC family tyrosine kinase inhibitors, includ 0593. In one alternative, when the drug combination is use ing, but not limited to, 1-(1,1-dimethylethyl)-3-(1-naphthale with an alkylating agent, the alkylating agent can be selected nyl)-1H-pyrazolo 3,4-dipyrimidin-4-amine, and 3-(4-chlo from the group consisting of BCNU, BCNU wafers (Gliadel), rophenyl)-1-(1,1-dimethylethyl)-1H-pyrazolo 3,4-d and CCNU. pyrimidin-4-amine; 0594. When the improvement is made by chemosensitiza 0577 (63) Syk tyrosine kinase inhibitors including, but tion, the chemosensitization can comprise, but is not limited not limited to, piceatannol; to, the use of indirubin, an analog of indirubin, or a derivative 0578 (64) Janus (JAK-2 and/or JAK-3) tyrosine kinase of indirubin or of an analog of indirubin as a chemosensitizer inhibitors, including, but not limited to, Tyrphostin AG 490, in combination with an agent selected from the group con and 2-maphthyl vinyl ketone; sisting of: 0579 (65) inhibitors of Ras oncogenic isoforms, includ 0595 (a) topoisomerase inhibitors; ing, but not limited to, (2S)-2-(2S)-2-(2S,3S)-2-(2R)-2- 0596 (b) fraudulent nucleosides; amino-3-mercaptopropylamino-3-methylpentylloxy)-1- 0597 (c) fraudulent nucleotides: oXo-3-phenylpropylamino-4-(methylsulfonyl)-butanoic 0598 (d) thymidylate synthetase inhibitors; acid 1-methylethyl ester (L-744832), DK8G557, and tipi 0599 (e) signal transduction inhibitors; farnib; 0600 (f) cisplatin or platinum analogs; 0580 (66) retinoids, including, but not limited to, isotret 0601 (g) alkylating agents; inoin and tretinoin, 0602 (h) anti-tubulin agents; 0581 (67) ribonucleotide reductase inhibitors, including, 0603 (i) antimetabolites: but not limited to, hydroxyurea and 2-hydroxy-1H-isoindole 0604) () berberine; 1,3-dione; 0605 (k) apigenin: 0582 (68) RNA polymerase II elongation inhibitors, 0606 (1) amonafide: including, but not limited to, 5,6-dichloro-1-beta-D-ribofura 0607 (m) vinca alkaloids; nosylbenzimidazole; 0608 (n) 5-fluorouracil; 0583 (69) S-adenosylmethionine decarboxylase inhibi 0609 (o) curcumin; tors, including, but not limited to, 5-amidino-1-tetralone-2'- 0610 (p) NF-kB inhibitors; amidinohydrazone and other compounds disclosed in U.S. 0611 (q) rosmarinic acid; Pat. No. 5,461,076 to Stanek et al., incorporated herein by this 0612 (r) mitoguaZone: reference; 0613 (s) tetrandrine; and 0584 (70) serine/threonine kinase inhibitors, including, 0.614 (t) proteasome inhibitors. but not limited to, Sorafenib and 2-aminopurine; 0615. When the improvement is made by chemopotentia 0585 (71) compounds which target, decrease, or inhibit tion, the chemopotentiation can comprise, but is not limited the activity or function of serine/threonine mTOR kinase, to, the use of indirubin, an analog of indirubin, or a derivative including, but not limited to, everolimus, temsirolimus, of indirubin or of an analog of indirubin as a chemopotentia Zotarolimus, rapamycin, derivatives and analogs of rapamy tor in combination with an agent selected from the group cin, deforolimus, AP23841, sirolimus, and everolimus; consisting of: 0586 (72) Somatostatin receptor antagonists, including, 0616 (a) topoisomerase inhibitors; but not limited to, octreotide and pasireotide (SOM230); 0.617 (b) fraudulent nucleosides; 0587 (73) sterol biosynthesis inhibitors, including, but 0618 (c) fraudulent nucleotides: not limited to, terbinadine; 0619 (d) thymidylate synthetase inhibitors; 0588 (74) telomerase inhibitors, including, but not lim 0620 (e) signal transduction inhibitors; ited to, telomestatin; and 0621 (f) cisplatin or platinum analogs; 0589 (75) topoisomerase inhibitors, including, but not limited to: 0622 (g) alkylating agents; 0590 (a) topoisomerase I inhibitors, including, but not 0623 (h) anti-tubulin agents; limited to, topotecan, gimatecan, irinotecan, camptoth 0624 (i) antimetabolites: ecin and its analogues, 9-nitrocamptothecin and the 0625 (i) berberine; macromolecular camptothecin conjugate PNU-16614, 0626 (k) apigenin: macromolecular camptothecin conjugates described in 0627 (1) amonafide: PCT Patent Application Publication No. WO99/17804 0628 (m) vinca alkaloids; by Angelucci et al., 10-hydroxycamptothecin acetate 0629 (n) 5-fluorouracil; salt, etoposide idarubicin hydrochloride, teniposide, 0630 (o) curcumin; doxorubicin; epirubicin hydrochloride, mitoxantrone 0631 (p) NF-kB inhibitors; hydrochloride, and daunorubicin hydrochloride; and 0632 (q) rosmarinic acid; 0591 (b) topoisomerase II inhibitors, including, but not 0633 (r) mitoguaZone: limited to, anthracyclines, such as doxorubicin, includ 0634 (s) tetrandrine; and ing liposomal formulations thereof, daunorubicin, 0635 (t) proteasome inhibitors. including liposomal formulations thereof, epirubicin, 0636. In one alternative, when the chemopotentiation idarubicin, nemorubicin, mitoxantrone, losoxantrone, involves chemopotentiation of an alkylating agent by the etoposide, and eniposide; activity of indirubin, an analog of indirubin, or a derivative of 0592 (76) VEGFR tyrosine kinase inhibitors, including, indirubin orofananalog of indirubin. the alkylating agent can but not limited to, 3-(4-dimethylaminobenzylidenyl)-2-in be selected from the group consisting of BCNU, BCNU dolinone; and (77) RANKL inhibitors, including, but not wafers (Gliadel), CCNU, bendamustine (Treanda), and temo limited to, denosumab. Zolimide (Temodar). US 2016/024.3077 A1 Aug. 25, 2016 42

0637. When the improvement is made by post-treatment 0664. When the improvement is made by use of a diluent, management, the post-treatment management can be, but is the diluent can be, but is not limited to, a diluent selected from not limited to, a method selected from the group consisting of: the group consisting of: 0638 (a) a therapy associated with pain management; 0665 (a) an emulsion; 0666 (b) dimethylsulfoxide (DMSO); 0639 (b) administration of an anti-emetic; 0667 (c) N-methylformamide (NMF) 0640 (c) an anti-nausea therapy: 0668) (d) DMF; 0641 (d) administration of an anti-inflammatory agent; 0669 (e) ethanol: 0642 (e) administration of an anti-pyretic agent; and 0670 (f) benzyl alcohol: 0643 (f) administration of an immune stimulant. 0671 (g) dextrose-containing water for injection; 0644) When the improvement is made by alternative medi 0672 (h) Cremophor; cine/post-treatment support, the alternative medicine/post 0673 (i) cyclodextrin; and 0674) (j) PEG. treatment support can be, but is not limited to, a method 0675 When the improvement is made by use of a solvent selected from the group consisting of: system, the solvent system can be, but is not limited to, a 0645 (a) hypnosis: solvent system selected from the group consisting of 0646) (b) acupuncture; 0676 (a) an emulsion; (0647 (c) meditation; 0677) (b) dimethylsulfoxide (DMSO); (0648 (d) a herbal medication created either syntheti 0678 (c) N-methylformamide (NMF) cally or through extraction; and 0679) (d) DMF; 0680 (e) ethanol: 0649 (e) applied kinesiology. 0681 (f) benzyl alcohol: 0650. In one alternative, when the method is a herbal 0682 (g) dextrose-containing water for injection; medication created either synthetically or through extraction, 0683 (h) Cremophor; the herbal medication created either synthetically or through 0684 (i) cyclodextrin; and extraction can be selected from the group consisting of: 0685 (i) PEG. 0651 (a) a NF-kB inhibitor; 0686. When the improvement is made by use of an excipi ent, the excipient can be, but is not limited to, an excipient 0652 (b) a natural anti-inflammatory; selected from the group consisting of: 0653 (c) an immunostimulant; group consisting of: 0654) (d) an antimicrobial; and 0687 (a) mannitol; 0655 (v) a flavonoid, isoflavone, or flavone. 0688 (b) albumin; 0656. When the herbal medication created either syntheti 0689) (c) EDTA; cally or through extraction is a NF-kB inhibitor, the NF-KB (0690 (d) sodium bisulfite: inhibitor can be selected from the group consisting of par 0691 (e) benzyl alcohol; thenolide, curcumin, and rosmarinic acid. When the herbal 0692 (f) a carbonate buffer; medication created either synthetically or through extraction (0693) (g) a phosphate buffer; and is a natural anti-inflammatory, the natural anti-inflammatory (0694 (h) methylcellulose. can be selected from the group consisting of rhein and par 0695. When the improvement is made by use of a dosage thenolide. When the herbal medication created either syn form, the dosage form can be, but is not limited to, a dosage thetically or through extraction is an immunostimulant, the form selected from the group consisting of: immunostimulant can be a product found in or isolated from 0696 (a) tablets; Echinacea. When the herbal medication created either syn (0697) (b) capsules; thetically or through extraction is an anti-microbial, the anti 0698 (c) topical gels; microbial can be berberine. When the herbal medication cre (0699 (d) topical creams; ated either synthetically or through extraction is a flavonoid (0700 (e) patches; or flavone, the flavonoid, isoflavone, or flavone can be 0701 (f) suppositories; and selected from the group consisting of apigenin, genistein, (0702 (g) lyophilized dosage fills. apigenenin, genistein, genistin, 6"-O-malonylgenistin, 6"-O- 0703 Formulation of pharmaceutical compositions in tab acetylgenistin, daidzein, daidzin, 6"-O-malonyldaidzin, lets, capsules, and topical gels, topical creams or supposito 6"-O-acetylgenistin, glycitein, glycitin, 6"-O-malonylgly ries is well known in the art and is described, for example, in citin, and 6-O-acetylglycitin. United States Patent Application Publication No. 2004/ 0657. When the improvement is made by a bulk drug 0023290 by Griffin et al., incorporated herein by this refer product improvement, the bulk drug product improvement CCC. can be, but is not limited to, a bulk drug product improvement 0704. Formulation of pharmaceutical compositions as selected from the group consisting of patches such as transdermal patches is well known in the art 0658 (a) salt formation; and is described, for example, in U.S. Pat. No. 7,728,042 to Eros et al., incorporated herein by this reference. 0659 (b) preparation as a homogeneous crystal struc (0705 Lyophilized dosage fills are also well known in the ture; art. One general method for the preparation of such lyo 0660 (c) preparation as a pure isomer; philized dosage fills, applicable to indirubin, an analog of 0661 (d) increased purity; indirubin, or a derivative of indirubin or of an analog of 0662 (e) preparation with lower residual solvent con indirubin, comprises the following steps: tent; and (0706 (1) Dissolve the drug in water for injection pre 0663 (f) preparation with lower residual heavy metal cooled to below 10°C. Dilute to final volume with cold water content. for injection to yield a 40 mg/mL solution. US 2016/0243077 A1 Aug. 25, 2016

(0707 (2) Filter the bulk solution through an 0.2-lum filter anhydride); polydioxanone; polyamines; polyamides; poly into a receiving container under aseptic conditions. The for urethanes; polyesteramides; polyorthoesters; polyacetals; mulation and filtration should be completed in 1 hour. polyketals; polycarbonates; polyorthocarbonates; polyphos (0708 (3) Fill nominal 1.0 mL filtered solution into steril phaZenes; poly(malic acid); poly(amino acids); polyvi ized glass vials in a controlled target range under aseptic nylpyrrolidone; poly(methyl vinyl ether); poly(alkylene conditions. oxalate); poly(alkylene Succinate); polyhydroxycellulose; (0709 (4) After the filling, all vials are placed with rubber chitin; chitosan; and copolymers and mixtures thereof. stoppers inserted in the “lyophilization position' and loaded 0726 Liposomes are well known as drug delivery in the prechilled lyophilizer. For the lyophilizer, shelf tem vehicles. Liposome preparation is described in European perature is set at +5°C. and held for 1 hour; shelf temperature Patent Application Publication No. EP 1332755 by Wenget is then adjusted to -5° C. and held for one hour, and the al., incorporated herein by this reference. condenser, set to -60° C., turned on. 0727 Slow release injectable gels are known in the art and 0710 (5) The vials are then frozen to 30° C. or below and are described, for example, in B. Jeong et al., “Drug Release held for no less than 3 hours, typically 4 hours. from Biodegradable Injectable Thermosensitive Hydrogel of 0711 (6) Vacuum is then turned on, the shelf temperature PEG-PLGA-PEG Triblock Copolymers,” J. Controlled is adjusted to -5°C., and primary drying is performed for 8 Release 63: 155-163 (2000), incorporated herein by this ref hours; the shelf temperature is again adjusted to -5°C. and CCC. drying is carried out for at least 5 hours. 0728. The use of microspheres for drug delivery is known 0712 (7) Secondary drying is started after the condenser in the art and is described, for example, in H. Okada & H. (set at -60° C.) and vacuum are turned on. In secondary Taguchi. “Biodegradable Microspheres in Drug Delivery.” drying, the shelf temperature is controlled at +5°C. for 1 to 3 Crit. Rev. Ther. Drug Carrier Sys. 12: 1-99 (1995), incorpo hours, typically 1.5 hours, then at 25° C. for 1 to 3 hours, rated herein by this reference. typically 1.5 hours, and finally at 35-40°C. for at least 5 0729. The use of vascular disrupting agents for delivery is hours, typically for 9 hours, or until the product is completely disclosed in United States Patent Application Publication No. dried. 2010/0272717 by Evans et al., incorporated herein by this 0713 (8) Break the vacuum with filtered inert gas (e.g., reference. Such vascular disrupting agents include, but are nitrogen). Stopper the vials in the lyophilizer. not limited to, 5,6-dimethylxanthenone-4-acetic acid. 0714 (9) Vials are removed from the lyophilizer chamber 0730. The use of polymer-coated stents for drug delivery and sealed with aluminum flip-offseals. All vials are visually is disclosed in U.S. Pat. No. 7,906,134 to Hauenstein, incor inspected and labeled with approved labels. porated by this reference. 0715. When the improvement is made by use of dosage 0731. When the improvement is made by use of a drug kits and packaging, the dosage kits and packaging can be, but conjugate form, the drug conjugate form can be, but is not are not limited to, dosage kits and packaging selected from limited to, a drug conjugate form selected from the group the group consisting of the use of amber vials to protect from consisting of: light and the use of stoppers with specialized coatings to 0732 (a) a polymer system; improve shelf-life stability. (0733) (b) polylactides; 0716. When the improvement is made by use of a drug 0734 (c) polyglycolides; delivery system, the drug delivery system can be, but is not (0735 (d) amino acids: limited to, a drug delivery system selected from the group 0736 (e) peptides; and consisting of: (0737 (f) multivalent linkers. 0717 (a) nanocrystals; 0738 Polylactide conjugates are well known in the art and 0718 (b) bioerodible polymers: are described, for example, in R. Tong & C. Cheng, “Con 0719 (c) liposomes: trolled Synthesis of Camptothecin-Polylactide Conjugates 0720 (d) slow release injectable gels; and Nanoconjugates. Bioconjugate Chem. 21: 111-121 0721 (e) microspheres; (2010), incorporated herein by this reference. 0722 (f) vascular disrupting agents; and 0739 Polyglycolide conjugates are also well known in the 0723 (g) polymer-coated stents. art and are described, for example, in PCT Patent Application 0724 Nanocrystals are described in U.S. Pat. No. 7,101, Publication No. WO 2003/070823 by Elmaleh et al., incor 576 to Hovey et al., incorporated herein by this reference. porated herein by this reference. 0725 Bioerodible polymers are described in U.S. Pat. No. 0740 Multivalent linkers are known in the art and are 7.318.931 to Okumu et al., incorporated herein by this refer described, for example, in United States Patent Application ence. A bioerodible polymer decomposes when placed inside Publication No. 2007/0207952 by Silva et al., incorporated an organism, as measured by a decline in the molecular herein by this reference. For example, multivalent linkers can weight of the polymer over time. Polymer molecular weights contain a thiophilic group for reaction with a reactive cys can be determined by a variety of methods including size teine, and multiple nucleophilic groups (such as NH or OH) exclusion chromatography (SEC), and are generally or electrophilic groups (such as activated esters) that permit expressed as weight averages or number averages. A polymer attachment of a plurality of biologically active moieties to the is bioerodible if, when in phosphate buffered saline (PBS) of linker. pH 7.4 and a temperature of 37° C., its weight-average 0741. Suitable reagents for cross-linking many combina molecular weight is reduced by at least 25% over a period of tions of functional groups are known in the art. For example, 6 months as measured by SEC. Useful bioerodible polymers electrophilic groups can react with many functional groups, include polyesters, such as poly(caprolactone), poly(glycolic including those present in proteins or polypeptides. Various acid), poly(lactic acid), and poly(hydroxybutryate); polyan combinations of reactive amino acids and electrophiles are hydrides, such as poly(adipic anhydride) and poly(maleic known in the art and can be used. For example, N-terminal US 2016/0243077 A1 Aug. 25, 2016 44 cysteines, containing thiol groups, can be reacted with halo N,N'-disuccinimidyl carbonate, N-hydroxysuccinimidyl gens or maleimides. Thiol groups are known to have reactiv chloroformate, periodate (for oxidation), alkyl halogens, or ity with a large number of coupling agents, such as alkyl isocyanates. Aldehyde and ketone groups can react with halides, haloacetyl derivatives, maleimides, aziridines, acry hydrazines, reagents forming Schiffbases, and other groups loyl derivatives, arylating agents such as aryl halides, and in reductive amination reactions or Mannich condensation others. These are described in G. T. Hermanson, “Bioconju reactions. Still other reactions suitable for cross-linking reac gate Techniques' (Academic Press, San Diego, 1996), pp. tions are known in the art. Such cross-linking reagents and 146-150, incorporated herein by this reference. The reactivity reactions are described in G. T. Hermanson, “Bioconjugate of the cysteine residues can be optimized by appropriate Techniques” (Academic Press, San Diego, 1996), incorpo selection of the neighboring amino acid residues. For rated herein by this reference. example, a histidine residue adjacent to the cysteine residue 0743. When the improvement is made by use of a com will increase the reactivity of the cysteine residue. Other pound analog, the compound analog can be, but is not limited combinations of reactive amino acids and electrophilic to, a compound analog selected from the group consisting of reagents are known in the art. For example, maleimides can 0744 (a) alteration of side chains to increase or react with amino groups, such as the e-amino group of the side decrease lipophilicity; chain of lysine, particularly at higher pH ranges. Aryl halides 0745 (b) addition of an additional chemical function can also react with Such amino groups. Haloacetyl derivatives ality to alter a property selected from the group consist can react with the imidazolyl side chain nitrogens of histidine, ing of reactivity, electron affinity, and binding capacity; the thioether group of the side chain of methionine, and the and .epsilon.-amino group of the side chain of lysine. Many other 0746 (c) alteration of salt form. electrophilic reagents are known that will react with the 0747 The addition of an additional chemical functionality e-amino group of the side chain of lysine, including, but not to alter reactivity, can be, but is not limited to, an moiety that limited to, isothiocyanates, isocyanates, acyl azides, N-hy acts as an alkylating agent and a colchicine binding site. droxysuccinimide esters, Sulfonyl chlorides, epoxides, Moieties that act as alkylating agents can include, for oxiranes, carbonates, imidoesters, carbodiimides, and anhy example, nitrogen atoms Substituted with two chloroalkyl drides. These are described in G. T. Hermanson, “Bioconju moieties, as occur in nitrogen mustards, nitrosourea moieties, gate Techniques' (Academic Press, San Diego, 1996), pp. and alkylsulfonate moieties. Moieties that act as colchicine 137-146, incorporated herein by this reference. Additionally, binding sites are described in A. Jordan et al., “Tubulin as a electrophilic reagents are known that will react with carboxy Target for Anticancer Drugs: Agents Which Interact With the late side chains such as those of aspartate and glutamate. Such Mitotic Spindle.” Med. Res. Rev. 18: 259-296 (1998), incor as diaZoalkanes and diazoacetyl compounds, carbony dilmi porated herein by this reference. The compound analog is dazole, and carbodiimides. These are described in G. T. Her typically a compound analog of meisoindigo; alternatively, manson, “Bioconjugate Techniques' (Academic Press, San the compound analog can be a compound analog of one of Diego, 1996), pp. 152-154, incorporated herein by this refer Alternatives (1)–(486) as described above. ence. Furthermore, electrophilic reagents are known that will 0748. When the improvement is made by use of a prodrug react with hydroxyl groups such as those in the side chains of system, the prodrug system can be, but is not limited to, a serine and threonine, including reactive haloalkane deriva prodrug system selected from the group consisting of: tives. These are described in G. T. Hermanson, “Bioconjugate 0749 (a) the use of enzyme sensitive esters: Techniques.” (Academic Press, San Diego, 1996), pp. 154 (0750 (b) the use of dimers: 158, incorporated herein by this reference. In another alter (0751 (c) the use of Schiffbases; native embodiment, the relative positions of electrophile and 0752 (d) the use of pyridoxal complexes: nucleophile (i.e., a molecule reactive with an electrophile) are 0753 (e) the use of caffeine complexes; reversed so that the protein has an amino acid residue with an 0754 (f) the use of N-substituted carbohydrate deriva electrophilic group that is reactive with a nucleophile and the tives; targeting molecule includes therein a nucleophilic group. (0755 (g) the use of Mannich N-oxides: This includes the reaction of aldehydes (the electrophile) with 0756 (h) the use of products of reaction with an acylat hydroxylamine (the nucleophile), described above, but is ing or carbamylating agent; more general than that reaction; other groups can be used as 0757 (i) the use of hexanoate conjugates; electrophile and nucleophile. Suitable groups are well known 0758 () the use of polymer-agent conjugates; and in organic chemistry and need not be described further in 0759 (k) the use of prodrugs that are subject to redox detail. activation. 0742 Additional combinations of reactive groups for 0760. The use of prodrug systems is described in T. Jarv cross-linking are known in the art. For example, amino inen et al., “Design and Pharmaceutical Applications of Pro groups can be reacted with isothiocyanates, isocyanates, acyl drugs' in Drug Discovery Handbook (S. C. Gad, ed., Wiley azides, N-hydroxysuccinimide (NHS) esters, sulfonyl chlo Interscience, Hoboken, N.J., 2005), ch. 17, pp. 733-796, rides, aldehydes, glyoxals, epoxides, oxiranes, carbonates, incorporated herein by this reference. This publication alkylating agents, imidoesters, carbodiimides, and anhy describes the use of enzyme sensitive esters as prodrugs. The drides. Thiol groups can be reacted with haloacetyl or alkyl use of dimers as prodrugs is described in U.S. Pat. No. 7.879, halide derivatives, maleimides, aziridines, acryloyl deriva 896 to Allegretti et al., incorporated herein by this reference. tives, acylating agents, or other thiol groups by way of oxi The use of peptides in prodrugs is described in S. Prasadet al., dation and the formation of mixed disulfides. Carboxy groups “Delivering Multiple Anticancer Peptides as a Single Prodrug can be reacted with diazoalkanes, diazoacetyl compounds, Using Lysyl-Lysine as a Facile Linker. J. Peptide Sci. 13: carbonyldiimidazole, carbodiimides. Hydroxyl groups can 458–467 (2007), incorporated herein by this reference. The be reacted with epoxides, oxiranes, carbonyldiimidazole, use of Schiff bases as prodrugs is described in U.S. Pat. No. US 2016/0243077 A1 Aug. 25, 2016

7,619,005 to Epstein et al., incorporated herein by this refer types (i.e., mixed-type). A mixed-type prodrug is one that is ence. The use of caffeine complexes as prodrugs is described bioactivated at multiple sites, either in parallel or sequential in U.S. Pat. No. 6,443,898 to Unger et al., incorporated herein steps. Many ADEPs, VDEPs, GDEPs and nanoparticle- or by this reference. nanocarrier-linked drug moieties can be sequential mixed 0761 Still other prodrug systems applicable to indirubin type prodrugs. Bioactivation of prodrugs can occur by many derivatives or analogs are carbohydrate derivatives Such as reactions, including bioactivation by esterases, hydrolysis, those described in U.S. Pat. No. 6,566,341 to Wang et al., bioactivation by decarboxylases, bioactivation by phos incorporated by this reference. Typically, these prodrugs are phatases, bioactivation by deacetylases, bioactivation by N-substituted carbohydrate derivatives. The carbohydrate N-dealkylases, and many other reactions. can be, for example, a monosaccharide or a disaccharide. The 0767. When the improvement is made by use of a multiple monosaccharide or disaccharide can be, for example, glu drug system, the multiple drug system can be, but is not cose, fructose, ribulose, galactose, mannose, cellobiose, limited to, a multiple drug system selected from the group allose, altrose, ribose, Xylose, arabinose. Sucrose, or lactose. consisting of: A particularly suitable prodrug is an N-triacetylxylopyrano 0768 (a) use of multi-drug resistance inhibitors; syl derivative of indirubin oran analog orderivative of indiru 0769 (b) use of specific drug resistance inhibitors; bin, including an N-triacetylxylopyranosyl derivative or 0770 (c) use of specific inhibitors of selective enzymes: meisoindigo. 0771 (d) use of signal transduction inhibitors; 0762. Still other prodrug systems applicable to indirubin 0772 (e) use of repair inhibition; and derivatives or analogs include Mannich N-oxides of indirubin 0773 (f) use of topoisomerase inhibitors with non derivatives or analogs, including meisoindigo, as described in overlapping side effects. United States Patent Application Publication No. 2010/ 0774) Multi-drug resistance inhibitors are described in 0016252 by Keana et al., incorporated herein by this refer U.S. Pat. No. 6,011,069 to Inomata et al., incorporated herein CCC. by this reference. 0763 Yet other prodrug systems applicable to indirubin 0775 Specific drug resistance inhibitors are described in derivatives or analogs include prodrugs that are prepared by T. Hideshima et al., “The Proteasome Inhibitor PS-341 Inhib reacting a compound with an acylating or carbamylating its Growth, Induces Apoptosis, and Overcomes Drug Resis agent, such as 1,1-acyloxyalkylcarbonochloridate, p-nitro tance in Human Multiple Myeloma Cells. Cancer Res.61: phenyl carbonate, or a similar acylating or carbamylating 3071-3076 (2001), incorporated herein by this reference. agent, as described in U.S. Pat. No. 8,076,375 to Sefton etal, PS-341 refers to bortezomib. incorporated herein by this reference. (0776 Repair inhibition is described in N. M. Martin, 0764 Still other prodrug systems applicable to indirubin “DNA Repair Inhibition and Cancer Therapy.J. Photochem. derivatives or analogs include hexanoate conjugates and Photobiol. B63: 162-170 (2001), incorporated herein by this polymer-agent conjugates as described in United States reference. Patent Application Publication No. 2011/0268658 by Craw (0777. When the improvement is made by biotherapeutic ford et al., incorporated herein by this reference. enhancement, the biotherapeutic enhancement can be per 0765 Still other prodrug systems applicable to indirubin formed by use in combination as sensitizers/potentiators with derivatives or analogs include the use of prodrugs that are atherapeutic agent or technique that can be, but is not limited Subject to redox activation. This utilizes the large quantities of to, a therapeutic agent or technique selected from the group reductase enzyme present in a hypoxic cell to bioactivate the consisting of: drug into its cytotoxic form, essentially activating it. (0778 (a) cytokines; 0766. In general, prodrugs can be classified into two major (0779) (b) lymphokines: types, based on their cellular sites of bioactivation into the 0780 (c) therapeutic antibodies; final active drug form, with Type I being those that are bio 0781 (d) antisense therapies; activated intracellularly (e.g., anti-viral nucleoside analogs, 0782 (e) gene therapies; lipid-lowering statins), and Type II being those that are bio activated extracellularly, especially in digestive fluids or the 0783 (f) ribozymes; and systemic circulation (e.g., etoposide phosphate, Valganciclo 0784 (g) RNA interference. Vir, foSamprenavir, antibody-gene- or virus-directed enzyme 0785 Antisense therapies are described, for example, in prodrugs ADEP/GDEP/VDEP for chemotherapy or immu B. Weiss et al., “Antisense RNA Gene Therapy for Studying notherapy). Both types can be further categorized into sub and Modulating Biological Processes. Cell. Mol. Life Sci. types, i.e. Type IA, IB and Type IIA, IIB, and IIC based on 55:334-358 (1999), incorporated herein by this reference. whether or not the intracellular bioactivating location is also 0786 Ribozymes are described, for example, in S. Pas the site of therapeutic action, or the bioactivation occurs in the colo, “RNA-Based Therapies” in Drug Discovery Handbook gastrointestinal (GI) fluids or systemic circulation. Type IA (S.C. Gad, ed., Wiley-Interscience, Hoboken, N.J., 2005), ch. prodrugs include many antimicrobial and chemotherapy 27, pp. 1273-1278, incorporated herein by this reference. agents (e.g., 5-flurouracil). Type IB agents rely on metabolic 0787 RNA interference is described, for example, in S. enzymes, especially in hepatic cells, to bioactivate the pro Pascolo, “RNA-Based Therapies’ in Drug Discovery Hand drugs intracellularly to active drugs. Type II prodrugs are book (S. C. Gad, ed., Wiley-Interscience, Hoboken, N.J., bioactivated extracelluarly, either in the milieu of GI fluids 2005), ch. 27, pp. 1278-1283, incorporated herein by this (Type IIA), within the systemic circulation and/or other extra reference. cellular fluid compartments (Type IIB), or near therapeutic 0788. When the biotherapeutic enhancement is use in target tissues/cells (Type IIC), relying on common enzymes combination as sensitizers/potentiators with a therapeutic Such as esterases and phosphatases or target directed antibody, the therapeutic antibody can be, but is not limited to, enzymes. Importantly, prodrugs can belong to multiple Sub a therapeutic antibody selected from the group consisting of US 2016/0243077 A1 Aug. 25, 2016 46 bevacizumab (Avastin), rituximab (Rituxan), trastuzumab 0812. When the improvement is by use of a novel mecha (Herceptin), and cetuximab (Erbitux). nism of action, the novel mechanism of action can be, but is 0789. When the improvement is made by use of biothera not limited to, a novel mechanism of action that is a thera peutic resistance modulation, the biotherapeutic resistance peutic interaction with a target or mechanism selected from modulation can be, but is not limited to, use against tumors the group consisting of resistant to a therapeutic agent or technique selected from the 0813 (a) inhibitors of poly-ADP ribose polymerase: group consisting of 0814 (b) agents that affect vasculature or vasodilation; 0790 (a) biological response modifiers; 0815 (c) oncogenic targeted agents; 0791 (b) cytokines: 0816 (d) signal transduction inhibitors; 0792 (c) lymphokines: 0817 (e) EGFR inhibition: 0793 (d) therapeutic antibodies: 0818 (f) protein kinase C inhibition; 0794 (e) antisense therapies; 0819 (g) phospholipase C downregulation; 0795 (f) gene therapies; 0820 (h) Jun downregulation; 0796 (g) ribozymes; and 0821 (i) histone genes; 0797 (h) RNA interference. 0822 () VEGF; 0798. When the biotherapeutic resistance modulation is 0823 (k) ornithine decarboxylase; use against tumors resistant to therapeutic antibodies, the 0824 (1) ubiquitin C: therapeutic antibody can be, but is not limited to, atherapeutic 0825 (m) Jun D; antibody selected from the group consisting of bevacizumab 0826 (n) v-Jun; (Avastin), rituximab (Rituxan), trastuzumab (Herceptin), and 0827 (o) GPCRs: cetuximab (Erbitux). 0828 (p) protein kinase A: 0799. When the improvement is made by radiation therapy 0829 (q) protein kinases other than protein kinase A: enhancement, the radiation therapy enhancement can be, but 0830 (r) prostate specific genes: is not limited to, a radiation therapy enhancement agent or 0831 (s) telomerase; and technique selected from the group consisting of 0832 (t) histone deacetylase. 0800 (a) hypoxic cell sensitizers; 0833 EGFR inhibition is described in G. Giaccone & J. A. 0801 (b) radiation sensitizers/protectors; Rodriguez, “EGFR Inhibitors: What Have We Learned from 0802 (c) photosensitizers; the Treatment of Lung Cancer.” Nat. Clin. Pract. Oncol. 11: (0803 (d) radiation repair inhibitors; 554-561 (2005), incorporated herein by this reference. Pro (0804) (e) thiol depleters: tein kinase C inhibition is described in N. C. Swannie & S. B. 0805 (f) vaso-targeted agents; Kaye, “Protein Kinase C Inhibitors. Curr. Oncol. Rep. 4: (0806 (g) DNA repair inhibitors: 37-46 (2002), incorporated herein by this reference. Phos (0807 (h) radioactive seeds: pholipase C downregulation is described in A. M. Martelli et (0808 (i) radionuclides: al., “Phosphoinositide Signaling in Nuclei of Friend Cells: (0809 () radiolabeled antibodies; and Phospholipase C B Downregulation Is Related to Cell Differ 0810 (k) brachytherapy. entiation.” Cancer Res. 54: 2536-2540 (1994), incorporated 0811 Hypoxic cell sensitizers are described in C. C. Ling herein by this reference. Downregulation of Jun (specifically, et al., “The Effect of Hypoxic Cell Sensitizers at Different c-Jun) is described in A. A. P. Zada et al., “Downregulation of Irradiation Dose Rates.” Radiation Res. 109:396–406 (1987), c-Jun Expression and Cell Cycle Regulatory Molecules in incorporated herein by this reference. Radiation sensitizers Acute Myeloid Leukemia Cells Upon CD44 Ligation. Onco are described in T. S. Lawrence, "Radiation Sensitizers and gene 22: 2296-2308 (2003), incorporated herein by this ref Targeted Therapies. Oncology 17 (Suppl. 13) 23-28 (2003), erence. The role of histone genes as a target for therapeutic incorporated herein by this reference. Radiation protectors intervention is described in B. Calabretta et al., “Altered are described in S. B. Vuyyuri et al., “Evaluation of D-Me Expression of G1-Specific Genes in Human Malignant thionine as a Novel Oral Radiation Protector for Prevention of Myeloid Cells.” Proc. Natl. Acad. Sci. USA 83: 1495-1498 Mucositis.” Clin. Cancer Res. 14: 2161-2170 (2008), incor (1986), incorporated herein by this reference. The role of porated herein by this reference. Photosensitizers are VEGF as a target for therapeutic intervention is described in described in R. R. Allison & C. H. Sibata, “Oncologic Pho A. Zielke et al., “VEGF-Mediated Angiogenesis of Human todynamic Therapy Photosensitizers: A Clinical Review.” Pheochromocytomas Is Associated to Malignancy and Inhib Photodiagnosis Photodynamic Ther. 7: 61-75 (2010), incor ited by anti-VEGF Antibodies in Experimental Tumors. Sur porated herein by this reference. Radiation repair inhibitors gery 132: 1056-1063 (2002), incorporated herein by this ref and DNA repair inhibitors are described in M. Hingorani et erence. The role of ornithine decarboxylase as a target for al., “Evaluation of Repair of Radiation-Induced DNA Dam therapeutic intervention is described in J. A. Nilsson et al., age Enhances Expression from Replication-Defective Aden “Targeting Ornithine Decarboxylase in Myc-Induced Lym oviral Vectors.” Cancer Res.68:9771-9778 (2008), incorpo phomagenesis Prevents Tumor Formation. Cancer Cell 7: rated herein by this reference. Thiol depleters are described in 433-444 (2005), incorporated herein by this reference. The K. D. Held et al., “Postirradiation Sensitization of Mamma role of ubiquitin C as a target for therapeutic intervention is lian Cells by the Thiol-Depleting Agent Dimethyl Fumarate.” described in C. Aghajanian et al., “A Phase I Trial of the Novel Radiation Res. 127:75-80 (1991), incorporated herein by this Proteasome Inhibitor PS341 in Advanced Solid Tumor reference. Vaso-targeted agents are described in A. L. Seyn Malignancies.” Clin. Cancer Res. 8: 2505-2511 (2002), haeve et al., “Tumor Necrosis Factor C. Mediates Homoge incorporated herein by this reference. The role of Jun D as a neous Distribution of Liposomes in Murine Melanoma that target for therapeutic intervention is described in M. M. Caf Contributes to a Better Tumor Response.” Cancer Res. 67: farel et al., “Juni) Is Involved in the Antiproliferative Effect of 9455-9462 (2007), incorporated herein by this reference. A-Tetrahydrocannibinol on Human Breast Cancer Cells.” US 2016/0243077 A1 Aug. 25, 2016 47

Oncogene 27: 5033-5044 (2008), incorporated herein by this - Continued reference. The role of V-Jun as a target for therapeutic inter RLOGLVLKCLAEQQKLOOENLOIFTOLOKLNKKLEGGQQVGMHSKGTOTA vention is described in M. Gao et al., “Differential and Antagonistic Effects of V-Jun and c-Jun, Cancer Res. 56: KEEMEMDPKPDLDSDSWCLLGTDSCRPSL. 4229-4235 (1996), incorporated herein by this reference. The role of protein kinase A as a target for therapeutic intervention 0840 Nek9 is a member of the NIMA family of protein is described in P. C. Gordge et al., “Elevation of Protein kinases. The NIMA family of protein kinases is named after Kinase A and Protein Kinase C in Malignant as Compared the Aspergillus nidulans protein kinase encoded by the nimA With Normal Breast Tissue. Eur: J. Cancer 12: 2120-2126 (never in mitosis A) gene. NIMA is required for entry into (1996), incorporated herein by this reference. The role of mitosis and is involved in the control of chromatin conden telomerase as a target for therapeutic intervention is described sation, spindle and nuclear envelope organization, and in E. K. Parkinson et al., “Telomerase as a Novel and Poten cytokinesis. Eleven protein kinases, Nek1 to Nek 11, with a tially Selective Target for Cancer Chemotherapy.” Ann. Med. catalytic domain related to NIMA have been identified in the 35: 466-475 (2003), incorporated herein by this reference. human genome. Nek9 is a 107-kDa polypeptide whose N-ter The role of histone deacetylase as a target for therapeutic minal catalytic domain is followed by a domain homologous intervention is described in A. Melnick & J. D. Licht, "His to regulator of chromatin condensation (RCC1). The Nek9 tone Deacetylases as Therapeutic Targets in Hematologic C-terminal coiled coil motif binds to DYNLL/LC8 and this Malignancies. Curr. Opin. Hematol. 9: 322-332 (2002), interaction is regulated by Nek9 activity through autophos incorporated herein by this reference. phorylation. Microinjection of anti-Nek9 antibodies in 0834. When the improvement is made by use of selective prophase results in spindle abnormalities and/or chromo target cell population therapeutics, the use of selective target Somal misalignment. Nek9 co-immunoprecipitates gamma cell population therapeutics can be, but is not limited to, a use tubulin and the activated Nek9 localizes to the centrosomes selected from the group consisting of: and spindle poles during early mitosis, indicating that active 0835 (a) use against radiation sensitive cells; Nek9 has important functions at the microtubular organizing 0836 (b) use against radiation resistant cells; center during cell division. Nek6 and Nek7 can bind strongly to RCC1 domain of Nek9 and are phosphorylated and acti 0837 (c) use against energy depleted cells; and vated by Nek9 and depletion of either Nek6 or Nek7 leads to 0838 (d) use against endothelial cells. defective mitotic progression. It is clear that Nek6, Nek7 and 0839. As detailed above, a particularly significant activity Nek9 contribute to the establishment of the microtubule of indirubin analogs and derivatives, particularly meisoin based mitotic spindle. Besides spindle tubule formation. It digo, is the inhibition of the kinase Nek9. Wild-type human has been reported that Nek9 activation by Polo-like kinase 1 Nek9 is a 979-amino-acid protein with the sequence: (PLK1) contributed to the phosphorylation of the mitotic kinesin Eg5 that is necessary for Subsequent centrosome separation and timely mitosis. In addition, Nek9 phosphory (SEQ ID NO: 1) lates NEDD1 which recruits gamma-tubulin to the cen MSWLGEYERHCDSINSDFGSESGGCGDSSPGPSASQGPRAGGGAAEQEEL trosome to ensure the formation of two dense microtubule HYIPIRWLGRGAFGEATLYRRTEDDSLWWWKEWDLTRLSEKERRDALNEI asters in cells entering mitosis. These data indicate that Nek9 has important function not only in spindle formation but also WILALLOHDNIIAYYNHFMDNTTLLIELEYCNGGNLYDKILROKDKLFEE in centrosome maturation and separation. Spindle assembly checkpoint (SAC) is a fail-safe mechanism that monitors the EMVWWYLFOIVSAVSCIHKAGILHRDIKTLNIFLTKANLIKLGDYGLAKK fidelity of chromosome segregation in space and time. Until LNSEYSMAETLVGTPYYMSPELCOGWKYNFKSDIWAVGCVIFELLTLKRT all chromosomes are properly aligned at the spindle equator, the mitotic checkpoint inhibits the anaphase promoting com FDATNPLNLCVKIVOGIRAMEVDSSOYSLELIOMVHSCLDODPEORPTAD plex/cyclosome (APC/C) that prevents cells from entering ELLDRPLLRKRRREMEEKWTLLNAPTKRPRSSTWTEAPIAWWTSRTSEWY anaphase. If the kinetochore-microtubule attachment is not correct, SAC generates the wait-anaphase signal that propa WWGGGKSTPOKLDVIKSGCSAROVCAGNTHFAVWTVEKELYTWVNMOGGT gates throughout the cell to inhibit the APC/C. In various human cancers, mitotic checkpoint function is partially com KLHGOLGHGDKASYROPKHWEKLOGKAIROVSCGDDFTWCVTDEGOLYAF promised, and altered expression or mutations in mitotic GSDYYGCMGVDKVAGPEVLEPMOLNFFLSNPVEOVSCGDNHVVVLTRNKE checkpoint genes have been shown to be related to chromo some instability (CIN) and aneuploidy, which frequently WYSWGCGEYGRLGLDSEEDYYTPOKVDVPKALI.IVAVOCGCDGTFLLTOS occurs in malignant cells. Nevertheless, no evidence for GKVLACGLNEFNKLGLNOCMSGI INHEAYHEVPYTTSFTLAKOLSFYKIR checkpoint malfunctions as a direct cause of CIN in tumor cells has been found. However, complete inactivation of the TIAPGKTHTAAIDERGRLLTFGCNKCGOLGWGNYKKRLGINLLGGPLGGK mitotic checkpoint results in gross chromosomal missegre gation and is not compatible with cell viability. This has led to OVIRVSCGDEFTIAATDDNHIFAWGNGGNGRLAMTPTERPHGSDICTSWP the Suggestion that inhibition of the mitotic checkpoint could RPIFGSLHHVPDLSCRGWHTILIVEKVLNSKTIRSNSSGLSIGTVFOSSS have therapeutic potential in cancer treatment. Moreover, tumor cells that have acquired a decreased checkpoint activity PGGGGGGGGGEEEDSOOESETPDPSGGFRGTMEADRGMEGLISPTEAMGN could be more sensitive to mitotic checkpoint inhibition when SNGASSSCPGWLRKELENAEFIPMPDSPSPLSAAFSESEKDTLPYEELOG compared to healthy, checkpoint proficient cells.

LKVASEAPLEHKPOVEASSPRLNPAVTCAGKGTPLTPPACACSSLOVEVE 0841 Nek9 was found to play an important role in the spindle checkpoint. Nek9 inhibition, through the use of siRNA, induced apoptosis through abnormal mitosis in tumor US 2016/0243077 A1 Aug. 25, 2016 48 cells, whereas the hTERT-RPE1 (REP1) normal human reti that cells exited from mitotic phase after Nek9 knockdown, nal pigment epithelium cell line was refractory to abnormal even when chromosomes were not aligned in the metaphase mitosis. plate. This observation suggests that Nek9 inhibition could 0842) To investigate whether Nek9 inhibition could impair compromise the functioning of the SAC and allow cells to cell proliferation, a growth assay was performed with Caki2 undergo abnormal mitosis. and U1242 cell lines by using two independent Nek9-specific 0847 First, to investigate whether Nek9 is implicated in siRNAs. The results showed that the siRNAs did inhibit the the mitotic checkpoint system, the cell cycle was analyzed production of Nek9 mRNA. In particular, it was observed that after nocodazole treatment, which inhibits the metaphase-to mRNA degradation by NEK9#115-siRNA always had a anaphase transition by preventing microtubule polymeriza stronger effect than NEK9#72-siRNA. Coincident with the tion. FCM analysis showed that NC-siRNA transfectants knockdown efficiency, all tested biological properties were arrested at G2/M phase by nocodazole treatment. By induced by NEK9#115-siRNA in the observed results were contrast, NEK9 transfectants did not stop the cell cycle like more pronounced than those induced by NEK9#72-siRNA. NC-siRNA transfectants. This data suggests that Nek9-deple Immunohistochemical analysis revealed that the fraction of tion compromised SAC function and forced cells to exit from multinuclear cells was increased in Caki2 and U1242 cells. mitosis. Moreover, micronucleated cells were observed in Caki2 cells, 0848. Immunohistochemical stain analysis was per which contained many nuclei. formed to observe morphological change. In an NC-siRNA 0843. To analyze the development of abnormal nuclear treated group, a Substantial number of typical prometaphase morphology, live cell imaging was performed after Nek9 cells with round shape and condensed chromatin in the center knockdown in U1242 cells. Mitosis cells were categorized were observed. On the other hand, the NEK9-siRNA trans into three groups; normal mitosis, abnormal mitosis (cytoki fectants contained fewer prometaphase cells and more adher nesis failure or chromosomes were not aligned in the ent cells, which have numerous micronuclei. The morphol metaphase plate) and death in mitosis (cells showed blebbing ogy of micronucleation was clearly different from that of and nuclear fragmentation before attachment onto plate). In chromatin condensation triggered by apoptosis. The structure cytokinesis failure mitosis, chromosomes were aligned in the of each Small nucleus in micronucleated cells was apparent, metaphase plate, followed by chromosome separation in whereas that in the apoptotic cells was ambiguous. anaphase. However, furrow ingression was repressed in 0849. These results showed that Nek.9 knockdown cytokinesis, resulting in fused daughter cells. These results induced cytokinesis failure and incorrect chromosome sepa suggest one of the reasons for the generation of multi-nuclear ration. It is assumed that Nek6 and Nek7 are activated via the cells is that Nek9 knockdown compromised cytokinesis. C-terminal domain of Nek9 in mitosis. Mouse embryonic 0844. In Nek9-siRNA transfected Caki2 cells, it was fibroblast (MEF) cells derived from Nek 7-deficient mice observed that 33.3% of mitotic cells were unable to undergo showed cytokinesis failure. Depletion of Nek6 by siRNA also normal mitosis. In addition to cytokinesis failure like U1242 resulted in a significant increase in cells with two or more cells, more abnormal mitotic cells were observed whose chro nuclei. These observations were consistent with prior reports mosomes could not be aligned correctly at the metaphase and indicated that the Nek6/779 pathway is involved in the plate. After remaining in mitosis for several hours, cells cytokinesis process. exited from mitosis without cytokinesis, generating micro 0850. In addition, it was found that Nek9 depletion over nucleated cells. This data Suggests that spindle tubules in rode mitotic arrest by nocodazole treatment, indicating that Nek9 depletion cells were imprecisely connected to chromo Nek9 function could be included in the SAC system. Somes and mis-regulated. Moreover, the spindle assembly 0851. It has been reported that overexpression of a kinase checkpoint (SAC) complex in NEK9-siRNA transfectants dead Nek9 mutant compromised spindle dynamics and seemed not to function correctly. microinjection of anti-Nek9 antibodies disrupted spindle for 0845. In order to explore the effect of Nek9 knockdown on mation. At prometaphase and metaphase, Nek9 localizes at normal cells, RPE1 cells (human retinal pigment cells) were the centrosome. It has previously been reported that Nek9 also analyzed. Similar to cancer cells, Nek9 depletion inhib depletion by siRNA resulted in slower cellular growth due to ited cellular growth of RPE1 cells. Interestingly, the nuclear prolonged retention of the cells in the G1 and S phase. It has shape in the NEK9-siRNA transfectants looked like normal been suggested that Nek9 is required for normal cell cycle cells, although the number of nuclei was much less than in the progression. However, it has also been reported that siRNA control group. Moreover, live cell imaging analysis revealed mediated knockdown of Nek9 does not impede cell cycle that normal cells were refractory to abnormal mitosis. The progression. This discrepancy might be explained by two insensitivity of non-malignant RPE1 cells to Nek9 knock possibilities. Firstly, cell cycle analysis based only on the down is not a consequence of insufficient mRNA deletion, as amount of DNA is not an accurate method of discrimination NEKH 115 and NEK9H72-SiRNA showed 89.75% and between diploid G2/M and tetraploid G1 populations. Live 89.66% knockdown, respectively; these values indicated that cell imaging is essential to analyze the detailed abnormalities knockdown was effective. These data Suggested that normal in mitosis. A second possible explanation is due to insufficient cells were less dependent on Nek9 at mitosis and Nek9 inhibi knockdown. High knockdown efficiency seems to be neces tors might be able to kill cancer cells selectively in preference sary to induce apoptosis, probably because Small amounts of to non-malignant cells. This would provide a new avenue of Nek9 protein were functionally sufficient to rescue severe treatment for malignancies. abnormalities. Indeed, the data described above also indi 0846. It is well established that if all chromosomes are not cated that approximately 80% knockdown efficiency was not connected with microtubules precisely, the SAC complex is enough to generate clear cellular growth retardation. activated and postpones the onset of anaphase to ensure seg 0852. Therefore, the Nek9 protein and/or the NEK9 gene regation of the correct number of chromosomes into daughter are proposed as a new target for anti-cancer therapy develop cells. However, live cell imaging data of Caki2 cells revealed ment that would act by inducing interference with normal US 2016/0243077 A1 Aug. 25, 2016 49 mitosis and spindle checkpoint control, leading to mitotic enously added sperm nuclei or plasmid DNA. However, its catastrophe. Mitotic catastrophe is often characterized by the ability to alter nucleosomal DNA structure is conserved. formation of giant micronucleated cells, which typically Studies on the corresponding Spt 16-Pob3 heterodimer in reflect the abnormal segregation of chromosomes induced in yeast have provided evidence on the pleiotropic nature of this a p53-independent manner. Mitotic catastrophe is considered heterodimeras well as its role as a chromatin structure modu to be one of the protective mechanisms reducing cancer inci lator. Mutations in either genes cause various allele-specific dence, leading to the application of the induction of mitotic defects in transcription initiation and elongation and DNA catastrophe as a strategy for the treatment of cancer. replication. Additionally, conditional mutants exhibit stalled 0853. It has also been previously been reported that deple progression in G and S phases, perhaps as the indirect con tion of either Mad2 or BubR1 was able to induce cell death sequence of misregulated processes involving chromatin. through abnormal mitosis. Mad2 loss effectively inactivates Genetic and physical interactions have been observed the checkpoint while inducing a checkpoint stress owing to between Spt 16-Pob3 and a substantial number of transcrip defective microtubule formation, resulting in efficient killing tion or replication proteins, including transcription elongator of tumor cells. It has also been reported that inactivation of an Paf1 complex, histone acetyltransferase complex NuA3, his essential checkpoint protein, Mps 1, induced cell death tone deacetylase Rpd3, general transcription factor TFIIE, accompanied by a chromosome segregation error. Addition putative chromatin factors San1 and Chd1, Nhp6, which is a ally, partial reduction of Mad2 or BubR1, which have dual high mobility group protein that forms the SPN complex with roles in checkpoint activation and chromosome alignment, a heterodimer, DNA polymerase a, and additional DNA rep resulted in sensitivity to taxol; this sensitization was observed lication factors. These observations link FACT/DUF/Spt16 only in tumor cells. Partial reduction of Mps1 or BubR1, Pob3 to replication and transcription and, more specifically, which have dual roles in checkpoint activation and chromo to the structural regulation of chromatin, as Such an entity Some alignment, resulted in sensitivity to taxol. The data serves as template for both processes. showed that Nek9 also has a dual function as a regulator of 0858. The existing evidence is consistent with the notion spindle formation and in the mitotic checkpoint, similarly to that FACT and homologous complexes possess a single cen Mad2, Mps 1 and BubR1. tral activity that is required for the progression of replication 0854. Remarkably, Nek9 depletion did not induce abnor and transcription. However, their roles at multiple steps mal mitosis in normal cells although cellular growth was within these two processes may be mediated by differential impaired. It is Supposed that the existence of multiple backup coordination with various factors. The potential existence of systems for the spindle checkpoint pathway in normal cells such an interaction network is substantiated by the aforemen makes them resistant to abnormal mitosis. These redundant tioned group of ySpt 16-Pob3-interacting proteins, which pathways seem to be lost or compromised in cancer cell lines Suggests further interaction partners. Hence, to further delin underlining the potential of spindle checkpoint interfering eate the role of human FACT in chromatin-related cellular drugs as an attractive new approach in anti-cancer therapy. processes, research efforts have been carried out to delineate 0855. It is considered that the cell cycle of most cancer the identification and functional characterization of its inter cells contains defects in the checkpoint system, which allows acting proteins. By generating monoclonal antibodies cancer cells to proliferate indefinitely without normal growth directed against SSRP1, and Subsequently performing anti control. To selectively target the abnormal cell cycle machin body affinity purification, researchers were able to isolate ery in cancer cells is a rational approach for anticancer FACT-associated protein complexes from HeLa nuclear therapy. The inhibition of NEK9 is therefore proposed as a extracts. One of the polypeptides identified in these searches novel anticancer therapeutic strategy by inducing mitotic is the Nek9 protein. catastrophe via impairment of spindle dynamics, cytokinesis 0859 Nek9 (previously termed Nek8) was originally and the mitotic checkpoint. A greater understanding of the identified as a B-casein kinase that associates with a putative mitotic progress in normal and cancer cells will improve substrate Biccd2. It was independently identified as a Nek6 strategic targeting of cancer cells by Nek9 inhibitors. and Ran GTPase-binding protein under a different name, 0856 Nek9 also has interactions with the human Spt 16 Nercc1. Based on kinase domain sequence similarity, Nek9 is and SSRP1 proteins. The human Spt 16 and SSRP1 proteins a recent addition to the NIMA-like kinase (Nek) family, constitute an abundant, nuclear, and heterodimeric complex whose dozen or so members have diverse but relatively called FACT. The complex FACT was initially discovered as unknown functions. In addition to the N-terminal catalytic a chromatin-specific elongation factor that facilitates tran domain, Nek9 contains a central, RCC1-like, seven-repeats Scription of chromatin templates in vitro. The exact mecha region, followed by a coiled-coil domain. Despite the pres nism by which FACT aids the movement of RNA polymerase ence of a nuclear localization signal, previous publications II and thus facilitates transcription along nucleosomal DNA is have reported a cytoplasmic localization of this kinase. Fur not known. By interacting with histone H2A/H2B dimers, thermore, Nek9 is putatively involved in the microtubule FACT has been postulated to render intranucleosomal histone dynamics and has been implicated in the regulation of mitotic connections more flexible. progression, as disclosed above. 0857. The genes that encode these two FACT polypeptides 0860. An in vivo interaction between Nek9 protein and and, more significantly, the existence of Such stable het FACT has been identified by immunoaffinity purification. erodimeric factors are highly conserved among eukaryotes. Nuclear extracts were subjected to immunoprecipitation with Homologous heterodimers have been purified and identified anti-SSRP1 monoclonal antibodies. The major protein bands from yeast and frog cells. The Xenopus DNA unwinding of 140 and 90 kDa corresponded to hSpt 16 and SSRP1, factor (DUF) complex is also composed of Spt 16 and SSRP1 respectively, as confirmed by Western blot and mass spectro homologs. Unlike the mammalian counterpart, DUF was metric analyses. Such affinity purification also recovers a directly implicated in DNA replication, as immunodepletion 120-kDa polypeptide, which associates with the FACT het of this complex led to a reduced ability to replicate exog erodimer with a molar ratio of ~1-5. Subsequent spectromet US 2016/0243077 A1 Aug. 25, 2016 50 ric analysis of this band revealed a sequence match of the throughout different phases of the cell cycle, the levels of identified spectra to the coding sequence of the human Nek9 association appeared to rise upon release from the G1/S phase protein. To further characterize this protein, polyclonal anti block and then to fall in the Mand G phases. This may reflect bodies against different parts of Nek9 were generated and an interphase-specific elevation in the formation of the active affinity-purified polyclonal antibodies, termed C.Nek9-M phosphorylated Nek9-FACT complex. Additionally, cells (spanning amino acids 291-559) and C.Nek9-N (spanning harboring Nek9-interfering dsRNA displayed significantly amino acids 1-291). Both antibodies specifically recognized a slower cell growth, leading to the conclusion that an adequate 120-kDa polypeptide in the HeLa cell extracts as well as in level of Nek9 is essential for G/S transition and S phase the monoclonal antibody-targeted immunocomplex. Other progression. Nek9 is located in the cytoplasm and likely plays anti-SSRP1 monoclonal antibodies, whose approximate significant roles in the regulation of microtubule dynamics epitope regions have been mapped by using deletion con and mitosis. structs of SSRP1 were subjected to immunoprecipitation 0862. Additionally, Nek9 is activated by the Polo-like experiments as well. kinase Plk1. The cyclin-dependent kinase CDK1 orchestrates 0861 Nek9 was shown to interact with FACT to forman the onset of mitosis through the regulation of multiple pro ~600-kDa complex. Myc-tagged Nek9 was found to bind teins either directly or in collaboration with a number of immobilized FLAG-FACT140 and His-SSRP1, whereas no helper kinases, including Plk1. Plk1 is involved in the com interaction between Myc-tagged Nek9 and other similarly plex mechanism that culminates in CDK1 activation during tagged proteins was observed. In a gel filtration chromatog mitotic entry and is crucial for different mitotic events includ raphy procedure, Nek9 eluted with a molecular mass of -600 ing the formation of the spindle. The molecular basis for some kDa. NEK9 and FACT partially co-localize in the interphase of Plk1 functions relies on the recognition of previously phos nucleus and are similarly free from condensed chromatin but phorylated proteins by the Polo-box domain (PBD) of Plk1. dissociated from each other during chromatin decondensa which also targets the kinase to different sites of action Such tion. Therefore, the subcellular locations of both Nek9 and as the centrosomes and centromeres. FACT alter through the cell cycle. Upon autoactivation, Nek9 0863. The functions of the various members of the NIMA undergoes a phosphorylation-dependent electrophoretic family of protein kinases, which includes Nek9, are still being mobility shift and phosphorylation on amino acid residue elucidated. Aspergillus nidulans NIMA, the founding mem Thr' '. Among Nek family proteins, there is extensive ber of the family, is necessary for entry into mitosis and has sequence conservation within the kinase domain, especially several roles during mitotic progression, including the regu in the activation loop region. It had previously been demon lation of chromosome condensation and spindle formation, strated that Nek9 was able to phosphorylate invitro the Thr' although it is not clear whether all these functions are shared residue within the activation loop, thus indicating the possible with its mammalian counterparts. Different Neks have been ability of Nek9 to autophosphorylate. To assess whether auto implicated in the control of the centrosome and microtubule phosphorylation indeed occurs on the highly-conserved resi cytoskeleton. However, Nek9, which is ~80% identical to due Thr" as well as the relationship between any such phos Nek6 and Nek7, is involved in the control of spindle structure phorylation and the activity of the kinase, polyclonal and function. antibodies that specifically recognize phosphorylated Thr' 0864 Nek9 is activated at centrosomes during early mito were generated. Endogenous, immunoprecipitated Nek9 sis, interacts with both Nek6 and Nek7 and directly phospho kinase became activated (i.e., catalyzed the phosphorylation rylates and activates them. Microinjection of anti-Nek9 anti of histone H1) upon preincubation with a high concentration bodies into prophase cells induces prometaphase arrest and in of ATP. The majority of Nek9 underwent the electrophoretic Some cases aberrant chromosome segregation, resulting in mobility shift only at the high ATP concentration. Phospho mitotic catastrophes or aneuploidy, while Nek9 depletion rylation on Thr210 accompanies this decrease in mobility, as from Xenopus meiotic egg extracts results in delayed spindle Such a slower-moving form was detectable using the antibody assembly, reduced number of bipolar spindles and appear specific for phosphorylated Thr'. The specificity of this ance of aberrant microtubule structures. Downregulation of antibody was confirmed by the lack of immunoreactive sig either Nek6 or Nek7 by RNA interference delays cells at nals from the non-shifted species of Nek9. This further sug metaphase with fragile mitotic spindles and for Nek7 has gests that phosphorylation at Thr' was absent or was at low been shown to result in an increased incidence of multipolar levels for non-phosphorylated Nek9 protein. The FACT-com spindle phenotypes. Mice lacking Nek7 die during late plexed Nek9 protein had similar enzymatic behavior upon embryogenesis or at early postnatal stages, and Nek7 ( / ) kinase activation. The ATP-induced electrophoretic retarda cells show increased tendency for chromosomal lagging as tion of Nek9 can be attributed to the addition of the nega well as abnormalities in primary cilia number. Nek6-deficient tively-charged phosphate moiety, in view of the fact that mice die early during embryogenesis. Thus, Nek9 together treatment with alkaline phosphatase, which removes the with Nek6/7 form a mitotically activated module with key phosphate residue by the enzymatic activity of alkaline phos roles during mitotic progression and more specifically phatase, following kinase autoactivation, abolished the posi spindle organization (Nek6 and Nek7 seem to be functionally tional shift on the gel. The immunoreactivity of Nek9 toward equivalent in most instances, thus when adequate the two the antibody specific for phosphorylated Thr' was directly kinases will be collectively referred to as Nek6/7). Neverthe linked to phosphorylation. Additionally, Nek9 phosphory less, to this date a clear picture of the module activation lated at Thr' associates with FACT in a cell-cycle-depen mechanism, integration with other mitotic signaling systems dent manner. In the presence of FACT, Nek9 undergoes a and precise functions during mitosis has been missing. It has marked elevation in phosphorylation at Thr'. The level of been Suggested that Nek9 could be controlling spindle orga phosphorylation at Thr' is regulated in a cell-cycle-depen nization in part through the action of Nek6/7 and their ability dent manner. Although there is a constitutive presence of to phosphorylate the kinesin Eg5 at a site necessary for nor phosphorylated Nek9 in FACT-Nek9 immunocomplexes mal mitotic progression. Recent work has shown that Plkl, in US 2016/0243077 A1 Aug. 25, 2016

conjunction with CDK1, activates Nek9 early in mitosis, and 0867 Specifically, the NIMA-family kinases Nek9, Nek6 that downstream of Plk1, Nek9 and Nek6/7 are responsible and Nek7 form a signaling module required for normal for centroSome separation during prophase through the con spindle assembly and function during mitosis. CDK1 and trol of Eg5 recruitment to centrosomes. Plk1 are physiologic activators for Nek9. A two-step activa tion mechanism for Nek9 has been proposed. In this activa 0865. In addition to Thr', described above, Nek9 can be tion mechanism, CDK1, together with cyclin B1 and perhaps phosphorylated at additional sites, including Ser’. Thr, cyclin A, phosphorylates Nek9 at Ser', inducing Plk1 bind Ser7, Ser7, Ser, and Thr. A number of these sites ing and Subsequent Plk1 phosphorylation and activation of conform to an STP sequence, and thus are potential phos Nek9. Nek9 activation could directly result from Plk1 phos phorylation sites for the protein kinase CDK1. This protein phorylation of Nek9 at Thr', although conceivably Plk1 kinase has been demonstrated to phosphorylate Nek9 in vitro. phosphorylation of additional sites outside the Nek9 activa Additionally, three potential phosphorylation sites, Ser', tion loop may also contribute to activation by releasing Nek9 Ser', and Ser', conform to an SS/TP sequence, a motif autoinhibition, thus triggering Thr' autophosphorylation. that when phosphorylated at the serine or threonine immedi Nek9 colocalizes at prophase centrosomes with active Plk1 ately preceding the proline can be recognized by the Plk1 and CDK1. protein-binding domain. Plk1 has been shown to specifically 0868. These observations clearly implicate Nek9 as co-immunoprecipitate with Nek9 in mitosis in HeLa cells. directly involved in mitosis and thus in cell division and This interaction was also detected using the yeast two-hybrid proliferation. Therefore, it is important to discover therapeu system, and was mapped to the carboxyl-terminal tail of tic agents that can inhibit the activity of Nek9 in order to more Nek9, where two of the three Nek.9 putative PBD-binding effectively treat diseases and conditions characterized by dys sites reside, and to the PBD domain of Plk1. The Nek.9-PBD regulation of cell division and proliferation, including, but not interaction was increased in mitotic extracts and was totally limited to, malignancies. abolished by mutation of Nek9 Ser' to alanine, which is 0869. A number of Nek9 inhibitors are known in the art. non-phosphorylatable. Mutation of Nek9 Ser’ to alanine did Nek9 inhibitors include, but are not limited to, the agents not have any effect, consistent with the two-hybrid results described below. above, while mutation of Nek9 Ser' had only a minor effect 0870 U.S. Pat. No. 8,591,943 to Deng et al., incorporated on the binding. Plk1 was also able to phosphorylate Nek9 in herein by this reference, discloses pyrazolo 1.5-alpyrim vitro with little or no synergy with CDK1. Specifically, Plk1 idines, including compounds of Formula (N-I): was able to modify Thr', in the kinase activation loop, which is required for Nek9 activation. However, CDK1/cy clin B and the unrelated kinase JNK1 were unable to phos (N-I) phorylate this residue. Phosphorylation of Thr' catalyzed R2 by the kinase Plk1 activated Nek9, as did autophosphoryla R! N tion of Thr' by Nek9 itself. However, CDK1 and Plk1 are 21 e necessary for Nek9 activation in vivo during mitosis. More M over, Plk1, Nek6, Nek7, Nek9, and the mitotic kinesin are R S NN necessary for normal centrosome separation during prophase. This was confirmed by the use of siRNA directed against the NH2 3'-UTR of Nek9 mRNA, which downregulated the level of endogenous Nek9. Additionally, active Nek9 and Nek6 wherein: induce centroSome separation in an Eg5-dependent manner. 0871 (i)R is independently selected from the group con Downregulation of Eg5 almost totally abrogated Nek6- or sisting of halo, hydroxyl, amino, cyano, hydrogen, —(C-C) Nek9-induced centroSome separation, Suggesting that Nek6 alkyl, alkoxy, —C(=O)alkyl, heteroaryl, and aryl, wherein and Nek9 induce centroSome separation through the regula each heteroaryl and aryl can be unsubstituted or substituted tion of the kinesin Eg5. Active Nek9 and Nek6 can rescue with one or more alkyl or halo; Plk1 but not Eg5 downregulation in prophase centrosome 0872 (i) R' is independently selected from the group separation. Specifically, Nek9, Nek6 and Nek7 act down consisting of heterocycloalkyl, heterocycloalkylalkyl, stream of Plk1 and upstream of Eg5 during early centroSome spiroheterocycloalkyl, heterocyclenyl, NR'R''. separation and Suggest that Plk1 inhibition precludes cen cycloalkyl, heteroaryl, aryl, alkyl, alkynyl, heterocycle troSome separation as a result of the failure to activate the nylalkyl, cycloalkylalkyl, heteroarylalkyl, heteroaryla Nek9/Nek677 module. lkynyl, spiroheterocycloalkylalkyl, - N-heteroaryl, 0866 Plk1 controls Eg5 phosphorylation at the Nek6 site -alkyl-NH-heterocyclyl and arylalkyl, wherein each het Ser', that, together with the CDK1 site Thr’, is necessary erocycloalkyl, heterocycloalkylalkyl, spiroheterocy for prophase centroSome separation and Eg5 recruitment. It cloalkyl, heterocyclenyl, cycloalkyl, heteroaryl, aryl, was shown that transfection of active Nek9D346-732 or alkyl, alkynyl, heterocyclenylalkyl, cycloalkylalkyl, Nek6 is able to induce ectopic Eg5 accumulation around heteroarylalkyl, heteroarylalkynyl, - N-heteroaryland centrosomes in parallel to centroSome separation even in arylalkyl can be unsubstituted or substituted with one or interphase and that this is accompanied with Eg5S1033 more moieties independently selected from the group X; phosphorylation. Thus, a physiological correlation exists 0873 (iii) X is alkoxy, alkyl, —C(O)alkyl, —C(O)- between Eg5 recruitment and centroSome separation in hydroxyalkyl, —C(O)alkyl, —C(O)H, hydroxyalkyl, prophase cells, and activation of the Nek9/Nek6 module is —S(O)-alkyl, hydroxyl, heterocycloalkyl, NH-het both necessary and Sufficient to induce both phenomena in a erocycloalkyl, -trihaloalkyl, -dihaloalkyl, -monoha cell-cycle-independent manner. Significantly, failure to phos loalkyl, —N S(O)-alkyl, —C(O)-heteroaryl, -alkyl phorylate Ser' of Eg5 results in a delay in prometaphase. C(O)H, -alkyl (CO)N(CH)—O CH, -alkyl (CO)- US 2016/0243077 A1 Aug. 25, 2016 52

heteroaryl, -alkyl-C(O) NH, amino, heteroaryl, 0884 (vi) R, is selected from hydrogen and C-C, -alkyl-CN. —C(O)-arylalkyl, halo, carboxyesteralkyl, alkyl; or —C(O)—NH, -alkyl-C(O)-alkyl, heteroarylalkyl, 0885 (vii) RandR, together with the nitrogenatom to —C(O)-heteroaryl, —C(O)-alkyl-O-alkyl, -alkyl (CO) which they are attached form an optionally substituted NS(O)-cycloalkyl, -alkyl(CO)N S(O). CF, —N- 5-, 6-, or 7-membered monocyclic heteroaryl or hetero alkyl, -SO-cycloalkyl, -alkyl(CO)NS(O)-alkyl, cyclyl ring, or an optionally Substituted 7- to 11-mem -alkyl-C(O) N(alkyl), -alkyl-NS(O)-alkyl, alkyl bered bicyclic heteroaryl or heterocyclyl ring; and (CO)NS(O)-cycloalkyl, —CO CO.H. —C(O)- 0886 (viii) Rs is selected from hydrogen and C-C, alkyl-aryl, -SO, CF or —C(O)H, wherein each of alkyl; and provided that if R is a group of Subformula heterocycloalkyl, heteroaryl or —C(O)-heteroaryl can (N-IIa) be unsubstituted or substituted with one or more alkyl: 10874) (iv) R is heteroaryl or aryl, wherein each of said heteroaryl or aryl can be unsubstituted or independently (N-IIa) substituted with one or more moieties independently Selected from the group consisting of alkyl, alkoxyl, —CN, aryloxyl, aryl, halo, hydroxyl, —C(CH),CN, trifluoromethyl, difluoromethyl, monofluoromethyl, heterocycloalkyl, and arylalkyl, Ra is a group of Subformula (N-IIb); 0875 (v) R is cycloalkyl or heteroaryl, wherein each cycloalkyl or heteroaryl can be unsubstituted or substi tuted with one or more moieties independently selected (N-IIb) from the group consisting of X; and 0876 (vi) R' is H. 0877 U.S. Pat. No. 8,586,719 to Chan et al., incorporated -( ) herein by this reference, discloses triterpenes inhibiting NEK9 gene expression, including carbohydrate-substituted triterpenes. R is a group of Subformula (N-IIc); 0878 U.S. Pat. No. 8,389,527 to Finket al., incorporated herein by this reference, discloses substituted imidazopy ridazines, including compounds of Formula (N-II): (N-IIc) -O- NH2: R4 R (N-II) NN1 5 then R is not hydrogen. R 0887 U.S. Pat. No. 8,354,408 to Bourke et al., incorpo NS1S-1 rated herein by this reference, discloses nitrogen-containing R N heterocyclic compounds, including compounds of Formula Naan-R6N N (N-III): R8 n NR, N (N-III) / 'o 21 wherein: -H-A-R). 0879 (i) R and R are each independently selected (R- ts n1s from the group consisting of hydrogen, halogen, cyano, M N and C-C alkyl; Z w1 0880 (ii) R is selected from substituted or unsubsti tuted cycloalkyl, substituted or unsubstituted heterocy wherein: clyl, substituted or unsubstituted aryl, and substituted or 0888 (i) Q, W, and Y are each independently selected from unsubstituted heteroaryl; the group consisting of N and CR; 0881 (iii) Ra is selected from hydrogen, substituted or 0889 (ii) Z is NR or S; unsubstituted alkyl, —C(=O)alkyl, —S(O)alkyl, sub 0890 (iii) L is absent, CO, SO, or substituted or unsub stituted or unsubstituted cycloalkyl, substituted or stituted C-C alkylene; unsubstituted heterocyclyl, substituted or unsubstituted 0891 (iv) A and B are each independently absent or sub aryl, and substituted or unsubstituted heteroaryl; stituted or unsubstituted C-C alkylene wherein one or more 0882 (iv) Rs is selected from hydrogen and C-C, carbon atoms can be optionally replaced with O, CO, NR, alkyl: NRCO, CONR, NRSO, S, or S(O), 0883 (v) R is selected from substituted or unsubsti (0892 (v) R' is independently selected from hydrogen, tuted alkyl, substituted or unsubstituted cycloalkyl, sub substituted or unsubstituted C-C alkyl, substituted or stituted or unsubstituted heterocyclyl, substituted or unsubstituted C-C alkenyl, substituted or unsubstituted unsubstituted aryl, and substituted or unsubstituted het C-C alkynyl. Substituted or unsubstituted C-C alkoxy, eroaryl; hydroxyl, halogen, CN, NO, NRR, SOR, SONRR, US 2016/0243077 A1 Aug. 25, 2016

CF. OCF, NRSOR, COR, COSR, CSR, COR, NR, 0901 (iv) R, at each occurrence, is independently NRCSR, CONRR, NRCOR, NRCONRR, SOR, selected from H and C-C alkyl substituted with 0-5 R: substituted or unsubstituted C-C cycloalkyl, substituted or 0902 (v) R, at each occurrence, is independently unsubstituted aryl, and substituted or unsubstituted heterocy selected from H and C-C alkyl substituted with 0-5 R: clyl having up to three heteroatoms or heteroatom-containing (0903 (vi) R is selected from H, F, Cl, Br, CN, OR, moieties selected from the group consisting of N, O, S, and - NRR = C(=O)NRR, NRS(O)R - NRC(=O) SO; R - NRC(=O)CR, and C-C alkyl substituted with 0-5 0893 (vi) R is selected from hydrogen, substituted or R; unsubstituted C-C alkyl, substituted or unsubstituted C-C, 0904 (vii) R is selected from H. C-C alkyl substituted alkenyl, Substituted or unsubstituted C-C alkynyl, Substi tuted or unsubstituted C-C alkoxy, hydroxyl, halogen, CN. with 0-5 R, (CR4.R.), OR, (CR4,R), S(O).R., NO, COR, CONRR, NRCOR, SOR, C-C, —(CR4R),C(=O)R. —(CR4R), NR.R. cycloalkyl, aryl, and heterocyclyl having up to three heteroa —(CR4R),C(=O)NRR —(CR4R), NRC(=O)R. toms or heteroatom-containing moieties selected from the —(CR4R), NRC(=O)CR, -(CR4R), OC(=O) group consisting of N, O, S, and SO, each of which may be NRR,-(CR4R),was OC(=O)NRR,-(CR4R), NRC substituted with up to three substituents independently selected from substituted or unsubstituted C-C alkyl, sub stituted or unsubstituted C-C alkenyl, substituted or unsub substituted with 0-5 R, and —(CR4R), C-C hetero stituted C-C alkynyl, Substituted or unsubstituted C-C, cyclyl substituted with 0-5 R: alkoxy, hydroxyl, OCF, halogen, cyano, NO, NRR, (0905 (viii) R, at each occurrence, is independently SO.R., SONRR, NRSOR, COR COR, NRCOR, selected from F, Cl, Br, C-C alkyl substituted with 0-5 R. RNHCOR, CONRR, NRCONRR, and substituted or C-C alkenyl, C-C alkynyl, NO =O, CN, SOH, unsubstituted heterocyclyl having up to three heteroatoms or S(O).R., S(O)NRR, NRS(O)2R, OR, heteroatom-containing moieties selected from the group con —NRR —NRC(=O)R. -NRC(=O)NRR, sisting of N, O, S, and SO; - C(=O)CR - C(=O)CR - OC(=O)R –C(=O) (0894 (vii)R and Rare eachindependently selected from NRR, C-C cycloalkyl, heterocyclyl, and aryl; hydrogen, Substituted or unsubstituted C-C alkyl, Substi tuted or unsubstituted C-C alkenyl, substituted or unsubsti (0906 (ix) R', at each occurrence, is independently tuted C-C alkynyl, Substituted or unsubstituted C-C, selected from H and C-C alkyl substituted with 0-5 R: alkoxy, cyano, C-C cycloalkyl, substituted or unsubstituted (0907 (x) R, at each occurrence, is independently aryl, and Substituted or unsubstituted heterocyclyl having up selected from H and C-C alkyl substituted with 0-5 R: to three heteroatoms or heteroatom-containing moieties 0908 (xi) Rs is independently selected from H and C-C, selected from the group consisting of N, O, S, and SO; alkyl substituted with 0-5 R: (0895 (viii) m is 1, 2, or 3; and (0909 (xii)R is independently selected from Hand C-C, 0896 (ix) n is 1 or 2; and salts, isomers, and prodrugs alkyl substituted with 0-5 R: thereof. 0910 (xiii) R, at each occurrence, is independently (0897 U.S. Pat. No. 8.252,795 to Finket al., incorporated selected from H. CN, C-C alkyl substituted with 0-5 R. herein by this reference, discloses imidazopyridazinecarbo C-C alkenyl substituted with 0-5 R. C-C alkynyl substi nitriles, including compounds of Formula (N-IV): tuted with 0-5R, —(CH), C-C carbocyclyl substituted with 0-5 R, and —(CH), Cs-Coheterocyclyl Substituted with 0-5R, or R, and R together with the nitrogen atom to R4 R (N-IV) which they are both attached form a heterocyclic ring substi NN1 5 tuted with 0-5 R: 0911 (xiv) R, at each occurrence, is independently R selected from H. CN, C-C alkyl substituted with 0-5 R. NS1 S1 C-C alkenyl substituted with 0-5 R. C-C alkynyl substi Ri \l R tuted with 0-5R, —(CH), C-C carbocyclyl substituted NN 2 y 2 with 0-5 R, and —(CH), Cs-Coheterocyclyl Substituted NC with 0-5 R: R6 (0912 (XV) R, at each occurrence, is independently selected from C-C alkyl substituted with 0-5 R. C-C, wherein: alkenyl substituted with 0-5R, and C-C alkynyl substituted 0898 (i) R is selected from H, F, Cl, Br, CN, and C-C, with 0-5 R, carbocyclyl, and heterocyclyl: alkyl: 0913 (xvi) R, at each occurrence, is independently 0899 (ii) R, is selected from aryl substituted with 0-5 R. selected from H. C-C alkyl substituted with 0-5 R. C-C, and heteroaryl substituted with 0-5 R: alkenyl substituted with 0-5 R. C-C alkynyl substituted 0900 (iii) R2, at each occurrence, is independently with 0-5 R —(CH), C-C carbocyclyl substituted with selected from H, F, Cl, Br, =O, CN, NO, -OR- S(O) 0-5R, and —(CH2), Cs-Coheterocyclyl Substituted with R., C(=O)R = NR,R, (CR2,R2),C(=O)NR.R., 0-5 R: -NRC(=O)R - NRC(=O)CR - OC(=O)NRR, (0914 (XVii) R, at each occurrence, is independently - NRC(=O)NRR —(CR2R2),C(=O)CR - S(O) selected from C-C alkyl substituted with 0-5 Ra C-C, NR.R.—NRS(O).R., C-C alkyl substituted with 0-5R, alkenyl, C-C alkynyl, -(CH2), C-C cycloalkyl, F, Cl, —(CRR), (C-C) carbocyclyl Substituted with 0-5R, Br, CN, NO. —O, COH, -(CH), OC-Cs alkyl, and —(CRR),-heterocyclyl Substituted with 0-5 R: -(CH2),OH, SH, and -(CH2).NRR, US 2016/0243077 A1 Aug. 25, 2016 54

(0915 (xviii) Ra at each occurrence, is independently N, respectively, and RandR'7 or RandR'7 together form a selected from H. C-C alkyl, C-C cycloalkyl, and phenyl, 5- to 8-membered ring system; or R is -(CH2), Z. or R, and Rf, together with the phenyl ring to which they are wherein p is 0, 1, 2, 3, 4, 5, or 6 and Z is selected from the both attached, form a heterocyclic ring: group consisting of Substituted or unsubstituted aryl, Substi 0916 (xix) p, at each occurrence, is independently tuted or unsubstituted heteroaryl, substituted or unsubstituted selected from 0, 1, and 2; and heterocyclyl, - N(RR), wherein R7 and R are indepen 0917 (XX) r, at each occurrence, is independently selected dently selected from H or linear or branched substituted or from 0, 1, 2, 3, and 4. unsubstituted C-C alkyl, or Z is selected from (0918 U.S. Pat. No. 8,084,457 to Choidas et al., incorpo —(CRR'R'') wherein R. R', and R'' are independently rated herein by this reference, discloses 4,6-disubstituted selected from H, linear or branched substituted or unsubsti aminopyridines, including compounds of Formula (N-V): tuted C-C alkyl, or - N(R'R') wherein R'' and Rare independently selected from H or linear or branched substi tuted or unsubstituted C-C alkyl; under the proviso that, if Z is—(CRR'R''), p is 0,1,2,3,4, 5, or 6; and if Z is selected from substituted or unsubstituted aryl, substituted or unsub N stituted heteroaryl, substituted or unsubstituted heterocyclyl, or - N(RR), p is 1, 2, 3, 4, 5, or 6: R3 2 -R 0924 (vi) L is a group of Subformula (N-V(a)), (N-V(b)), (N-V(c)), (N-V(d)), (N-V(e)), (N-V(f)), (N-V(g)), (N-V(h)), R R5-HL-R (N-V(i)), (N-V(i)), (N-V(k)), (N-V(1)), or (N-V(m)): wherein: (0919 (i) R' is selected from the group consisting of H, (N-V(a)) linear or branched C-C substituted or unsubstituted alkyl, - - - -NH---CO-NH-; linear or branched C-C alkenyl, or linear or branched C-C, (N-V(b)) alkynyl: - - - -NH-CS-NH-; 0920 (ii) Rand Rare each independently selected from (N-V(c)) the group consisting of H. linear or branched C-C substi - - - -NH---CO-O-, tuted or unsubstituted alkyl, linear or branched C-C alkenyl, (N-V(d)) linear or branched C-C alkynyl, aryl, F, Cl, Br, I, CN, - - - -CO- - - -: —NH, or - NO; (N-V(e)) 0921 (iii)R’ is selected from the group consisting of F, Cl, - - - - CO-O- - - -: Br, I, substituted or unsubstituted aryl, substituted or unsub (N-V(f)) stituted —O-aryl, substituted or unsubstituted NH-aryl, SO- - - -: substituted or unsubstituted —S-aryl, substituted or unsub (N-V(g)) stituted aryl, substituted or unsubstituted —O-heterocyclyl, - - - -SO----. substituted or unsubstituted —NH heterocyclyl, substi (N-V(h)) tuted or unsubstituted —S-heterocyclyl, substituted or - - - -SO3- - - -: unsubstituted —CH=CH-aryl, substituted or unsubstituted (N-V(i)) heteroaryl, substituted or unsubstituted heterocyclyl, substi tuted or unsubstituted C-C cycloalkyl, or—NH-(CH2)— X, wherein n is 0, 1, 2, 3, 4, 5, or 6 and X is selected from the --- NH group consisting of hydroxyl, amino, or Substituted or unsub (N-V(i)) stituted C-C cycloalkyl; -NR-SO-: 0922 (iv) R is selected from the group consisting of sub (N-V(k)) stituted or unsubstituted aryl, substituted or unsubstituted -NR-SO-: heteroaryl, substituted or unsubstituted heterocyclyl, substi (N-V(1)) tuted or unsubstituted C-C cycloalkyl, or NH-(CH2) -SO-NR8-; or Y, wherein o is 1, 2, 3, 4, 5, or 6 and Y is substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, (N-V(m)) substituted or unsubstituted heterocyclyl, substituted or ----CO-NR'---. unsubstituted C-C cycloalkyl: 0923 (v) R' is selected from the group consisting of sub stituted or unsubstituted C-C alkyl, substituted or unsubsti wherein R'' is selected from H. linear or branched substituted tuted aryl, substituted or unsubstituted heteroaryl, substituted or unsubstituted C-C alkyl, -SO, R', wherein R' is or unsubstituted heterocyclyl, substituted pyrrolidinyl, sub linear or branched C-C alkyl, or R'' is —(CH), stituted or unsubstituted C-C cycloalkyl, disubstituted COOR', whereinris 0,1,2,3,4,5, or 6 and R' is Horlinear cyclohexyl, cyclopentyl, Substituted or unsubstituted Cs-C or branched substituted or unsubstituted C-C alkyl, bicycloalkyl, substituted or unsubstituted adamantyl, - NR7 CO , wherein R7 is H. linear or branched sub -(CH2)-group, where q is 1, 2, or 3, under the proviso that, stituted or unsubstituted C-C alkyl, or a —(CH2)-group, if R is selected to be a methylene chain—(CH.) -group, R17 whereins is 1, 2, or 3; and wherein R'' and R7 both represent or R'' are selected to be a —(CH-)-group whereins is 1, 2, a methylene chain group, RandR'7 may form together a 5 or 3 or a —(CH)-A group whereint is 1, 2, or 3 and A is 0 or to 8-membered ring system of Subformula (N-V(n))

US 2016/0243077 A1 Aug. 25, 2016 59 amino)-7-isopropylimidazo 1.2-bipyridaZin-8-yl)amino)-N- from N, O, and S, and optionally contains one double bond; ethylbenzenesulfonamide, 4-((6-((trans-4-aminocyclohexyl) (b) a 6- to 11-membered heterobicycloalkyl which optionally amino)-7-isopropylimidazo 1.2-bipyridaZin-8-yl)amino)-N- contains one to two additional heteroatoms selected from N. phenylbenzamide; N'-(trans-4-aminocyclohexyl)-7- O, and S; or (c) a 6- to 12-membered heterospirocycloalkyl isopropyl-N-3-pyridinylimidazo[-1,2-bipyridazine-6.8- which optionally contains one to two additional heteroatoms diamine; N'-(trans-4-aminocyclohexyl)-N'-(3- selected from N, O, and S, and wherein the structures of (a), ethoxyphenyl)-7-isopropylimidazo[1,2-bipyridazine-6,8- (b), or (c) is optionally substituted with one to three R: diamine; N'-(trans-4-aminocyclohexyl)-N'-(4- 0929 (ii) R is phenyl or a 5- to 10-membered heteroaryl ethoxyphenyl)-7-isopropyl-imidazo 1.2-bipyridazine-6,8- which contains one to four heteroatoms selected from N, O, diamine; 4-((6-((trans-4-Aminocyclohexyl)amino)-7- and S, and wherein the phenyl or the 5- to 10-membered isopropylimidazo[1,2-bipyridazin-8-yl)amino)phenol; N heteroaryl are optionally substituted with one to three R and (trans-4-aminocyclohexyl)-N'-(4-(aminomethyl)phenyl)-7- wherein the phenyl is optionally fused with a C-C, methylimidazo 1.2-bipyridazine-6,8-diamine; 6-(3-amino cycloalkyl or a 5- to 6-membered heterocycloalkyl which 1,2-benzisoxazol-5-yl)-N-phenylimidazo[1,2-bipyridazin contains one to three heteroatoms selected from N, O, and S 8-amine; 6-(3-amino-1,2-benzisoxazol-6-yl)-N- and which is optionally substituted with oxo; phenylimidazo 1.2-bipyridazin-8-amine; N'-(trans-4- 0930 (iii) R' and Rare each independently hydrogen or aminocyclohexyl)-7-(3-chlorophenyl)-N'-phenylimidazo-6, C-C alkyl; 8-diamine; N'-(trans-4-aminocyclohexyl)-7-(4- 0931 (iv) R' at each occurrence is selected from C-C, chlorophenyl)-N'-phenylimidazo[1,2-bipyridazine-6,8- alkyl, C-C alkoxy, hydroxyl, halo, NR'R'', —C(O) diamine 8-anilino-6-((3S)-3-piperidinylamino)imidazol-2- NR'R'', or a 4- to 7-membered heteroycloalkyl which con bipyridazine-3-carbonitrile; 3-((6-((trans-4- tains one to three heteroatoms selected from N, O, and S; aminocyclohexyl)amino)-7-isopropylimidazo 1.2-b 0932 (v) Rat each occurrence is independently selected pyridaZin-8-yl)amino)phenol; N'-(trans-4- from halo, hydroxyl, cyano, NR'R'', —C(O)NR'R'', C-C, aminocyclohexyl)-3-cyclopropyl-N-phenylimidazo[1,2-b) alkyl, C-C alkoxy, phenyl, a 5- to 6-membered heteroaryl pyridazine-6,8-diamine; N-(4-aminocyclohexyl)-N'- containing one to four heteroatoms selected from N, O, or S, phenyl-3((E)-2-(4-pyridinyl) vinyl)imidazo 1,2-b or two R7 when attached to the same carbon atom and taken pyridazine-6,8-diamine; N'-(trans-4-aminocyclohexyl)-N'- together can be oxo, wherein the C-C alkyl, phenyl, or 5- to phenyl-3-(1-propyn-1-yl)imidazo[1,2-bipyridazine-6.8- 6-membered heteroaryl are optionally substituted with one to diamine; 6-((trans-4-aminocyclohexyl)amino)-8-anilino-N- three R: 4-pyridinylimidazo 1.2-bipyridazine-3-carboxamide, 6-((4- 0933 (vi) R' at each occurrence is independently amino-1-piperidinyl)methyl)-N-phenylimidazo 1.2-b hydroxyl, halo, cyano, C-C alkoxy, NR'R'', C-C alkyl pyridaZin-8-amine; 6-((trans-4-aminocyclohexyl)amino)-8- optionally substituted with one to three halo, C-C, anilinoimidazo[1,2-bipyridazine-7-carbonitrile; 8-anilino-6- cycloalkyl, phenoxy optionally Substituted with cyano, or a 5 ((3S)-3-piperidinylamino)imidazo[1,2-bipyridazine-7- to 6-membered heteroaryloxy containing one to four heteroa carbonitrile; and 4-((6-((trans-4-aminocyclohexyl)amino)-3- toms selected from N, O, and S and which is optionally cyanoimidazo 1.2-bipyridazin-8-yl)amino)-N- substituted with one or two halo or C-C alkyl: methylbenzenesulfonamide. 0934) (vii) Rat each occurrence is independently cyano, 0927 United States Patent Application Publication No. halo, hydroxyl, C-C alkyl-S , carboxyl, —C(O)NH2. 2014/0005183 by Galatsis et al., incorporated herein by this —S(O)NH2, C-C alkyl optionally substituted with one to reference, discloses 4-(substituted amino)-7H-pyrrolo2,3-d three halo or hydroxyl; and pyrimidines, including compounds of Formula (N-VI): 0935 (viii) R' and Rat each occurrence are each inde pendently hydrogen, C-C alkyl, C-C, cycloalkyl, or —C(O)C-C alkyl: (N-VI) 0936 or pharmaceutically acceptable salts thereof. RN-R 0937 United States Patent Application Publication No. N R3 2013/0331359 by Yun et al., incorporated herein by this ref erence, discloses phosphorus-containing-group-substituted quinolines, including: N1-4-7-3-(diethoxyphosphorylm Ny R4 ethylamino)propoxyl-6-methoxyl-4-quinolyloxyl-3-fluo R5 - N - NH rophenyl)-N1-(4-fluorophenyl)cyclopropane-1,1-dicar boxamide; N 1-4-7-3-(diethoxyphosphorylmethylamino) ethoxyl-6-methoxyl-4-quinolyloxyl-3-fluorophenyl)-N1'- wherein: (4-fluorophenyl)cyclopropane-1,1-dicarboxamide; N 1-4- 0928 (i) R' and R are each independently hydrogen, 7-3-(diethoxyphosphorylmethyl)(methyl)amino) C-C alkyl, C-C cycloalkyl, a 4- to 7-membered heterocy propoxyl-6-methoxyl-4-quinolyloxyl-3-fluorophenyl cloalkyl which contains from one to three heteroatoms N1'-(4-fluorophenyl)cyclopropane-1,1-dicarboxamide: N1 selected from N, O, and S, or a 5- to 6-membered heteroaryl 4-7-3-(diethoxyphosphorylmethyl)(ethyl)amino) which contains one to four heteroatoms selected from N, O, propoxyl-6-methoxyl-4-quinolyloxyl-3-fluorophenyl and S.; wherein the C-C alkyl, C-C cycloalkyl, 4- to N1'-(4-fluorophenyl)cyclopropane-1,1-dicarboxamide: N1 7-membered heterocycloalkyl, and 5- to 6-membered het 4-7-3-(diethoxyphosphorylmethyl)(n-propyl)amino) eroaryl are optionally substituted with one to three R'; or R' propoxyl-6-methoxyl-4-quinolyloxyl-3-fluorophenyl and R taken together with the nitrogen to which they are N1'-(4-fluorophenyl)cyclopropane-1,1-dicarboxamide: N1 attached form: (a) a 4- to 7-membered heterocycloalkyl 4-7-3-(diethoxyphosphorylmethyl)(ethyl)amino) which optionally contains one to two heteroatoms selected propoxyl-6-methoxyl-4-quinolyloxyl-3-fluorophenyl

US 2016/0243077 A1 Aug. 25, 2016

(morpholinomethyl)phenylamino)thieno3,2-dipyrimidin-7- 0946 (vi) R' is absent, or RandR', together with the atoms yl)-5,6-di-hydropyridine-1 (2H)-carboxylate: tert-butyl to which they are attached, form a cycloalkyl, heterocy morpholinomethyl)phenylamino)thieno 3.2-dpyrimidin-7- cloalkyl, aryl, or heteroaryl ring, each of which is optionally yl)benzylcarbamate; N-(3-(2-(4-(morpholinomethyl)pheny substituted; lamino)thieno3,2-dipyrimidin-7-yl)phenyl)acetamide; (0947 (vii) Z is NR 0, NRC(O), C(O)NR, CRR, or N-(4-(2-(4-(morpholinomethyl)phenylamino)thieno3,2-d S(O); pyrimidin-7-yl)phenyl)acetamide: N-(3-(2-(4-(morpholi 0948 (viii) R is hydrogen or alkyl: nomethyl)phenylamino)thieno 3.2-dpyrimidin-7-yl)phe 0949 (ix) R is hydrogen, alkyl, or absent, or R and R. nyl)methanesulfonamide; 7-(4-(4-methylpiperazin-1-yl) together with the carbon to which each is attached form C(O); phenyl)-N-(4-(morpholinomethyl)phenyl)thieno3,2-d 0950 (x) ring E is monocyclic or bicyclic heteroaryl; pyrimidin-2-amine; N-(2-methoxy-4-(2-(4- 0951 (xi) R is NRR: (morpholinomethyl)phenylamino)thieno3,2-dipyrimidin-7- 0952 (xii) R is hydrogen, optionally substituted aryl, yl)phenyl)acetamide; 7-bromo-N-(3,4,5-trimethoxyphenyl) optionally substituted heteroaryl, optionally substituted het thieno3.2-dpyrimidin-2-amine; (3-(2-(3,4,5- erocycloalkyl, C(O)R. C(O)CRC(O)NRR, C(NR)R. trimethoxyphenylamino)thieno3,2-dipyrimidin-7-yl) or C(NR)OR; phenyl)methanol: (4-(2-(3,4,5-trimethoxyphenylamino) 0953 (xiii) Rs is hydrogen, halo, alkyl, alkoxy, or thio thieno 3.2-dpyrimidin-7-yl)phenyl)methanol: (3-(2-(4- alkoxy; morpholinophenylamino)thieno3.2-dpyrimidin-7-yl) 0954 (xiv) R is hydrogen, NRR, or OR; phenyl)methanol; (4-(2-(4-morpholinophenylamino)thieno 0955 (XV) each R is independently hydrogen, alkyl, alk 3,2-dpyrimidin-7-yl)phenyl)methanol; N-(pyrrolidin-1-yl) enyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl, each of ethoxy)phenylamino)thieno3,2-dipyrimidin-7-yl)benzyl) which is optionally substituted; methanesulfonamide: tert-butyl-morpholinomethyl) phenylamino)thieno3,2-dpyrimidin-7-yl) 0956 (xvi) each R is independently hydrogen, alkyl, alk benzylcarbamate; N-(4-(morpholinomethyl)phenyl)-7-(3- enyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl, each of (piperazin-1-yl)phenyl)thieno3,2-dipyrimidin-2-amine; which is optionally substituted; 7-(6-(2-morpholinoethylamino)pyridin-3-yl)-N-(3,4,5-tri 0957 (xvii) or, for each occurrence of NRR, R and R. methoxyphenyl)thieno 3.2-dpyrimidin-2-amine; 7-(2-eth are taken together with the nitrogen atom to which they are ylphenyl)-N-(4-(2-(pyrrolidin-1-yl)ethoxy)phenyl)thieno3, attached to form a 3- to 7-membered heterocycloalkyl ring: 2-dipyrimidin-2-amine; 7-(4-(aminomethyl)phenyl)-N-(4- 0958 (xviii) each m is independently 0, 1, or 2; and (morpholinomethyl)phenyl)thieno 3.2-dpyrimidin-2- 0959 (xix) each n is independently 0 or 1. amine; N-(4-(1-ethylpiperidin-4-yloxy)phenyl)-7-(1H 0960 United States Patent Application Publication No. pyrazol-4-yl)thieno3.2-dpyrimidin-2-amine; N-(2,4- 2013/0023514 by Markwalder et al., incorporated herein by dimethoxyphenyl)-7-phenylthieno 3.2-dpyrimidin-2- this reference, discloses Substituted pyrrolotriazines, includ amine; 7-bromo-N-(3,4-dimethoxyphenyl)thieno 3.2-d ing compounds of Formula (N-VIII): pyrimidin-2-amine; and N-(3,4-dimethoxyphenyl)-7- phenylthieno3.2-dpyrimidin-2-amine. (N-VIII) (0940 United States Patent Application Publication No. R4 2013/0040949 by Gray et al., incorporated herein by this YNH O reference, discloses substituted bicyclic compounds of For Y R mula (N-VII): N21 e N R2 lsN --/Y

(N-VII) R3

wherein: 0961 (i) R is selected from hydrogen, NRR, C-C, alkyl substituted with 0-5R, C-C alkenyl substituted with 0-5 R. C-C alkynyl substituted with 0-5 R —(CHR)- carbocyclyl substituted with 0-5R, or—(CHR)-heterocy clyl substituted with 0-5 R: wherein: 0962 (ii) R, at each occurrence, is independently 0941 (i) ring D is aryl or heteroaryl; selected from C-C alkyl substituted with 0-5 R. C-C, alkenyl substituted with 0-5 R. C-C alkynyl substituted 0942 (ii) R is hydrogen, halo, or -A-B; with 0-5 R. C-C haloalkyl, F, Cl, Br, NO, CN, —O, 0943 (iii) A is NRC(O), O, S(O), C(O), C(O)O, C(O) —(CHR), OH, -(CHR), SH, (CHR), OR, -(CHR), S(O) NR, NRC(O)NR, or absent: R. (CHR),C(O)R —(CHR),NR.R. (CHR),C(O) 0944 (iv) B is hydrogen, alkyl, alkoxy, cycloalkyl, or aryl, NRR, -(CHR),C(O)NRNRR, -(CHR), NRC(O)R. -(CHR), NRC(O)OR -(CHR), NRC(O)(CRR), OC(O) wherein each of alkyl, alkoxy, cycloalkyl, oraryl is optionally NRR,-(CHR), NRC(O)(CRR), NRR,-(CHR), NRC substituted; (O)(CRR), NRC(O)OR —(CHR), OC(O)NRR, (0945 (v) R is hydroxyl, alkyl, alkoxy, C(O)OR, C(O) —(CHR),C(O)OR —(CHR), S(O)NR,R, -(CHR).N- NRR, or NRR, each of which may be optionally substi RS(O).R., (CHR)-carbocyclyl substituted with 0-5 R. tuted, or hydrogen or halo: and —(CHR),-heterocyclyl substituted with 0-5 Re; US 2016/0243077 A1 Aug. 25, 2016 64

0963 (iii) R is selected from hydrogen and C-C alkyl substituted with 0-3 R: R4 R (N-IX) 0964 (iv) R, is selected from F, Cl, and Br; NN1 5 0965 (v) alternatively, R and Rare taken together with R the nitrogen atom to which they are attached to form a het NS1 S1 erocyclyl substituted with 0-5 R: R \, 2 R 0966 (vi) R is selected from aryl substituted with 0-5R, NN N- N2 and heteroaryl substituted with 0-5 R: NC R6 0967 (vi) R, at each occurrence, is independently selected from C-C alkyl, C-C alkenyl, C-C alkynyl, wherein: C-C haloalkyl, F, Cl, Br, NO, CN, hydroxyl, sulfhydryl, 0979 (i)R is selected from hydrogen, Cl, Br, F, CN, and —OR, S(O).R, C(O)R. N.R.R. C(O)NR,R, C-C alkyl; - NRC(O)R - NRC(O)OR - OC(O)NRR = C(O) 0980 (ii) R is selected from aryl substituted with 0-5R, OR S(O)NR,R or NRS(O).Rb; and heteroaryl substituted with 0-5 R: 0968 (vii) Ra is selected from hydrogen, C-C alkyl, and 0981 (iii) R, at each occurrence, is independently C-C cycloalkyl, selected from hydrogen, F, CI, Br, —O, CN, NO. —OR, 0969 (viii) R, at each occurrence, is independently S(O).R., C(=O)R - NR.R. (CR2,R2),C(=O) selected from hydrogen, amino, C-C alkyl substituted with NRR, NRC OR NRC(OOR,6Ce O) 0-3 R. C-C alkenyl substituted with 0-3 R, C-C alkynyl substituted with 0-3 R, C-C haloalkyl, —(CH),OH, C-C alkyl substituted with 0-5 R —(CR2,R2), C-C, —(CH),-carbocyclyl substituted with 0-3 R, and —(CH) carbocyclyl substituted with 0-5 R, and —(CR2R2)-het -heterocyclyl substituted with 0-3 R, or R, and R, together erocyclyl substituted with 0-5 R: with the nitrogen atom to which they are attached form a 0982) (iv) R2, at each occurrence, is independently heterocyclyl substituted with 0-3 R: selected from hydrogen and C-C alkyl substituted with 0-5 0970 (ix) R, at each occurrence, is independently R; selected from C-C alkyl substituted with 0-3 R. C-C, 0983 (v) R2, at each occurrence, is independently haloalkyl, C-C alkenyl substituted with 0-3 R.C.-C alky selected from hydrogen and C-C alkyl substituted with 0-5 nyl substituted with 0-3 R—(CH),-carbocyclyl substituted R; with 0-3 R, and —(CH)-heterocyclyl substituted with 0-3 0984 (vi) R is selected from H, F, Cl, Br, CN, OR,

R;8 - NRR-C(=O)NRR-NRS(O)2R - NRC(=O) R - NRC(=O)CR, and C-C alkyl substituted with 0-5 0971 (x) Rat each occurrence, is independently selected R; from hydrogen, C-C alkyl substituted with 0-3 R, C-C, 0985 (vii) R is selected from H. C-C alkyl substituted haloalkyl, C-C alkenyl substituted with 0-3 R.C.-C alky with 0-5 R. —(CR4R), OR, —(CR4R), S(O).R., nyl substituted with 0-3 R—(CH),-carbocyclyl substituted —(CR4R),C(=O)R. —(CR4R), NRR, with 0-3 R, and —(CH),-heterocyclyl substituted with 0-3 —(CR4R),C(=O)NRR —(CR4R), NRC(=O)R. R;8 —(CR4R), NRC(=O)CR, -(CR4R), OC(=O) 0972 (xi) R, at each occurrence, is independently NRR —(CR4R), NRC(=O)NRR —(CR4R),C selected from C-C alkyl, C-C alkenyl, C-C alkynyl, (=O)CR,-(CR4R), S(O)2NRR,-(CR4R), NRS —(CH), C-C cycloalkyl, F, Cl, Br, CN, NO, COH =O. (O)2NRR —(CR4R), NRS(O)2R —(CR4R)- —C(O)NRR (CF2), CF3,-(CH2), OC-C, alkyl, -(CH2) C-C carbocyclyl substituted with 0-5 R —(CR4,R)- ,OH, SH,-(CH2), SC-C alkyl, -(CH2).NRRA -(CH2) heterocyclyl substituted with 0-5 R: -phenyl, and (CH2)-heterocyclyl: 10986) (viii) R, at each occurrence, is independently selected from F, Cl, Br, C-C alkyl substituted with 0-5 R. 10973 (xii) Ra at each occurrence, is independently C-C alkenyl, C-C alkynyl, NO =O, CN, SOH, selected from hydrogen, C-C alkyl, C-C cycloalkyl, and S(O).R., S(O)NRR, NRS(O).R, OR, phenyl: —NRR —NRC(=O)R. -NRC(=O)NRR, 0974 (xiii)R, at each occurrence, is independently —C(=O)CR —C(=O)R —OC(=O)R. —C(=O) selected from hydrogen, —(CH2), OH, C-C alkyl, C-C, NRR, C-C cycloalkyl, heterocyclyl, and aryl; haloalkyl, and —(CH2),-aryl; 0987 (ix) R, at each occurrence, is independently selected from hydrogen and C-C alkyl substituted with 0-5 0975 (xiv) p, at each occurrence, is independently R; selected from 0, 1, and 2: 10988 (x) R, at each occurrence, is independently 0976 and selected from hydrogen and C-C alkyl substituted with 0-5 R; 0977 (XV) r, at each occurrence, is independently selected 0989 (xi) Rs is selected from hydrogen and C-C alkyl from 0, 1, 2, 3, and 4. substituted with 0-5 R: 0978 United States Patent Application Publication No. 0990 (xii) R is selected from hydrogen and C-C alkyl 2012/0283241 by Fink et al., incorporated herein by this substituted with 0-5 R: reference, discloses imidazopyridazinecarbonitriles, includ 0991 (xiii) R, at each occurrence, is independently ing compounds of Formula (N-IX): selected from hydrogen, CN, C-C alkyl substituted with 0-5

US 2016/0243077 A1 Aug. 25, 2016 69

6,7-tetrabromo-1-methyl-benzimidazole: 2-(methyl(4,5,6,7- 1025 (ii) each R is independently selected from hydro tetrabromo-1-methyl-1H-benzodimidazol-2-yl)amino) gen, C-C alkyl, C-C alkenyl, hydroxyl, halogen, nitro, ethanol: (2-dimethylamino-4,5,6,7- Substituted or unsubstituted amino, cyano, nitro, trifluorom tetrabromobenzoimidazol-1-yl)acetic acid ethyl ester; and ethyl, methoxy, trifluoromethoxy, aryl and substituted or (2-dimethylamino-4,5,6,7-tetrabromobenzoimidazol-1-yl) unsubstituted 5 or 6 membered heterocyclyl containing 1 to 2 acetic acid. Natoms; 1020 United States Patent Application Publication No. 1026 (iii) R is selected from hydrogen, C-C alkyl, 2011/0034459 by Adibhatla et al., incorporated herein by this C-C alkyl-CN, C-C cycloalkyl, C-C alkylene-cy reference, discloses 6,7-dialkoxyquinazoline derivatives, cloalkyl, aryl, C-C alkylene-aryl, heterocyclyl and C-C, including compounds of Formula (N-XII): alkylene-heterocyclyl, wherein C-C alkyl, C-Cs cycloalkyl, heterocyclyl and aryl may be optionally Substi tuted with 1 to 3 substituents selected from R or Ro: (N-XII) 1027 (iv) R is independently selected from halogen, sub stituted or unsubstituted C-C alkyl, OH. (O), OCN, substi tuted or unsubstituted O C-C alkyl, CN, CF, CFCN, SCN, SONRR, SR, CHO, COR, COR, CONRR, HN Šs CONRR, NRCOR, NO, NRR, NRCN, CH(CN) NRR, NRS02R, COCF, COCHF, NRCOCOR, RO n N NRCOOR, NRCONRR, heterocyclyl and CO-heterocy clyl, wherein each heterocyclyl may be optionally substituted e with 1 to 4 substituents selected from NH, CN, OH, CO.R., R2O N CHCN and 5-membered N-containing heterocyclyl; 1028 (v) R is C-C alkylene-R, O C-C alkylene-Rs, wherein: except when R is NRs R or O C-C alkyl, then R is 1021 (i) R' is a group of Subformula (N-XII(a)), (N-XII O—C-C alkylene-Ro); (b)), or (N-XII(c)); 1029 (vi) or R and R together with the groups to which they are attached form a substituted or unsubstituted 5- or 6-membered N-containing heterocyclyl: (N-XII(a)) 1030 (vii)Rs and Rare each independently selected from H. C-C alkyl, C-C alkyl-CN, C-C cycloalkyl, aryl, het rO, erocyclyl. Substituted or unsubstituted C-C alkylene, cycloalkyl, SO C-C alkyl and C-C-alkylene-heterocy CHCHCH-1 N clyl; or (N-XII(b)) 1031 (viii) Rs and R together with the nitrogen to which they are attached form a 4-8 membered ring having 1 to 3 heteroatoms independently selected from NRs 0, S(O), o CHCH-CH-1 O wherein m is 0, 1 or 2 and wherein the ring may be optionally (N-XII(c)) substituted with C-C alkyl or NRR: 1032 (ix) Rs is selected from H. C-C alkyl, C-C alky NH: lene-OH, C-C alkylene-NRRC-Cs cycloalkyl, aryl, het erocyclyl, C-C alkylene-cycloalkyl, C-C alkylene-aryl, -CHCHCH-1 N C-C alkylene-heterocyclyl and C-C-alkyleneCN: 1033 (x) R, is selected from H, substituted or unsubsti tuted C-C alkyl, substituted or unsubstituted O—C-C, and alkyl, substituted or unsubstituted S C-C alkyl, CNOH, 1022 (ii) R is CH or -CHCH. C-C-alkylene-CN, substituted or unsubstituted C-Cs 1023 United States Patent Application Publication No. cycloalkyl, substituted or unsubstituted aryl, substituted or 2010/0239631 by Bourke et al., incorporated herein by this unsubstituted heterocyclyl, C-C alkylene-cycloalkyl, reference, discloses thiopyrimidines, including compounds C-C alkylene-aryl, C-C alkylene-heterocyclyl, C-C alk of Formula (N-XIII): enyl, C-C alkynyl, NRR, C-C alkylene-NRRs and C-C alkylene-ORs: 1034 (xi)W is absent, CO, SO, or C1-C6 alkylene: (N-XIII) H 1035 (xii) R is selected from H. C-C alkyl, C-Cs R2 N N R3 cycloalkyl, aryl and heterocyclyl, each of which may be optionally substituted with 1 to 4 substituents selected from R X 2Y and Ro, and 1036 (xiii) wherein each alkenyl and alkynyl may be optionally substituted with 1 to 3 substituents independently S selected from C-C alkyl, COR7 CONRR aryl, hetero YR cyclyl, C-C alkylene, OH, and C-C alkylene-NH. 1037 United States Patent Application Publication No. wherein: 2010/0152434 by Peterson, incorporated herein by this ref 1024 (i) X and Y are independently selected from N and erence, discloses kinase-binding nucleosides and derivatives, CR; including compounds of Formula (N-XIV): US 2016/0243077 A1 Aug. 25, 2016 70

(N-XIV) (N-XV)

R15 R14 a. R

R2,

RO R. Rio

wherein: 1049 (i) R is selected from hydrogen, hydroxyl, wherein: O-acetyl, O-angeloyl, O-tigloyl, O-Senecioyl, O-alkyl, 1038 (i) R', R. R. and Rare members selected inde O-dibenzoyl, O-benzoyl, O-alkanoyl, O-alkenoyl, O-ben pendently from the group consisting of H. HO—, CHO—, Zoyl-alkyl substituted alkanoyl, O-aryl, O-acyl, O-heterocy CH , HOCHCH , HOCH2CHOCHCH clyl, O-heteroaryl and O-Sugar moiety, wherein the Sugar NHCHCH-, RNHCHCH , (R)-NCHCH moiety is substituted with two groups selected from benzoyl, NHCHCH-NHCHCH alkanoyl, alkenoyl, benzoyl, alkyl-substituted alkanoyl, aryl, RNHCHCH-NHCHCH , (R7) acyl, heterocyclyl, heteroaryl, and derivatives thereof; NCHCH-NHCHCH , RCO , a mono-, di-, or tri 1050 (ii) R is selected from hydrogen, hydroxyl, O-an cyclic aryl from C to C, a mono-, di-, or tri-cyclic aryl from geloyl, O-tigloyl, O-senecioyl, O-alkyl, O-dibenzoyl, O-ben C to C mono-, di-, tri-, or poly-substituted with a member Zoyl, O-alkanoyl, O-alkenoyl, O-benzoyl-alkyl-substituted selected independently from the group consisting of F, Cl, Br, alkanoyl, O-aryl, O-acyl, O-heterocyclyl, O-heteroaryl, and I, alkoxy (RO ), nitro (NO), nitroso (NO), azido (N), O-Sugar moiety, wherein the Sugar moiety is substituted with C2-C2 alkyl, C-C2 alkenyl, C-C2 alkynyl, C2-C2 acyl; an two groups selected from benzoyl, alkanoyl, alkenoyl, ben O, N, or S mono- or bi-cyclic heterocycle having from 2 to 9 Zoylalkyl Substituted alkanoyl, aryl, acyl, heterocyclyl, het carbon atoms, and an O, N, or S mono- or bi-cyclic hetero eroaryl, and derivatives thereof; cycle having from 2 to 9 carbonatoms and mono-, di-, tri-, or 1051 (iii) R is selected from CHR and COR, wherein poly-substituted with a member selected independently from R comprises a group selected from hydroxyl, O-angeloyl, the group consisting of F, Cl, Br, I, alkoxy (RO ), nitro O-tigloyl, O-senecioyl, O-alkyl, O-dibenzoyl, O-benzoyl, (NO), nitroso (NO), azido (N), C-C alkyl, C-C alk O-alkanoyl, O-alkenoyl, O-benzoyl-alkyl-substituted enyl, C2-C2 acyl: alkanoyl, O-aryl, O-acyl, O-heterocylic, O-heteroraryl, and 1039 (ii)R’ is C-C alkyl: derivatives thereof; or wherein at least one of R', RandR is 1040 (iii) R is H.N. , HOHN , C, -Co alkyl, C.-Co selected from O-angeloyl, O-tigloyl, O-senecioyl, O-diben alkenyl, or phenyl: Zoyl, O-benzoyl, O-alkanoyl, O-alkenoyl, O-benzoyl alkyl 1041 (iv) R is C-C alkyl: substituted alkanoyl, O-aryl, O-acyl, O-heterocyclyl, O-het 1042 (v) Rand Rare members selected independently eroaryl, and derivatives thereof; from the group consisting of H. HO—, CH , or 1052 (iv) or wherein at least two of R. R. and Ra are CHCH : selected from O-angeloyl, O-tigloyl, O-senecioyl, O-diben 1043 (vi) X" and X’ are members selected independently Zoyl, O-benzoyl, O-alkanoyl, O-alkenoyl, O-benzoyl alkyl from the group consisting of O and S. substituted alkanoyl, O-aryl, O-acyl, O-heterocylic, O-heter oraryl, and derivatives thereof, orany of R. RandR has two 1044 (vii) U is a member selected from the group consist angeloyl, tigloyl, Senecioyl, dibenzoyl, benzoyl, alkanoyl, ing of H, HO F, CF : alkenoyl, O-benzoyl alkyl Substituted alkanoyl, aryl, acyl, 1045 (viii) W is a member selected from the group con heterocyclyl, heteroaryl, and derivatives thereof; sisting of H, HO , F, CF , CHCHOCCH , CH 1053 (v) R is H or OH: (CHO)NCOCH , HOCHCH-O-, NH COCH , 1054 (vi) Rs is H or OH: CHNHCOCH, , (CH), NCOCH , 1055 (vii) Rs is a hydrogen, or sugar moiety(ies) or acid HOCHCH-NHCOCH HSCHCH-NHCOCH thereof, wherein the Sugar moiety(ies) is/are selected from a RO , and an O-trialkylsilyl containing 3-16 carbons; group consisting of glucose, galactose, rhamnose, arabinose, 1046 (ix)Y is a member selected from the group consist Xylose, fucose, allose, altrose, gulose, idose, lyxose, man ing of H. HO , F, CF, , HOCH2CH2O , RO , and an nose, psicose, ribose, Sorbose, tagatose, talose, fructose, aldu O-trialkylsilyl containing 3-16 carbons; and ronic acid, glucuronic acid, galacturonic acid, and derivatives 1047 (x) Z is a member selected from the group consisting or combination thereof; of H, F, HO CF, , and RO-. 1056 (viii) wherein R. R. R. R. R. Ra, and Rs of 1048 United States Patent Application Publication No. the compound are independently selected from CH, 2010/0004190 by Chan et al., incorporated herein by this CHOH, CHO, COOH, COO-alkyl, COO-aryl, COO-hetero reference, discloses polycyclic kinase inhibitors, including cyc, COO-heteroaryl, CH, O-aryl, CH, O-heterocyclyl, compounds of Formula (N-XV): CH-O-heteroaryl, alkyl, hydroxyl, and acetyl.

US 2016/0243077 A1 Aug. 25, 2016 74 wherein X is a bond or C-C alkylene and Rs is selected from 1079 (v) each occurrence of X is independently NH, O, or hydrogen, C-C alkyl, and C-C cycloalkyl-Co-C alkyl; S; and 1069 (x) the N-oxide derivatives, prodrug derivatives, 1080 (vi) R' is H, C-C alkyl, halo-(C-C alkyl), C-C, protected derivatives, individual isomers and mixtures of iso cycloalkyl, halo-(C-C cycloalkyl), heterocyclyl, heterocy mers thereof; clyl-(C-C alkyl), aryl, aryl-(C-C alkyl), heteroaryl, or het and the pharmaceutically acceptable salts thereof. eroaryl-(C-C alkyl); 1070 PCT Patent Application Publication No. WO 2011/ 1081) (vii)R’ is H, C-C alkyl, halo-(C-C alkyl), C-C, 126882 by Gelberet al., incorporated herein by this reference, cycloalkyl, halo-(C-C cycloalkyl), heterocyclyl, heterocy discloses therapeutic peptides with Nek9-inhibitory activity, clyl-(C-C alkyl), aryl, aryl-(C-C alkyl), heteroaryl, or het including analogs and derivatives of SGRPPMIVWFNRP eroaryl-(C-C alkyl); FLIAVSHTHGQTILFMAKVINPVGA (SEQ ID NO:5). 1071 PCT Patent Application Publication No. WO 2013/ 1082 (viii) R is R, CO-R, or a moiety of Subfor 148864 by Gudkov et al., incorporated herein by this refer mula (N-XVIII(a)), (N-XVIII(b)), (N-XVIII(c)), (N-XVIII ence, discloses curaxins and derivatives thereof, including (d)), (N-XVIII)(e)), (N-XVIII.(f)), (N-XVIII.(g)), (N-XVIII compounds of Formula (N-XVII): (h)), (N-XVIII(i)), (N-XVIII()), (N-XVIII(k)), or (N XVIII(1))

(N-XVII) (N-XVIII(a)) wO R6; (N-XVIII(b)) - wherein: 1072 (i) each of R', R,R,R,R,R,R,R, and Rare independently hydrogen, hydroxyl, or alkyl, and w 1073 (ii) n is 0, 1, 2, 3, 4, or 5. (N-XVIII(c)) 1074 PCT Patent Application Publication No. WO 2009/ 032694 by Gray et al., incorporated herein by this reference, r discloses pyrimidine, pyrrolopyridine, and pyrazolopyrimi dine derivatives, including compounds of Formula ". N (N-XVIII): 7 (N-XVIII(d)) R'; (N-XVIII) Ra 1 HN vC 1. Rb X N1 N (N-XVIII(e)) 1 R R1 2 Y N Z N Y R; H H * r wherein: (N-XVIII(f)) 1075 (i) Z is N or CH: 1076 (ii) A is aryl or heteroaryl optionally substituted with one or more R groups; 1077 (iii) B is phenyl when Z is N; or B is phenyl, pyra Zolyl or thiazolyl when Z is CH; wherein B is optionally \C. substituted with one or more R groups; 1078 (iv) Y is -SO. , -SONH-, -NH SO , NH C(O) , —C(O) NH-, -O-, or NR; US 2016/0243077 A1 Aug. 25, 2016

-continued cyano, acyloxy, carboxy, ester, carbamoyl, carboxamide, ure (N-XVIII(g)) ido, amidino, guanidine, Sulfonyl, Sulphonylamino, or ami R7 nosulfonyl: 1085 (xi) each occurrence of R is independently (C-C) alkoxy, a moiety of Subformula (N-XVIII(ml)), hydroxy, a moiety of Subformula (N-XVIII(a)), a moiety of Subformula v. s (N-XVIII(n)), dialkylamino, or —NR': (N-XVIII(h)) R7 (N-XVIII(ml))

(N-XVIII(n)) - (C-C6 alkyl): vC N (N-XVIII(i)) R7

1086) (xii) each occurrence of R is independently O hydroxy, Subformula (N-XVIII(o)), Subformula (N-XVIII) '' (d)), or -CONH; (N-XVIII()) R7 ry N ): R7; ". Nulls 1087 (xiii) each occurrence of R is independently hydro (N-XVIII(k)) gen, (C-C) alkyl, (C-C) alkoxy, hydroxyl, or (C-C) O hydroxyalkyl: with the provisos that: NH 1088 (a) when Z is N. B is phenyl, and —X-R is NH R, then X-R is bound in the meta or para N- N position on the phenyl group and RandR, if present on B, are not bound to the meta or para position and are not —COR, C-C alkyl, halo-(C-C alkyl), Cs-Co N cycloalkyl, halo-(C-C cycloalkyl), heterocyclyl, het (XIX(1)) O erocyclyl-(C-C alkyl), aryl, aryl-(C-C alkyl), het eroaryl, heteroaryl-(C-C alkyl), C-C alkoxy, C-C, alkylthio, cyano, acyloxy, carboxyl, ester, carbamoyl, carboxamide, or amidino; 1089 (b) when Z is N, B is phenyl, and X-R is not NH R, then R is bound to the meta or paraposition and is a moiety of Subformula (XIX(a)), (XIX(c)), (XIX (e)), (XIX(f)), (XIX(g)), or (XIX()), or if R and R, if present on B, are each bound to the meta or paraposition 1083 (ix) R' is hydrogen and R is halogen, or RandR', and are independently nitro, azido, ureido, guanidine, taken together with the atoms to which they are bound form: Sulphonylamino; and (i) a pyrazolo ring fused to the pyrimidine ring when Zis Nor 1090 (c) when Z is CH, and R and R are taken (ii) a pyrrolo ring fused to the pyrimidine ring when Z is CH, together with the atoms to which they are bound to form the pyrazolo or pyrrolo ring optionally bearing one or two R' a pyrrolo ring fused to the pyridine ring, then: (1) X-R is not isopropoxy bound to the orthoposition groups: and R or R', if present on B, is methyl, ethyl, methoxy, 1084 (x) each occurrence of R is independently halogen, ethoxy, chloro or bromo any of which is bound to the C-C alkyl, halo-(C-C alkyl), C-C cycloalkyl, halo-(C- ortho position; or (2) R is bound to the meta or para C. cycloalkyl), heterocyclyl, heterocyclyl-(C-C alkyl). position and is a moiety of Subformula (XIX(b)), (XIX aryl, aryl-(C-C alkyl), heteroaryl, heteroaryl-(C-C alkyl). (i)), (XIX(k)), or (XIX(1)) or R or R, if present on B, (C-C) alkoxy, (C-C) alkylthio, hydroxyl, nitro, azido, are each bound to the meta or para position and are US 2016/0243077 A1 Aug. 25, 2016 76

independently C-C alkoxy, C-C alkylthio, hydroxyl, nitro, azido, cyano, acyloxy, carboxy, ester, carbamoyl, (N-XXI) carboxamide, ureido, amidino, guanidine, Sulfonyl, Sul phonylamino, aminosulfonyl, -CO-R, or phenyl O-L-R): Substituted with aminosulfonyl, amino, alkynyl or car L4 boxamide. N1 N-\ 1091 PCT Patent Application Publication No. WO 2009/ (R-L'H X 017795 by Ajamiet al., incorporated herein by this reference, NeN^N V discloses indazole compounds of Formulas (N-XIX) or L2 V (N-XX) R2 (N-XXII)

(N-XIX)

(N-XXIII)

wherein, for compounds of Formula (N-XXI). (N-XXII), and (N-XXIII): 1100 (i) X is =N or=C(L-R) ; 1101 (ii) Ring A is, in each instance, independently selected from cycloalkyl, heterocycloalkyl, aryl, or het eroaryl; 1102 (iii) L', L. L. L. L., and Lare, in each instance, independently selected from a bond, —C(O)— —C(O)N wherein, for both compounds of Formula (N-XIX) and (R) , —C(O)C) , S(O) , S(O)N(R) , —O , (N-XX): NR7 N(R')C(O)N(R), substituted or unsubsti 1092 (i) R' is hydrogen or NR'R'': tuted alkylene, substituted or unsubstituted heteroalkylene, substituted or unsubstituted cycloalkylene, substituted or 1093 (ii) R' and R', independently, are hydrogen or an unsubstituted heterocycloalkylene, substituted or unsubsti optionally substituted alkyl, or RandR, taken together with tuted arylene, or substituted or unsubstituted heteroarylene, the nitrogen to which they are attached, are attached, form a whereing is 0, 1, or 2; non-aromatic heterocycle, optionally substituted at one or more substitutable carbon atoms with methyl, hydroxyl, or 1103 (iv) R', R. R. R. R. R. R7, and Rare, in each methoxy, and optionally N-substituted with C-C alkyl or instance, independently selected from hydrogen, halogen, C-C alkyl substituted with - NR'R'': cyano, hydroxyl, amino, carboxyl, -CONH2, nitro, Sulfhy dryl, -SOCl. —SOH, -SOH, -SONH, substituted or 1094) (iii)R and Rare independently hydrogen, methyl, unsubstituted alkyl, substituted or unsubstituted heteroalkyl, or ethyl: substituted or unsubstituted cycloalkyl, substituted or unsub stituted heterocycloalkyl, substituted or unsubstituted aryl, or 1095 (iv) R is hydrogen or C-C alkyl: substituted or unsubstituted heteroaryl; 1096] (v) R is hydrogen or C-C alkyl: 1104 (v) a is 0, 1, or 2: 1097 (vi) R' is hydrogen or C-C alkyl; and 1105 (vi) b is 0, 1, 2, 3, 4, or 5; and 1098 (vii) n is 2, 3, 4, or 5. 1106 (vii) c is 0, 1, 2, 3, or 4. 1099 PCT Patent Application Publication No. WO 2011/ 1107 PCT Patent Application Publication No. WO 2013/ 153553 by Garske et al., incorporated herein by this refer 006758 by Kahne et al., incorporated herein by this reference, ence, discloses electrophilic Nek9 inhibitors of Formula discloses diphosphate mimetics, including compounds of (N-XXI), (N-XXII), or (N-XXIII): Formula (N-XXIV): US 2016/0243077 A1 Aug. 25, 2016 77

atom, or two R' groups are joined to form an optionally (N-XXIV) Substituted heterocyclic ring; m is 0, 1, 2, 3, or 4: R1-0 1115 (iii) Ring B is a moiety of Subformula (N-XXV(a)) RS NX=x (N-XXV(a)) Y 9 \ RR N R3 Y n WBeN wherein: 1108 (i) X is O or S; 1109 (ii)Y is O or S; 1110 (iii) R' and Rare independently optionally substi 1116 (iv) R' is selected from the group consisting of tuted alkyl, optionally substituted alkenyl, optionally substi hydrogen, halogen, optionally Substituted acyl, optionally tuted alkynyl, optionally Substituted carbocyclyl, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted heterocyclyl, optionally substituted aryl, or substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heteroaryl, or R' and R are taken optionally substituted heterocyclyl, optionally substituted together to form a 5-membered oxazolidin-2-one ring or a aryl, optionally substituted heteroaryl, OR1a, N(R') 6-membered 1.3-oxazinan-2-one ring, wherein the ring is and SR'', wherein each occurrence of R' is indepen optionally substituted or fused to an optionally substituted dently selected from the group consisting of hydrogen, acyl, carbocyclyl, optionally Substituted heterocyclyl, optionally optionally substituted alkyl, optionally substituted alkenyl, Substituted aryl, or optionally Substituted heteroaryl ring; and optionally substituted alkynyl, optionally substituted car 1111 (iv) R is optionally substituted alkyl, optionally bocyclyl, optionally substituted heterocyclyl, optionally sub Substituted alkenyl, optionally Substituted alkynyl, optionally stituted aryl, optionally substituted heteroaryl, a nitrogen pro substituted carbocyclyl, optionally substituted heterocyclyl, tecting group when attached to a nitrogen atom, an oxygen optionally substituted aryl, or optionally substituted het protecting group when attached to an oxygen atom, a Sulfur eroaryl. protecting group when attached to a sulfur atom, or two R' 1112. PCT Patent Application Publication No. WO 2013/ groups are joined to form an optionally Substituted heterocy 074986 by Baker, incorporated herein by this reference, dis clic ring: closes compounds of Formula (N-XXV): 1117 (v) W is N or CR', wherein R’ is selected from the group consisting of hydrogen, halogen, optionally Substi tuted acyl, optionally substituted alkyl, optionally substituted (N-XXV) alkenyl, optionally Substituted alkynyl, optionally Substituted carbocyclyl, optionally Substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl, -OR’", - N(R'), and -SR'', wherein each occurrence of R' is independently selected from the group consisting of hydro I gen, acyl, optionally Substituted alkyl, optionally Substituted alkenyl, optionally Substituted alkynyl, optionally Substituted carbocyclyl, optionally Substituted heterocyclyl, optionally (R'), () Substituted aryl, optionally Substituted heteroaryl, a nitrogen protecting group when attached to a nitrogenatom, an oxygen protecting group when attached to an oxygen atom, a Sulfur protecting group when attached to a sulfur atom, or two R' wherein: groups are joined to form an optionally Substituted heterocy 1113 (i) Ring A is a carbocyclic, heterocyclic, heteroaryl, clic ring: or aryl ring; 1118 (vi) optionally wherein R'' and R' are joined to 1114 (ii) each instance of R is independently selected forman optionally substituted carbocyclyl, optionally Substi from the group consisting of hydrogen, halogen, optionally tuted heterocyclyl, optionally substituted heteroaryl, or substituted acyl, optionally substituted alkyl, optionally sub optionally substituted aryl ring; stituted alkenyl, optionally Substituted alkynyl, optionally 1119 (vii) L is a bond directly attaching Ring A to Ring B; substituted carbocyclyl, optionally substituted heterocyclyl, or L is =C(R) - O - S - NR'' - N(R') optionally substituted aryl, optionally substituted heteroaryl, C(=O) , C(=O)NR'' , SC(=O) , C(=O) —OR'', N(R'), and - SR", wherein each occurrence S OC(=O) C(=O)C NRC(=S) , of R' is independently selected from the group consisting of —C(=S)NR'' , trans-CH=CH-, cis-CH=CH-, hydrogen, acyl, optionally Substituted alkyl, optionally Sub —S(=O).O. , OS(=O), , -S(-O)NR'', stituted alkenyl, optionally Substituted alkynyl, optionally NR''S(=O) , or an optionally substituted C-C, substituted carbocyclyl, optionally substituted heterocyclyl, hydrocarbon chain, optionally wherein one methylene unit of optionally substituted aryl, optionally substituted heteroaryl, the hydrocarbon chain is replaced with=C(R') , —O , a nitrogen protecting group when attached to a nitrogenatom, an oxygen protecting group when attached to an oxygen atom, a Sulfur protecting group when attached to a Sulfur US 2016/0243077 A1 Aug. 25, 2016

CH=CH-, cis-CH=CH-, - S(=O),O OS(=O) optionally substituted aryl, optionally substituted heteroaryl, , S(=O)2NR'', or NRS(=O) ; wherein R.' —ORP', N(R'), and—SRP', wherein each occurrence is hydrogen, halogen, optionally Substituted acyl, optionally of R' is independently selected from the group consisting of substituted alkyl, optionally substituted alkenyl, optionally hydrogen, acyl, optionally Substituted alkyl, optionally Sub substituted alkynyl, optionally substituted carbocyclyl, stituted alkenyl, optionally substituted alkynyl, optionally optionally substituted heterocyclyl, optionally substituted substituted carbocyclyl, optionally substituted heterocyclyl, aryl, optionally Substituted heteroaryl, cyano, or nitro, and optionally substituted aryl, optionally substituted heteroaryl, R'' is hydrogen, C-C alkyl, or a nitrogen protecting group; a nitrogen protecting group when attached to a nitrogenatom, 1120 (viii) - - - - represents a single or double bond; an oxygen protecting group when attached to an oxygen 1121 (ix) X is an optionally substituted C-C hydro atom, a Sulfur protecting group when attached to a Sulfur carbon chain, optionally wherein one or more carbon atom, or two R' groups are joined to form an optionally units of the hydrocarbon chain is replaced with —O—, Substituted heterocyclic ring; —S—, or—NR , where R is hydrogen, C-C alkyl, 1126 (xiv) p is 0, 1, 2, 3, or 4: or a nitrogen protecting group: 1127 (XV) R' is a moiety of Subformula (N-XXV(b)), 1122 (x) L is a bond, —O , —S , NR' , (N-XXV(c)), (N-XXV(d)), (N-XXV(e)), (N-XXV(f)), N(R')C(=O) C(=O)NR' , SC (N-XXV(g)), (N-XXV(h)), (N-XXV(i)), (N-XXV()), (=O)— —C(=O)S —OC(=O)— —C(=O)C)—, (N-XXV(k)), (N-XXV(1)), (N-XXV(m)), (N-XXV(n)), (N-XXV(o)), (N-XXV(p)), (N-XXV(q)), or (N-XXV(r)), depicted below in sequence: C(R2)NR2 , SC(R') , C(R').S. , —S(=O).O. , —OS(=O) , S(=O)NR', —NR'S(=O) , or an optionally substituted C-C, hydrocarbon chain, optionally wherein one methylene unit of the hydrocarbon chain is replaced with —O—,

wherein L' is hydrogen, C-C alkyl, or a nitrogen pro tecting group, and wherein each occurrence of L' is independently selected from the group consisting of hydrogen, halogen, optionally Substituted alkyl, option N ally substituted alkenyl, optionally substituted alkynyl, N optionally substituted carbocyclyl, optionally Substi tuted heterocyclyl, optionally substituted aryl, and L4 L4 Y L3 optionally substituted heteroaryl, or two R' groups are joined to form an optionally substituted carbocyclic or Y N RE3, Y. N Y, s optionally substituted heterocyclic ring; 1123 (xi) each instance of R is independently selected y RE4 from the group consisting of hydrogen, halogen, optionally RE RE2 RE RE2 substituted acyl, optionally substituted alkyl, optionally sub stituted alkenyl, optionally Substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl, —OR', N(R'), and - SR", wherein each occurrence of R is independently selected from the group consisting of hydrogen, acyl, optionally Substituted alkyl, optionally Sub stituted alkenyl, optionally Substituted alkynyl, optionally Y L Y L RR substituted carbocyclyl, optionally substituted heterocyclyl, 3 3 21 NREI optionally substituted aryl, optionally substituted heteroaryl, s s S(O s a nitrogen protecting group when attached to a nitrogenatom, RE2 REI RE2 REI RE31 (O) an oxygen protecting group when attached to an oxygen F C atom, a Sulfur protecting group when attached to a Sulfur O atom, or two R' groups are joined to form an optionally RE2 Substituted heterocyclic ring; RE 1124 (xii) n is 0, 1, 2, 3, or 4: 21 RFI, or 1125 (xiii) each instance of RP is independently selected from the group consisting of hydrogen, halogen, optionally RE3 REI Y RE3 substituted acyl, optionally substituted alkyl, optionally sub O stituted alkenyl, optionally Substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, US 2016/0243077 A1 Aug. 25, 2016 79

enyl, optionally Substituted alkynyl, optionally Substituted carbocyclyl, optionally Substituted heterocyclyl, optionally substituted aryl, and optionally substituted heteroaryl, or two R" groups are joined to form an optionally substituted het (—S) C(=S)NR'' , trans-trans-CR'—CR , erocyclic ring; cis-CR-CR C–C OC(R), C(R) 1133 (xxi) optionally wherein RandR or RandR O. NR?.3°C(R?.3). , C(Rt. 3b)NR's SC(Rt. 3b) or R'' and R'' are joined to form an optionally substituted , —C(R)-S , S(=O).O. , —OS(=O) , carbocyclic or optionally Substituted heterocyclic ring; —S(=O)2NR, NRS(=O), , oran optionally sub 1134) (xxii) R'' is a leaving group; stituted C-C hydrocarbon chain, optionally wherein one 1135 (xxiii)Y is 0, S, or NR'', wherein R is hydrogen, methylene unit of the hydrocarbon chain is replaced with C-C alkyl, or a nitrogen protecting group: - O - S - NR - N(R)C(=O) - C(=O) 1136 (xxiv) a is 1 or 2; and 1137 (XXV) Z is 0, 1, 2, 3, 4, 5, or 6. 1138 PCT Patent Application Publication No. WO 2012/ 158843 by Taunton et al., incorporated herein by this refer –OS(=O), , S(=O)2NR'', or NRS(=O) ; ence, discloses compounds of Formula (N-XXVI) wherein L" is hydrogen, C-C alkyl, or a nitrogen protecting group, and wherein each occurrence of L is independently selected from the group consisting of hydrogen, halogen, (N-XXVI) R3 optionally substituted alkyl, optionally substituted alkenyl, R! L optionally substituted alkynyl, optionally substituted car bocyclyl, optionally substituted heterocyclyl, optionally sub N 71 S-N- stituted aryl, and optionally substituted heteroaryl, or two CN R4 R’ groups are joined to form an optionally substituted car bocyclic or optionally substituted heterocyclic ring; wherein: 1129 (xvii) La is a bondoran optionally substituted C-C, 1139 (i) R' is substituted or unsubstituted alkyl, substi hydrocarbon chain; tuted or unsubstituted heteroalkyl, substituted or unsubsti 1130 (xviii) R'' is selected from the group consisting of tuted cycloalkyl, substituted or unsubstituted heterocy hydrogen, halogen, optionally substituted alkyl, optionally cloalkyl, substituted or unsubstituted aryl, or substituted or Substituted alkenyl, optionally Substituted alkynyl, optionally unsubstituted heteroaryl; substituted carbocyclyl, optionally substituted heterocyclyl, 1140 (ii) Z is a bond, —O— —N(R)— —S , optionally substituted aryl, optionally substituted heteroaryl, —SO— —SO. , Substituted or unsubstituted alkylene, CHOR1, CHN(R)2, CHSR, OR1, substituted or unsubstituted heteroalkylene, substituted or -N(R'), and—SR'', wherein each occurrence of R''“ unsubstituted cycloalkylene, substituted or unsubstituted het is independently selected from the group consisting of hydro erocycloalkylene, substituted or unsubstituted arylene or sub gen, optionally Substituted alkyl, optionally Substituted alk stituted or unsubstituted heteroarylene; enyl, optionally Substituted alkynyl, optionally Substituted 1141 (iii) R' is hydrogen or unsubstituted alkyl: carbocyclyl, optionally Substituted heterocyclyl, optionally 1142 (iv) L' is a bond, C(O) , – N(LR)C(O) , substituted aryl, and optionally substituted heteroaryl, or two OC(O) , S(O), (OLR)OP(O) , N(LR) R" groups are joined to form an optionally substituted het SO-, -(N(LR))NP(O) , substituted or unsubstituted erocyclic ring; alkylene, substituted or unsubstituted heteroalkylene, substi 1131) (xix) R' is selected from the group consisting of tuted or unsubstituted cycloalkylene, substituted or unsubsti hydrogen, halogen, optionally substituted alkyl, optionally tuted heterocycloalkylene, substituted or unsubstituted Substituted alkenyl, optionally Substituted alkynyl, optionally arylene, Substituted or unsubstituted heteroarylene, a moiety substituted carbocyclyl, optionally substituted heterocyclyl, of Subformula (N-XXVI(a)) or Subformula (N-XXVI(b)) optionally substituted aryl, optionally substituted heteroaryl, CHOR2, CHN(R2)2, CHSR2, OR2, -N(R'), and - SR", wherein each occurrence of R’ is independently selected from the group consisting of hydro gen, optionally Substituted alkyl, optionally Substituted alk enyl, optionally Substituted alkynyl, optionally Substituted carbocyclyl, optionally Substituted heterocyclyl, optionally /o/ substituted aryl, and optionally substituted heteroaryl, or two R’" groups are joined to form an optionally substituted het erocyclic ring; 1132 (XX) R' is selected from the group consisting of hydrogen, halogen, optionally substituted alkyl, optionally Substituted alkenyl, optionally Substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, wherein A is substituted or unsubstituted heterocycloalkylene optionally substituted aryl, optionally substituted heteroaryl, or substituted or unsubstituted heteroarylene; CHOR., CHN(R')2, CHSR'', OR*, 1143 (v) n is 0, 1 or 2: -N(R), and - SR", wherein each occurrence of R' (1144 (vi) L is a bond, substituted or unsubstituted alky is independently selected from the group consisting of hydro lene, substituted or unsubstituted heteroalkylene, substituted gen, optionally Substituted alkyl, optionally Substituted alk or unsubstituted cycloalkylene, substituted or unsubstituted US 2016/0243077 A1 Aug. 25, 2016

heterocycloalkylene, substituted or unsubstituted arylene, or form a 3- to 8-membered ring having from 0-2 additional substituted or unsubstituted heteroarylene; heteroatoms as ring members selected from N, O, or S; and Y 1145 (vii) R is hydrogen, substituted or unsubstituted is optionally substituted with one to three moieties each inde alkyl, substituted or unsubstituted heteroalkyl, substituted or pendently selected from R: unsubstituted cycloalkyl, substituted or unsubstituted hetero 1152 (iii) Q is selected from hydrogen, F, Cl, or methyl: cycloalkyl, substituted or unsubstituted aryl, or substituted or 1153 (iv) Z is N(R)(R) or C(R)(R)(R), wherein unsubstituted heteroaryl; R" and R are each independently selected from the group 1146 (viii) R' and R are independently substituted or consisting of hydrogen, optionally Substituted C-C alkyl, unsubstituted alkyl, substituted or unsubstituted heteroalkyl, optionally Substituted C-C cycloalkyl, optionally Substi substituted or unsubstituted cycloalkyl, substituted or unsub tuted heterocycloalkyl, optionally substituted heterocy stituted heterocycloalkyl, substituted or unsubstituted aryl, or cloalkylalkyl, optionally Substituted aryl, optionally Substi substituted or unsubstituted heteroaryl, or R and R are tuted arylalkyl, optionally substituted heteroaryl, or optionally joined together to form a Substituted or unsubsti optionally substituted heteroarylalkyl, or R and R taken tuted cycloalkyl, substituted or unsubstituted heterocy together with the nitrogen atom to which they are attached cloalkyl, substituted or unsubstituted aryl, or substituted or form a 4- to 8-membered ring having from 0 to 2 additional unsubstituted heteroaryl; heteroatoms as ring members selected from N, O, and S. 1147 (ix) R' is independently hydrogen, NR'R'', wherein the 4- to 8-membered ring is optionally substituted; —OR', SR', cyano, substituted or unsubstituted alkyl, 1154) (v) R. RandR are each independently hydrogen, substituted or unsubstituted heteroalkyl, substituted or optionally substituted C-C alkyl, optionally substituted unsubstituted cycloalkyl, substituted or unsubstituted hetero C-C haloalkyl, optionally substituted C-C haloalkoxy, cycloalkyl, substituted or unsubstituted aryl, or substituted or optionally Substituted C-C cycloalkyl, optionally Substi unsubstituted heteroaryl; and tuted C-C cycloalkylalkyl, optionally substituted aryl. 1148 (x) R' and Rare independently hydrogen, sub optionally substituted arylalkyl, optionally substituted het stituted or unsubstituted alkyl, substituted or unsubstituted erocycloalkyl, optionally substituted heterocycloalkylalkyl, heteroalkyl, substituted or unsubstituted cycloalkyl, substi optionally substituted heteroaryl, optionally substituted het tuted or unsubstituted heterocycloalkyl, substituted or unsub eroarylalkyl or XR, wherein X is Ne, 0 or S; stituted aryl, or substituted or unsubstituted heteroaryl. 1155 (vi) R' is hydrogen, C-C alkyl, or aryl; 1149 PCT Patent Application Publication No. WO 2012/ 1156 (vii) R is hydrogen, C-C alkyl, C-C haloalkyl, 109075 by Ibrahim et al., incorporated herein by this refer C-Chaloalkoxy, C-C cycloalkylalkyl, aryl, arylalkyl, het ence, discloses 1 H-pyrrolo-2.3b pyridine compounds of eroaryl, or heteroarylalkyl, where R is optionally substituted Formula (N-XXVII) with 1 to 3 R Substituents; or 1157 (viii) any two of the R. R', and R groups taken together with the carbonatom to which they are attached form (N-XXVII) a 3- to 8-membered optionally Substituted non-aromatic ring having from 0 to 2 heteroatoms selected from N, O, or S; provided that, at each occurrence, at least two of the R. R. and R groups are not simultaneously hydrogen, and with the provisos that: (i) when Y is halogen, methyl, cyano. —OMe, or 2-methoxypyrimidin-5-yl, Z is other than dimethylamino, diethylamino. 1-pyrrolidine, 1-piperidinyl, 4-morpholinyl, isopropyl. —CH(CH)(CHCH), —CH(CH) (CH2CH2CH), cyclobutyl, cyclopentyl, or cyclohexyl; and (ii) when Y is 1-methyl-4-pyrazolyl, 3-methylsulfonylphe wherein: nyl or 3-methylsulfonylaminophenyl, Z is other than cyclo 1150 (i)Y' is selected from the group consisting of cyano, propyl, cyclobutyl, cyclopentyl or cyclohexyl. halogen, hydroxyl, optionally Substituted C-C alkyl, 1158 PCT Patent Application Publication No. WO 2013/ optionally substituted C-C alkoxy, optionally substituted 147711 by Nacro et al., incorporated herein by this reference, C-C cycloalkyl, optionally Substituted C-C haloalkyl, discloses bicyclic heterocyclic derivatives, including com optionally Substituted C-C haloalkoxy, optionally Substi pounds of Formula (N-XXVIII): tuted aryl and optionally substituted heteroaryl: optionally wherein the two adjacent substituents on the aryl or heteroaryl ring together with the atoms to which they are attached form X (N-XXVIII) an optionally substituted 5- or 6-membered ring having from R3 xé "Sx2-y6 N 0 to 3 additional heteroatoms selected from N, O or S; 1151 (ii)Y is hydrogen, halogen, cyano, hydroxyl, C-C, R2 N s 3sNx, alkyl, C-C alkoxy, C-Chaloalkyl, C-Chaloalkoxy, aryl, R5 arylalkyl, heteroaryl, heteroarylalkyl, C-C cycloalkyl-Co 2 X7 R9 Calkyl, or (R)(R)N , wherein R and Rare each inde R1 Xs pendently selected from the group consisting of hydrogen, R6 C-C alkyl, C-C alkoxy, aryl, arylalkyl, heteroaryl, het eroarylalkyl, C-C cycloalkyl-Co-Calkyl, heterocycloalkyl R8 and heterocycloalkyl-C-C alkyl: or R and R taken together with the nitrogen atom to which they are attached US 2016/0243077 A1 Aug. 25, 2016

wherein: These agents include, but are not limited to, short interfering 1159 (i) X, X, and X are independently nitrogen or RNA (sRNA), microRNA (miRNA), synthetic hairpin RNA carbon; (shRNA), antisense nucleic acids, and complementary DNA 1160 (ii) X, Xs, and X are independently nitrogen or (cDNA). Other agents that can inhibit expression by Sup CR'; wherein at least one of X1, X2, Xs, X, Xs, and X is pressing transcription of the NEK9 gene are known in the art nitrogen; and are disclosed in PCT Application Publication No. 2011/ 070150 by Ullrich et al., including precursors of sRNA or 1161 (iii) X, is nitrogen or CR': miRNA, oligonucleotide aptamers, ribozymes, polypeptides 1162 (iv) - - - - is a single or double bond, as required by binding to target structures, including antibodies or peptidic Valency; aptamers, and low-molecular-weight organic molecules. The 1163 (v) R', R. R. R. R. R7, R, R, R 0, and R'' are use of siRNAs for inhibiting the expression of the NEK9 gene independently hydrogen, halogen, optionally Substituted is also disclosed in PCT Patent Application Publication No. alkyl, optionally substituted aryl, optionally Substituted car 2006/128063 by Gao et al., incorporated herein by this refer bocyclyl, optionally substituted heteroaryl, optionally substi ence. RNA interference is the process whereby the introduc tuted heterocyclyl, cyano. —SR, N(R), nitro, —C(O)R, tion of double stranded RNA into a cell inhibits the expression —COR, —C(O)N(R), —S(O)R. —SOR. - SON(R), of a gene corresponding to its own sequence. RNAi is usually OC(O)R, NRC(O)R, NRSOR, or NRC(O)N(R): described as a post-transcriptional gene-silencing (PTGS) 1164 (vi) each R is independently hydrogen, optionally mechanism in which double Stranded RNA triggers degrada Substituted C-C alkyl, optionally Substituted aryl, option tion of homologous messenger RNA in the cytoplasm. The ally substituted carbocyclyl, optionally substituted het siRNAs bind to a ribonuclease complex called RNA-induced eroaryl, or optionally substituted heterocyclyl, or two R silencing complex (RISC) that guides the small dsRNAs to its groups on the same nitrogen atom may be taken together to homologous mRNA target. Consequently, RISC cuts the form an optionally substituted heterocyclyl moiety; mRNA approximately in the middle of the region paired with 1165 (vii) each R" is independently hydrogen, halogen, or the antisense sRNA, after which the mRNA is further optionally substituted C-C alkyl; and degraded. 1166 (viii) R' and R, R and R, R and R, R and R'', 1170. When antibodies are used, such antibodies can be R'' and R'', RandR, R and R7, R7 and R, or RandR polyclonal or monoclonal. As used herein in this context, the may be optionally taken together to form an optionally Sub term "monoclonal antibodies' includes, but is not limited to, stituted 5-membered or 6-membered carbocyclyl, aryl, het chimeric antibodies, humanized antibodies, antibody frag erocyclyl, or heteroaryl ring. ments such as scFv fragments, diabodies, heavy chain anti 1167 For these Nek9 inhibitors, the compounds described bodies (HCAbs), and single-domain antibodies (sdAbs). Such above also include pharmaceutically acceptable analogues, monoclonal antibodies are not necessarily produced as the congeners, bioisosteres, hydrolysis products, metabolites, result of cell fusion between B cells and myeloma cells, and precursors, and prodrugs thereof. For compounds having one can be produced in other eukaryotic cells or even bacterial or more chiral centers, the compounds include all possible cells according to methods known in the art. General tech Stereoisomers including enantiomers and/or diastereomers niques of antibody engineering are described in C. A. K. unless it can be shown that one or more specific stereoisomers Borrebaeck, Antibody Engineering” (2d ed., Oxford Univer lack pharmacological activity. sity Press, New York, 1995), incorporated herein by this ref 1168 Additionally, the kinases Mad2, BubR1, and Mps 1 CCC. interact with Nek9 and inhibitors of one or more of those 1171 Screening methods for Nek9 inhibitors are kinases can promote inhibition of the activity of Nek9 in this described in United States Patent Application Publication No. complex regulatory network. Inhibitors of Mad2 are dis 2011/0229484 by Baumert et al., incorporated herein by this closed in the following United States patents, all of which are reference. A variety of assay protocols and detection tech incorporated herein by this reference: U.S. Pat. No. 8,629,118 niques are well known in the art and can easily be adapted for to Pellman; U.S. Pat. No. 8,586.297 to Pagano et al.; U.S. Pat. this purpose by one skilled in the art. Such methods include, No. 8,470,798 to Tagaki et al.; U.S. Pat. No. 8,399,659 to but are not limited to, high-throughput assays (e.g., microar Claiborne et al.; U.S. Pat. No. 8,354,386 to Lee et al.; U.S. ray technology, phage display technology) and in vitro and in Pat. No. 7,718,648 to Claiborne et al.; and U.S. Pat. No. Vivo cellular and tissue assays. One alternative comprises 7,572,784 to Claiborne et al. Inhibitors of BubR1 are dis incubating a biological system, which expresses (or can closed in the following United States patents, all of which are express) Nek9, with a candidate compound under conditions incorporated herein by this reference: U.S. Pat. No. 8,026,355 and for a time Sufficient for the candidate compound to modu to Hansen et al.; U.S. Pat. No. 7,943,629 to Luecking et al.: late the kinase activity; and measuring the activity of the U.S. Pat. No. 7,897,568 to Jia et al.; and U.S. Pat. No. 7,781, kinase. Candidate compounds that decrease the activity of the 580 to Lee et al. Inhibitors of Mps 1 are disclosed in the kinase are identified as kinase inhibitors. In certain embodi following United States patents, all of which are incorporated ments, a method according to the invention more specifically herein by this reference: U.S. Pat. No. 8,551,980 to Schulzeet includes incubating a biological system, which expresses (or al.; U.S. Pat. No. 8,541,576 to Casucelli et al.; U.S. Pat. No. can express) Nek9 with a candidate compound under condi 8,513,241 to Cerviet al.; U.S. Pat. No. 8,394,802 to Mirizziet tions and for a time Sufficient for the candidate compound to al.; U.S. Pat. No. 8,207,165 to Cervi et al.; U.S. Pat. No. modulate the activity of the kinase, thereby obtaining a test 8,106,069 to Salom et al.; U.S. Pat. No. 6,593,098 to Yen et system; incubating the biological system under the same con al.; and U.S. Pat. No. 5,972,640 to Drubin et al. ditions and for the same time absent the candidate compound, 1169. In another alternative, the derivative or analog of thereby obtaining a control system; measuring in the test meisoindigo, including one of Alternatives (1)–(486) above, system, at least one factor that is representative of the activity can be used together with an agent that inhibits Nek9 expres of the Nek9 kinase; measuring that factor in the control sys Sion, Such as by Suppressing transcription of the NEK9 gene. tem; comparing the factor measured in the test system and the US 2016/0243077 A1 Aug. 25, 2016

control system; and determining that the candidate compound al., incorporated herein by this reference, which discloses inhibits the activity of the kinase if the factor measured in the cyclopentyl (2S)-cyclohexyl ({6-3-(hydroxyamino)-3-oxo test system is less than or greater than the factor measured in propylpyridin-3-yl)methyl)aminoacetate; U.S. Pat. No. the control system. Other methods are known in the art. 8,614.223 to van Duzer et al., incorporated herein by this 1172 Accordingly, the following compositions and meth reference, which discloses pyrimidine hydroxy amide com ods are within the scope of the present invention: (1) compo pounds; U.S. Pat. No. 8,609.678 to van Duzer et al., incorpo sitions and methods employing a derivative or analog of rated herein by this reference, which discloses reverse amide meisoindigo, including one of Alternatives (1)–(486) above, compounds including a Zinc chelator group; U.S. Pat. No. together with an Nek9 inhibitor; (2) compositions and meth 8,603.521 to Loury et al., incorporated herein by this refer ods employing a derivative or analog of meisoindigo, includ ence, which discloses N-hydroxy-4-2-3-(N,N-dimethy ing one of Alternatives (1)–(486) above, together with an laminomethyl)benzofuran-2-ylcarbonylaminolethoxy-ben anti-Nek9 antibody; (3) compositions and methods employ Zamide: U.S. Pat. No. 8,598,168 to Moradei et al., ing a derivative or analog of meisoindigo, including one of incorporated herein by this reference, which discloses ortho Alternatives (1)–(486) above, together with an agent that aminobenzamides; U.S. Pat. No. 8,592,444 by Varasi et al., inhibits Nek9 expression; and (4) compositions and methods incorporated herein by this reference, which discloses spiro employing a derivative or analog of meisoindigo, including cyclic derivatives; U.S. Pat. No. 8,592,441 to Verdoncket al., one of Alternatives (1)–(486) above, together with an inhibitor incorporated herein by this reference, which discloses substi of at least one of Mad2, BubR1, and Mps1. tuted indolylalkylamino derivatives; U.S. Pat. No. 8,557,825 1173 Another class of agents that can be used together to Van Emelen et al., incorporated herein by this reference, with derivatives or analogs of meiso indigo and Nek9 inhibi which discloses sulfonyl derivatives; U.S. Pat. No. 8,546,588 tors or other agents inhibiting the expression of Nek9 are by Blackburn et al., incorporated herein by this reference, histone deactylase inhibitors (“HDAC inhibitors'). Suitable which discloses substituted hydroxamic acids; U.S. Pat. No. HDAC inhibitors are disclosed in PCT Patent Application 8,524,711 to Angibaud et al., incorporated herein by this Publication No. WO 2013/040286 by Chen et al., incorpo reference, which discloses Substituted amino compounds; rated herein by this reference, and include compounds of U.S. Pat. No. 8,513,237 to Van Emelen et al., incorporated Formula (H-I): herein by this reference, which discloses sulfonylamino derivatives; U.S. Pat. No. 8,501,737 to Van Emelen, incorpo rated herein by this reference, which discloses piperazinyl, (H-I) piperidinyl, and morpholinyl derivatives; U.S. Pat. No. 8,492, 401 to Oalmann et al., incorporated herein by this reference, which discloses thiazolopyridine derivatives; U.S. Pat. No. 8,491.927 to Epner et al., incorporated herein by this refer ence, which discloses entinostat, panobinostat, Vorinostat, and romedepsin; and U.S. Pat. No. 8,476.255 by Rajgopal et al., incorporated herein by this reference, which discloses stilbene-like compounds. Other histone deacetylase inhibi tors are known in the art. 1179 Accordingly, the following compositions and meth ods are within the scope of the present invention: (1) compo wherein: sitions and methods employing a derivative or analog of 1174 (i) m is 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, or 16; meisoindigo, including one of Alternatives (1)–(486) above, 1175 (ii) Z is absent, C(RR), O, S, C(O), N(R), SO, together with an HDAC inhibitor; (2) compositions and meth OC(O), C(O)O, OSO, S(O)O, C(O)S, SC(O), C(O)C(O), ods employing a derivative or analog of meisoindigo, includ C(O)N(R), N(R)C(O), S(O)N(R), N(R)S(O), OC(O)N ing one of Alternatives (1)–(486) above, together with an (R), N(R)C(O)O, N(R)C(O)S, or N(R)C(O)N(R), in HDAC inhibitor and an Nek9 inhibitor; (3) compositions and which each of IR, and R, independently, is H. alkyl, alkenyl, methods employing a derivative or analog of meisoindigo, or alkynyl, including one of Alternatives (1)–(486) above, together with 1176 (iii) X and X independently, is halo or OSOR, in an HDAC inhibitor and an anti-Nek9 antibody; (4) composi which R is alkyl, alkenyl, or alkynyl; and tions and methods employing a derivative or analog of 1177 (iv) Q is cycloalkyl, heterocycloalkyl, cycloalkenyl, meisoindigo, including one of Alternatives (1)–(486) above, heterocycloalkenyl, aryl, or heteroaryl, each of which, inde together with an HDAC inhibitor and an agent that inhibits pendently, is optionally Substituted with alkyl, alkenyl, alky Nek9 expression; and (4) compositions and methods employ nyl, cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocy ing a derivative or analog of meisoindigo, including one of cloalkenyl, aryl, heteroaryl, halo, nitro, oxo. —C=NH. Alternatives (1)–(486) above, together with an HDAC inhibi cyano, alkyl-Rd, OR OC(O)R OC(O)OR OC(O)SR. tor and an inhibitor of at least one of Mad2, BubR1, and SR C(O)R C(O)OR, C(O)SR, C(O)NR.R. SOR, Mps1. SO.R.NR,R or N(R)C(O)R in which each of R. R., and 1180 Another aspect of the present invention is a compo Ra independently, is H, alkyl, alkenyl, alkynyl, cycloalkyl, sition to improve the efficacy and/or reduce the side effects of heterocycloalkyl, aryl, heteroaryl, halo, cyano, amine, nitro, Suboptimally administered drug therapy comprising an alter hydroxy, or alkoxy. native selected from the group consisting of 1178. Other HDAC inhibitors are known in the art, and are 1181 (i) a therapeutically effective quantity of a modified disclosed in U.S. Pat. No. 8,648,092 to Lee et al., incorpo therapeutic agent or a derivative, analog, or prodrug of a rated herein by this reference, which discloses imidazo[1,2- therapeutic agent or modified therapeutic agent, wherein the alpyridine derivatives; U.S. Pat. No. 8,637,547 to Davidsonet modified therapeutic agent or the derivative, analog or pro US 2016/0243077 A1 Aug. 25, 2016 drug of the therapeutic agent or modified therapeutic agent 1189. In one alternative, the composition comprises a drug possesses increased therapeutic efficacy or reduced side combination comprising: effects as compared with an unmodified therapeutic agent; 1190 (i) indirubin, an analog of indirubin, or a derivative of indirubin or of an analog of indirubin; and 1182 (ii) a composition comprising: 1191 (ii) an additional therapeutic agent selected from the 1183 (a) a therapeutically effective quantity of a thera group consisting of: peutic agent, a modified therapeutic agent or a deriva 1192 (a) topoisomerase inhibitors; tive, analog, or prodrug of a therapeutic agent or modi 1193 (b) fraudulent nucleosides; 1194 (c) fraudulent nucleotides: fied therapeutic agent; and 1195 (d) thymidylate synthetase inhibitors; 1184 (b) at least one additional therapeutic agent, 1196 (e) signal transduction inhibitors; therapeutic agent Subject to chemosensitization, thera 1197 (f) cisplatin or platinum analogs; peutic agent Subject to chemopotentiation, diluent, 1198 (g) alkylating agents; excipient, solvent system, or drug delivery system, 1199 (h) anti-tubulin agents; wherein the composition possesses increased therapeu 1200 (i) antimetabolites: tic efficacy or reduced side effects as compared with an 1201 () berberine; unmodified therapeutic agent; 1202 (k) apigenin: 1203 (1) amonafide: 1185 (iii) a therapeutically effective quantity of a thera 1204 (m) colchicine or an analog thereof; peutic agent, a modified therapeutic agent, or a derivative, 1205 (n) genistein; analog, or prodrug of a therapeutic agent or modified thera 1206 (o) etoposide; peutic agent that is incorporated into a dosage form, wherein 1207 (p) cytarabine; the therapeutic agent, the modified therapeutic agent, or the 1208 (q) a camptothecin; derivative, analog, or prodrug of a therapeutic agent or modi 1209 (r) a vinca alkaloid; fied therapeutic agent incorporated into the dosage form pos 1210 (s) 5-fluorouracil; sesses increased therapeutic efficacy or reduced side effects 1211 (t) curcumin; as compared with an unmodified therapeutic agent; 1212 (u) an NF-kB inhibitor; 1213 (V) rosmarinic acid; 1186 (iv) a therapeutically effective quantity of a thera 1214 (w) mitoguaZone; peutic agent, a modified therapeutic agent, or a derivative, 1215 (x) tetandrine; analog, or prodrug of a therapeutic agent or modified thera 1216 (y) an antineoplastic agent not metabolized by peutic agent that is incorporated into a dosage kit and pack cytochrome P450 CYP1A2 or CYP2C19; aging, wherein the therapeutic agent, the modified therapeu 1217 (Z) a biological therapy: tic agent, or the derivative, analog, or prodrug. ofatherapeutic 1218 (aa) a tyrosine kinase inhibitor; agent or modified therapeutic agent incorporated into the 1219 (ab) all-trans-retinoic acid; 1220 (ac) an arsenical; dosage kit and packaging possesses increased therapeutic 1221 (ad) hydroxyurea; efficacy or reduced side effects as compared with an unmodi 1222 (ae) thioguanine; fied therapeutic agent; and 1223 (af) mercaptopurine; 1187 (v) a therapeutically effective quantity of a thera 1224 (ag) homoharringtonine; peutic agent, a modified therapeutic agent, or a derivative, 1225 (ah) oridonin; analog, or prodrug of a therapeutic agent or modified thera 1226 (a) uracil mustard; peutic agent that is Subjected to a bulk drug product improve 1227 (ak) nilotinib: ment, wherein the therapeutic agent, the modified therapeutic 1228 (al) dasatinib; agent, or the derivative, analog, or prodrug of a therapeutic 1229 (am) lonidamine: agent or modified therapeutic agent Subject to the bulk drug 1230 (an) 5-azacytidine: product improvement possesses increased therapeutic effi 1231 (ao) thalidomide or an analog thereof; cacy or reduced side effects as compared with an unmodified 1232 (ap) an EGFR inhibitor such as erlotinib, afatinib, therapeutic agent. lapatinib, or dacomitinib; 1233 (aq) a gold salt Such as aurothiomalate or 1188 Typically, the composition possesses increased effi aurothioglucose: cacy or reduced side effects for cancer therapy. Typically, the 1234 (ar) dibromodulcitol; unmodified therapeutic agent is indirubin, an analog of 1235 (as) dianhydrogalactitol; indirubin, or a derivative of indirubin or of an analog of 1236 (at) decitabine; and indirubin, as described above. In another alternative, the 1237 (au) a proteasome inhibitor. therapeutically active agent is an agent selected from the 1238. In this alternative, when the additional therapeutic group consisting of Alternatives (1)–(486), the modified agent is an alkylating agent, the alkylating agent can be, but is therapeutic agent is a modification of an agent selected from not limited to, an alkylating agent selected from the group the group consisting of Alternatives (1)–(486), and the deriva consisting of BCNU, BCNU wafers, CCNU, bendamustine tive, analog, or prodrug is a derivative, analog, or prodrug of (Treanda), and temozolimide (Temodar). an agent selected from the group consisting of Alternatives 1239. In another alternative, the composition comprises: (1)–(486) or of a modification of an agent selected from the 1240 (i) indirubin, an analog of indirubin, or a derivative group consisting of Alternatives (1)–(486). Alternatives (1)- of indirubin or of an analog of indirubin; and (486) are described above. Typically, the unmodified thera 1241 (ii) a therapeutic agent Subject to chemosensitiza peutic agent is meisoindigo, and, where the composition tion selected from the group consisting of: includes a modified therapeutic agent, the modified therapeu 1242 (a) topoisomerase inhibitors; tic agent is typically a modification of meisoindigo. 1243 (b) fraudulent nucleosides; US 2016/0243077 A1 Aug. 25, 2016 84

1244 (c) fraudulent nucleotides; 1292 (e) preparation with lower residual solvent con 1245 (d) thymidylate synthetase inhibitors; tent; and 1246 (e) signal transduction inhibitors; 1293 (f) preparation with lower residual heavy metal 1247 (f) cisplatin or platinum analogs; COntent. 1248 (g) alkylating agents; 1294. In still another alternative, the therapeutic agent is 1249 (h) anti-tubulin agents: indirubin, an analog of indirubin, or a derivative of indirubin 1250 (i) antimetabolites: or of an analog of indirubin and the composition comprises a 1251 (i) berberine; diluent, wherein the diluent is selected from the group con 1252) (k) apigenin: sisting of: 1253 (1) amonafide: 1295 (a) an emulsion; 1254 (m) vinca alkaloids; 1296 (b) dimethylsulfoxide (DMSO); 1255 (n) 5-fluorouracil; 1297 (c)N-methylformamide (NMF) 1256 (o) curcumin; 1298 (d) DMF; 1257 (p) NF-kB inhibitors: 1299 (e) ethanol: 1258 (q) rosmarinic acid; 1300 (f) benzyl alcohol: 1259 (r) mitoguaZone: 1301 (g) dextrose-containing water for injection; 1260 (s) tetrandrine; and 1302 (h) Cremophor; 1261 (t) a proteasome inhibitor. 1303 (i) cyclodextrin; and 1262. In still another alternative, the composition com 1304 (i) PEG. prises: 1305. In still another alternative, the therapeutic agent is 1263 (i) indirubin, an analog of indirubin, or a derivative indirubin, an analog of indirubin, or a derivative of indirubin of indirubin or of an analog of indirubin; and or of an analog of indirubin and the composition comprises a 1264 (ii)atherapeutic agent Subject to chemopotentiation solvent system, wherein the solvent system is selected from selected from the group consisting of: the group consisting of 1265 (a) topoisomerase inhibitors: 1306 (a) an emulsion; 1266 (b) fraudulent nucleosides; 1307 (b) dimethylsulfoxide (DMSO); 1267 (c) fraudulent nucleotides; 1308 (c)N-methylformamide (NMF) 1268 (d) thymidylate synthetase inhibitors; 1269 (e) signal transduction inhibitors; 1309 (d) DMF; 1270 (f) cisplatin or platinum analogs; 1310 (e) ethanol: 1271 (g) alkylating agents; 1311 (f) benzyl alcohol: 1272 (h) anti-tubulin agents: 1312 (g) dextrose-containing water for injection; 1273 (i) antimetabolites: 1313 (h) Cremophor; 1274 () berberine; 1314 (i) cyclodextrin; and 1275 (k) apigenin: 1315 (i) PEG. 1276 (1) amonafide; 1316. In yet another alternative, the therapeutic agent is 1277 (m) vinca alkaloids; indirubin, an analog of indirubin, or a derivative of indirubin 1278 (n) 5-fluorouracil; orofananalog of indirubin and the composition comprises an 1279 (o) curcumin; excipient, wherein the excipient is selected from the group 1280 (p) NF-kB inhibitors: consisting of: 1281 (q) rosmarinic acid; 1317 (a) mannitol; 1282 (r) mitoguaZone: 1318 (b) albumin; 1283 (s) tetrandrine; 1319 (c) EDTA; 1284 (t) biotherapeutics; and 1320 (d) sodium bisulfite: 1285 (u) a proteasome inhibitor. 1321 (e) benzyl alcohol: wherein the indirubin, an analog of indirubin, or a derivative 1322 (f) a carbonate buffer; of indirubin or of an analog of indirubin acts as a chemopo 1323 (g) a phosphate buffer; and tentiator. 1324 (h) methylcellulose. 1286. In this alternative, wherein the additional therapeu 1325 In still another alternative, the therapeutic agent is tic agent is a biotherapeutic, the biotherapeutic can be, but is indirubin, an analog of indirubin, or a derivative of indirubin not limited to, a biotherapeutic selected from the group con or of an analog of indirubin and the indirubin, an analog of sisting of Avastin, Herceptin, Rituxan, and ErbituX. indirubin, or a derivative of indirubin or of an analog of 1287. In yet another alternative, the therapeutic agent is indirubin is incorporated into a dosage form selected from the indirubin, an analog of indirubin, or a derivative of indirubin group consisting of: or of an analog of indirubin and the indirubin, an analog of 1326 (a) tablets; indirubin, or a derivative of indirubin or of an analog of 1327 (b) capsules; indirubin is subjected to a bulk drug product improvement, 1328 (c) topical gels: wherein the bulk drug product improvement is selected from 1329 (d) topical creams: the group consisting of 1330 (e) patches: 1288 (a) salt formation; 1331 (f) suppositories; and 1289 (b) preparation as a homogeneous crystal struc 1332 (g) lyophilized dosage fills. ture; 1333. In yet another alternative, the therapeutic agent is 1290 (c) preparation as a pure isomer; indirubin, an analog of indirubin, or a derivative of indirubin 1291 (d) increased purity: or of an analog of indirubin and the indirubin, an analog of