US007575876B2
(12) United States Patent (10) Patent No.: US 7,575,876 B2 Zhang (45) Date of Patent: Aug. 18, 2009
(54) BIOMARKERS FOR NEURODEGENERATIVE Smallet al., “In Vivo BrainImaging of Tangle Burden in Humans', J DSORDERS Mol Neurosci, 2002. 19(3): p. 323. Jankovic et al., “The Evolution of Diagnosis in Early Parkinson (75) Inventor: Jing Zhang, Mercer Island, WA (US) Disease”, Arch Neurol, 2000. 57(3): p. 369. Hughes et al., “The accuracy of Diagnosis of Parkinsonian Syn (73) Assignee: The University of Washington, Seattle, dromes in a Specialist Movement Disorder Service'. Brain, 2002. 125(Pt 4): p. 861. WA (US) Litvan et al., "Accuracy of the Clinical Diagnoses of Lewy Body Disease, Parkinson Disease, and Dementia with Lewy Bodies'. Arch *) NotOt1Ce: Subjubject to anyy d1Sclaimer,disclai theh term off thithis Neurol, 1998.55(7): p. 969. patent is extended or adjusted under 35 Rajput et al., "Accuracy of Clinical Diagnosis in Parkinsonism—A U.S.C. 154(b) by 0 days. Prospective Study”. Can J Neurol Sci, 1991. 18(3): p. 275. Hughes et al., "Accuracy of Clinical Diagnosis of Idiopathic (21) Appl. No.: 11/441,384 Parkinson's Disease: a clinico-Pathological Study of 100 Cases”. J Neurol Neurosurg Psychiatry, 1992. 55(3): p. 181-184. (22) Filed: May 24, 2006 McKeith et al., “Dementia with Lewy Bodies'. Semin Clin Neuropsychiatry, 2003. 8(1): p. 46. (65) Prior Publication Data Love et al., “PostMortem Sampling of the Brain and other Tissues in US 2007/OO992O3 A1 May 3, 2007 Neurodegenerative Disease'. Histopathology, 2004. 44(4): p. 309. Olsson et al., “Simultaneous Measurement off-Amyloid-2. Total Tau, and Phosphorylated Tau (Thr') in Cerebrospinal Fluid by the Related U.S. Application Data XMAP Technology”, Clin Chem, 2005. 51(2): p. 336. (60) Provisional application No. 60/731,339, filed on Oct. Zhang et al., “Quantitative Proteomic Analysis of Age-Related 27, 2005. Changes in Human Cerebrospinal Fuid'. Neurobiol Aging, 2005. 26(2): p. 207. (51) Int. Cl. Zhang et al., “Quantitative Proteomic of Cerebrospinal Fluid from A6 IB5/00 (2006.01) Patients with Alzheirmer Disease”, J Alzheimers Dis, 2005. 7(2): p. GOIN 33/00 (2006.01) 125. GOIN 33/53 (2006.01) * cited by examiner GOIN 33/566 (2006.01) GOIN 33/48 (2006.01) Primary Examiner Olga NChernyshev GOIN 3L/00 (2006.01) Assistant Examiner Stacey MacFarlane A6 IK 38/00 (2006.01) (74) Attorney, Agent, or Firm Edward J. Baba; Bozicevic, C07K 6/00 (2006.01) Field & Francis LLP (52) U.S. Cl...... 435/7.1:436/86; 436/501; 435/7.9; 530/300: 530/387.9; 600/300; 702/19; (57) ABSTRACT 702/23 (58) Field of Classification Search ...... None The present invention provides methods for diagnosing neu See application file for complete search history. rodegenerative disease. Such as Alzheimer's Disease, Parkin son's Disease, and dementia with Lewy body disease by (56) References Cited detectingapattern of gene product expression in a cerebrospi nal fluid sample and comparing the pattern of gene product OTHER PUBLICATIONS expression from the sample to a library of gene product Blennow, Journal of Internal Medicine 256:224-234, Sep. 2004.* expression pattern known to be indicative of the presence or Earley et al., Cytometry 50(5): 239-42, Oct. 15, 2002.* absence of a neurodegenerative disease. The methods also Otto et al., Methods, 44; 289-298 (2008).* provide for monitoring neurodegenerative disease progres Hickey et al., Current Alzheimer Research, 5:52-60, 2008.* sion and assessing the effects of therapeutic treatment. Also Bacskai et al., “Imaging Amyloid-f Deposits InVivo”. JCereb Blood provided are kits, systems and devices for practicing the Flow Metab, 2002. 22(9): p. 1035-41. Klunk et al., “Imaging A Plaques in Living Transgenic Mice with Subject methods. Multiphoton and Methoxy-X04, a Systemically Administered Congo Red Derivative”. JNeuropathol ExpNeurol, 2002.61 (9): p. 797-805. 6 Claims, 97 Drawing Sheets U.S. Patent Aug. 18, 2009 Sheet 1 Of 97 US 7,575,876 B2
Cell cycle/death
Cell structure?motility/transport/traffic
O Extracellular matrix/Cell Adhesion
Developmental process
immunity/defense
Metabolism S. Neuronal activities/signal transduction
Unknown functions
FIG. 1 U.S. Patent Aug. 18, 2009 Sheet 2 of 97 US 7,575,876 B2
A: Pooled samples
AD PD OLB CT AO PD DLB CT
Protein armout CSF volume B: individual Samples
FIG. 2 U.S. Patent Aug. 18, 2009 Sheet 3 Of 97 US 7,575,876 B2
A B
C
3. ri t -
A. s oi. 3 9. s: } c. o
c c " " - Beta-Fibrinogen
l -T- d TH -3 -2 2 3 O.O 0.2 0.4 0.6 0.8 1.O Standardized Vito BP 1-Specificity
FIG. 3 U.S. Patent Aug. 18, 2009 Sheet 4 of 97 US 7,575,876 B2
A B a is Pd w AD a DLB 4 Control
- O 2 4 Standardized ApoH 0.0 0.2 0.4 0.6 0.8 10 1-Specificity
FIG. 4 U.S. Patent Aug. 18, 2009 Sheet 5 of 97 US 7,575,876 B2
Proteins With Changes. In Expression Level
Protein Name Peptide Sequences
AD PD | DLB peptides
Brain abundant, membrane attached AEPPKAPEQEQAAPGPAAGGEAPK 0.39 signal protein l (SEQID NO:1) EADVVAR (SEQ ID NO:2) EKPDQDAEGK (SEQID NO:3) EsEPQAAAEPAEAK (SEQID No.4) Brain-derived neurotrophic factor IPI00336003 AAPMK (SEQID NO:5) BDNF1 GQGGLAYPGVR (SEQ ID NO:6) IDTSCVCTLTIK (SEQID NO:7) NYLDAANMSMRVR (SEQ ID NO:8) Cell growth regulator with EF hand domain|IPI00337548 ELPGETLESK (SEQID NO:9) 1 0.77 | | | 1.67 line ESLDPVOEPGGQAEADGDVPGPR (SEQID NO:10) GEAEGQAEAK (SEQID NO:11)
HVEPGEPLAPSPQEPQAVGR (SEQ ID NO:14)
QETQEAPGPR (SEQID NO:16) RESLDPVQEPGGQAEADGDVPGPR (SEQID NO:17)
Chromogranin. A io9 Ncos NC EAVEEPSSK (SEQ ID NO:20) EEEEEMAVVPQGLFR (SEQ ID ELQDLALQGAK (SEQID No.22) EWEDSK (SEQID NO:23) | | | | | | GEQEHSQQKEEEEEMAVVPQGLFR | | | | |
F.G. 5A
U.S. Patent Aug. 18, 2009 Sheet 10 of 97 US 7,575,876 B2
Ad Pd DLB
LIDDYDSTK(SEQID No.147) || | | | | |
NIEWLK (SEQID No. 148) | | | |
(SEQID NO:149) Splice isoform 2 of insulin-like growth IPI00215977 ELEAFR (SEQID NO:150) 1.57 | f | 1.07 NC factor II precursor FFQYDTwk (SEQID No.151) GFYFR (SEQ ID NO:152)
GHVLAK (SEQ ID NO: 153) GIVEECCFR (SEQID NO:154) GLPALLR (SEQID NO:155)
SCDLALLETYCATPAK (SEQID
NO:156) YPVGK (SEQ ID NO:157) Splice isoform 3 of calcium/calmodulin dependent protein IPIO021916.5 FYFENLLAK (SEQID NO:158) 0.64 0.85 NC kinase type II beta chain
GAILTTMLATR (SEQID NO:159) member 10E""'IPI00011167 FDELDMSPGDPK (SEQID No.160) 1.25 | f | 1.03 sale GGHGQQDLFRVLK (SEQID NO:161)
GIPPSLR (SEQID NO:162)
LRSLSPK (SEQ ID NO:163) YLPGYYSEK (SEQ ID NO:164) Cadherin-13 IPIO0024046 DIQGSLQDIFK (SEQ ID NO:165) O 5 5 1 .1 1 NC O 8 4. OcCuSO u
GIFRINENTGSVSVTR (SEQID NO:166)
F TLFVHAR (SEQID NO:167) TPHAEDMAELVIVGGK (SEQID NO:168) E VDCNAAGALR (SEQ ID NO:169) VNSDGGLVALR(SEQID NO:170) YEVSSPYFK (SEQID NO:171) subunitOS precursor PI00026154 ILIEDwk (SEQID No. 172) 0. 64 l l 1 O 7 TvKEEAEKPER (SEQID No.173) Golgi autoantigen, IPI00004671. AEMEEK (SEQID NO:174) O. 4. 7 2 . 4. tC l 7 NNNCCC FIG. 5F U.S. Patent Aug. 18, 2009 Sheet 11 of 97 US 7,575,876 B2
iTRAQ Ratios Protein Name IPI # Peptide Sequences AD PD DLB golgin subfamily B member 1 AQEIYEK (SEQID NO:175) DLVEMEQK (SEQID NO:176) | | | DVQLQQK (SEQID No.177) | | | EALKENKSLQEELSLAR (SEQ ID NO:178 EEDVSYLSGQLSEKEAALTK (SEQID NO: 179 EIKELENLLSQEEEENIVLEEENK H (SEQID NO: 180 ELLOR (SEQ ID NO:181) ELLSQLEETR (SEQ ID NO:182) EMKQMEGEGIAPIKMK (SEQ ID NO: 183) ENENIGDQLR (SEQ ID NO:184) ENLAQAVEHR (SEQ ID NO:185) LDELQK (SEQ ID NO:186) LLMVTK (SEQID NO:187) NETETAEER (SEQ ID NO:188) QDGDK (SEQID NO:189) SMSSLQNDRDR (SEQ ID NO:190) SSKIAESTEWQEK (SEQ ID NO:191) SSWEIHER (SEQ ID NO:192) Latent transforming growth factor beta IPIOO4651.45 AQPGWGSPR (SEQ ID NO.193) 0.82 binding protein 2
EQDAPVAGLQPVER (SEQ ID NO:194) RPGGSYPAAAAAK (SEQID No.195) STPLGQQQPAPR (SEQ ID NO:196) YEPAGGDANR (SEQ ID NO:197) Neuroblastoma suppressor of tumorigenicity l IPIO0013299 LALFPDK (SEQ ID NO:198) 1.55 NC 109 OCCTSO sAwcEAK (SEQID No.199) Splice isoform 1 of SWI/SNF-related, matrix associated, IPIO02201 19 ANTPDSDITEK (SEQID NO:200) actin-dependent allelson.
FIG.5G U.S. Patent Aug. 18, 2009 Sheet 12 of 97 US 7,575,876 B2
re. Peptide Sequences TRAQ Ratios regulator HGLNGILADEMGLGK (SEQ ID NO:201) NLFNLDR (SEQ ID NO:202) QELREVLK (SEQ ID NO:203) SADEQSIYEKER (SEQID NO:204) SLFRRLK (SEQ ID NO:205) VFAEDQDMQYASQSEVPNGK (SEQ ID NO:206) YQHLMTINANNR (SEQ ID NO:207) Cell Structure/Motility/Transport/Traffic H 107 kDa protein poo76999 (EPSVIDFNQNIR (SEQID 0.28 NC 0. 99 AHGLIGOFMQEPK (SEQ ID NO:209) AHVSFKPTVAQQR (SEQID NO:210) FQLVAENR (SEQID No.211) FYNQVSTPLLR (SEQ ID NO:212) IQPSGGTNINEALLR (SEQID NO:213) IYGNQDTSSQLK (SEQED NO:214) SLAPTAAAK (SEQ ID NO:215) SSALDMENFR (SEQ ID NO:216) YEK (sEQID No.217) Alpha-l-acid glycoprotein 1 IPIO0022429 EQLGEFYEALDCLR (SEQ ID NO:218) 2.49 precursor KQEEGES (SEQID NO:219) TEDTIFLR (SEQID NO:220) WFYIASAFR (SEQ ID NO:221)
APPOP',OreclSO 'Poo021854 DLMEK (SEQID No.222)
0. 7 8 1 O 5
FIG. 5H CC U.S. Patent Aug. 18, 2009 Sheet 13 of 97 US 7,575,876 B2
Apollipoprotein D yeCS
VLNQELR (SEQID NO:238) WYEIEK (SEQID NO:239) AATVGSLAGQPLQER (SEQ ID NO:240)
LASHLR (SEQ ID NO:249) LEEQAQQIR (SEQID NO:250)
SELEEQLTPVAEETR (SEQ ID NO:256 SWFEPLVEDMQR (SEQ ID NO:257) VEQAVETEPEPELR (SEQ ID NO:258) WELALGR (SEQID NO:259)
FIG. 5
U.S. Patent Aug. 18, 2009 Sheet 17 Of 97 US 7,575,876 B2
Peptide Sequences iTRAO Ratios
SSGLVSNAPGVQIR (SEQ ID NO:356) STELCOLWQGR (SEQ ID NO:357) 0.656 l AEAQEAEDQQAR (SEQID NO:359) AGSPLLWGPR (SEQ ID NO:360) ALAHLLEAER (SEQ ID NO:361) ARAEAQEAEDQQAR (SEQ ID NO:362 GEAAGAVQELAR (SEQ ID NO:363) ILAGSADSEGVAAPR (SEQ ID NO:364 ILAGSADSEGVAAPRR (SEQ ID NO:365) LETPAPQVPAR (SEQID NO:366) MAGSPLLWGPR (SEQID NO:367) NSDPALGLDDDPDAPAAQLAR (SEQ VLAQLLR (SEQID NO:369) VWGAPR (SEQID NO:370) SPSS"SPIO0218733l, soluble AvCVLK (SEQID No.371) GDGPVQGIINFEQK (SEQ ID NO:372) GGNEESTK (SEQ ID NO:373) LACGVIGIAQ (SEQ ID NO:374) VWGSIK (SEQ ID NO:375) Transcription elongation regulator PIO0247871 FKAIEK (SEQ ID NO:376) 2.33 t 1 YLVLDCVPEERR (SEQID No.377) Vacuolar ATP synthase subunit Sl IPI00020430 5:NP EVIGQVLSTLK (SEQ ID OneClSO
FIG.5M U.S. Patent Aug. 18, 2009 Sheet 18 of 97 US 7,575,876 B2
Protein Name Peptide Sequences iTRAQ Ratios DLB LGASPLHVDLATLR (SEQ ID NO:379) LPYTASSGLMAPR (SEQ ID NO:380) LSIEDFTAYGGVFGNK (SEQ ID MMAAMATARVRMGPR (SEQ ID NO:382) NVLLFLQDK (SEQ lD NO:383) TTT SETALTavers stop | | | | | | EExtracellular Matrix/Cell Adhesion | | | | | | Cochlin precursor IPI00012386 GVISNSGGPVR (SEQID NO:385) 1.22 NC TFEISDIGAK (SEQ ID NO:386) | | | | | VYSLPGR (SEQ ID NO:387) | | | | | | WSASFTVTK (SEQ ID NO:388) | | | | Matrix Gla-protein precursor IPIO0028714 ERSKPVHELNR (SEQ ID NO:389) 1.51 NANTFISPQQR (SEQ ID NO:390) | | | | Spondin 1 precursor IPI00171473 LCGGGIQER (SEQ ID NO:391) 176 it 09 NC 0.93 SEQLKEESEGEQFPGCR (SEQ ID NO:392) SSQFTSCK (SEQID NO:393) | | | | | | VVIERIAR (SEQID NO:394) Immunity/Defense 24 kDa protein IPIOO479531 EQLGEFYEALDCLR (SEQ ID NO:395) 3.02 t 10 5 N C KQEEGES (SEQ ID NO:396) NEEYNK (SEQID NO:397) NWGLSVYADKPETTK (SEQ ID NO:398) SDVVYTDWK (SEQID NO:399) TEDTIFLR (SEQ ID NO:400) TYMLAFDVNDEK (SEQID NO:401) WFYIASAFR (SEQ ID NO:402) YVGGQEHFAHLLILR (SEQ ID NO:403) AMBP protein 2.09 OecurSO IPIO0022426 AFIQLWAFDAVK (SEQ ID NO:404) f t C C
ETLLQDFR (SEQ ID NO:405) 0.80. EYCGVPGDGDEELLR (SEQ ID 87 FYSEK (SEQ ID NO:407)
FIG. 5N U.S. Patent Aug. 18, 2009 Sheet 19 of 97 US 7,575,876 B2
Protein Name Peptide Sequences AD iTRAOPD Ratios | DLB GECVPGEQEPEPILIPR (SEQ ID NO:408) GPCRAFIQLWAFDAVKGK (SEQID NO:409) TVAACNLPIVR (SEQID NO:410) VVAQGVGIPEDSIFTMADR (SEQID NO:41 l) Complement C2 OreCuSOr IPIO0303963 DFHINLFR (SEQ ID NO:412) 0.59 0.89 NC | 1.07 NC QHLGDVLNFLPL (SEQ ID NO:413) Cytokine-like protein C17 precursor IPIO0032876 ALSQEITR (SEQ ID NO:414) NC DFNLLQVSEPSEPCVR (SEQ ID
Fibrinogen beta chain IPIOO298497 AHYGGFTVONEANK (SEO ID OrecurSOr NO:416) Q (SEQ 1.55 0.97 NC 1.14 NC EDGGGWWYNREEAPSLRPAPPPISGGGYR (SEQID NO:417) (SEQ ID | | | | | NO:418) GSWYSMR (SEQID NO:419) KGGETSEMYLIQPDSSVKPYR (SEQ ID NO;420) KWDPYK (SEQ ID NO:421) MGPTELLIEMEDWK (SEQ ID QDGsvDFGR (SEQID No.423) QGFGNVATNTDGK (SEQID No.424) SILENLR (SEQID NO:425) | | YQISVNK (SEQID NO:426) YYWGGQYTWDMAK (SEQID HLA class I histocompatibility antigen, B-27 alpha IPIOO47 1986 AQTDREDLR (SEQ ID NO:428) chain precursor FDsDAASPR (SEQID No.429) sinces LLRGYHQDAYDGK (SEQID No.430) YTCHVOHEGLPKPLTLR (SEQ ID NO:43 HLA class I histo"Flyantigen, E alpha IPI0001.0362 FDNDAASPR (SEQID No.432) 1.28 NC 1.18 NC chain precursor
FIG. 5O
U.S. Patent Aug. 18, 2009 Sheet 22 of 97 US 7,575,876 B2
iTRAO Ratios
Peptide Sequences L
FAHYVVTSQVVNTANEAR (SEQ ID NO:492)
GSLVQASEANLQAAQDFVR (SEQ ID NO:493 QYYEGSEIVVAGR (SEQ ID NO:494) KIAA 1318 protein IPI00002353 APASGGVSSPLVR (SEQ ID NO:495) .2 3 1 7 NC
ASVSGSMPMPLPR (SEQID NO:496) FLEGLSEAVTTKMGRIFLK (SEQ ID NO:497) MATAPIRAsAsGAR (SEQID No.498) VTSTSQMMPTASGDMCTLPVR (SEQ ID NO:499 Uveal autoantigen IPI00173359 EHEK (SEQ ID NO:500) l NCC
EHLTSEAASGNHR (SEQID NO:501) CC ELEAMR (SEQ ID NO:502) IQQEQRILLDK (SEQID NO:503) KELEAMR (SEQID NO:504) LEMEK (SEQID NO:505) LMKAAER (SEQ ID NO:506) NLTHMQDEVNVK (SEQ ID NO:507)
SLNGTIENLK. (SEQ ID NO:508) TNRELLDVK (SEQ ID NO:509) VPS10 domain containing receptor IPIOOO 10381 AGPELLPQQGGGR (SEQ ID NO:510) N C 105 NC SorcS3 precursor AQMCPGKAPRGLHVVTTDGR (SEQ ID NO:511
AVASQWPEELASAR (SEQ ID
GGEMQVEAGGTSPAGER (SEQ ID NO:513 GGEMQVEAGGTSPAGERR (SEQ ID NO:514) E GLHVVTTDGRGIPAPAK (SEQ ID(SEQID NO:515) NO:516) II. Proteins unique to AD and identified by single peptide Neuronal Activities/Signal Transduction
protein""PhOS 15 precursor PI00001485 VLLSILR (SEQID No.517) ll NC 1.08
sphingolipidE"E"P.PI00015343 receptor SSVSDYVNYDIVR (SEQID No.518) 0.62- 0.86 NC
FIG. 5R U.S. Patent Aug. 18, 2009 Sheet 23 of 97 US 7,575,876 B2
Protein Name P. Peptide Sequences iTRAO Ratios DL B IL-7RC IPIO0303074 DDVLLLETR (SEQID NO:519) 0.6 Interleukin 1 receptor-associated IPI00304986 DFSTSIPK (SEQID NO:520) 0.5 1 C 0.88 NC kinase-like 2 Metallothionein-III IPI00016666 GGEAAEAEAEK (SEQ ID NO:521) 0.4 0. 9 5 0.0. 9877 Neural proliferation differentiation and control protein-1 IPIO0299699 LEDEIDFLAQELAR (SEQ ID NO:522) 0.51 1.05 OTCCUISO Potassium voltage gated channel IPIO0012857 DGTLLLEGGGR (SEQID NO:523) J. 0. 9 6 NC subfamily KQT member 3 Putative 4 repeat voltage-gated ion IPIO02 17996 SLQLEELLAR (SEQID NO:524) 1.26 0.91 NC channel Splice Isoform 1 of protachykinin 1 IPIO0023571 EELPEPFEHLLOR (SEQ ID NO:525) NC 0.87 NC OCuSO Splice isoform 2 of UDP-N- peptidea" N PI00219856 IVLNGIDLK (SEQID No.526) NC 101 NC acetylglucosaminyltr ansferase 11 Voltage-dependent calcium channel IPIO0011072 GAEFLLR (SEQ ID NO:527) NC 1.2 NC gamma-6 subunit Cell Cycle/Death | | | | collagenl'E' isoform 3 PI00375409 GEAGLDGAK (SEQID No.528) 1.26 | f | 0.63 NC Isambrane IPIO0031821 IENIDHLGFFIYR (SEQ ID NO:529) 1.8 | f | 0.94 | NC NC Cell structure/Motility/Transport?Traffic | | | | | |
muscleActin, aortic smooth pooooo SYELPDGQVITIGNERNO:530 (SEQID 1.72 | f | 1.05 NC NC
1 hesis"b"S^ P100025 15s. ToIDDR (SEQID No.531) 1.64 0.91 Hepatocellular carcinoma associated IPI00293898. LFAEEK (SEQID NO:532) 1.57 | f | 1.01 NC 1.05 NC protein TB6
SAA1 protein
ID NO:533) isomors opinc SAYY8238 IPIO0.432771 LQDMEK (SEQID NO:534) Splice isoform 1 of IPI00337307 VILAIR (SEQID NO:535)
FIG. 5S U.S. Patent Aug. 18, 2009 Sheet 24 of 97 US 7,575,876 B2
Peptide Sequences iTRAQ Ratios PD DLB hpalracinnam Itiny fragments re. locus 9c protein Splice isoform 2 of development and differentiation IPIOO4096.13 EIISEVQR (SEQID NO:536) 1.96 f 0.93 NC 103 NC enhancing factor 2 Splice isoform 2 of putative polypeptide N acetylgalactosaminylt IPIOO456715 AYLSAK (SEQID NO:537) 0.66 1.07 NC ransferase-like protein Eated 'P' PI003292.81. INSCPLSLSWGKR (SEQID No.538) 1.87 Metabolism ADAM 10 precursor IPIOOO13897 DTSLFSDEFK (SEQ ID NO:539) to 99 NC 0.91 NC Alpha-1,3-mannosyl glycoprotein 2-beta N IPIO0000138 WALGOVFR (SEQ ID NO:540) 0.49 1.02 NC 0.91 NC acetylglucosaminyltr ansferase Schrome ' PI00218839 VIIFGMGK (SEQID No.541) 103 NC 0.88 NC DNA-directed RNA polymerase I largest IPIO0031960 SITNPR (SEQID NO:542) 0.59 0.89 1.32 subunit Heat shock 10kDa protein 1 (chaperonin IPI00220362 FLPLFDR (SEQID NO:543) 10) rescore E. domain " PI00333067 KSDFFINK(SEQID NO:544) t 0.67 0.9 NC - More poisos NKPGVYTK(SEQ ID NO:545) t 1.12 NC 106 NC PPIB protein IPIO0419262 VIFGLFGK (SEQID NO:546) 0.55 l 1.03 NC 0.88 NC
Splice isoform 2 of insulin receptor IPIO0220325 VTDLMR (SEQID NO:547) 59 NC OCCUSO Transcriptional activator SRCAP IPIO0009101| LEAEGMRGR (SEQ ID NO:548) 0.59 0.94 NC 0.84 NC Eger P'"'PI00032316 ELEERR (SEQID No.549) 19 NC ZNF627 protein IPIO0029023. RNISHIPER (SEQID NO:550) 3.04 0 || 0 || 0 || 0 Extracellular Matrix/Celi Adhesion | | | | hit "PIO0028670. EGSDLSvvER (SEQID No.551) 0.66 l, 14 NC 106 NC
FIG. 5T U.S. Patent Aug. 18, 2009 Sheet 25 Of 97 US 7,575,876 B2
DLB EAEAWAKPGAAARR (SEQ ID KIAA 1730 protein IPIOO155199 NO:552) 1.63 misncos l PREDICTED: KIAA0527 protein IPIO0297224 GSGEQQIMR (SEQID NO:553) 0.66 NC PREDICTED: odz, odd Oz/ten-m NTMAMK (SEQID NO:554) 0.72 NC 1.91 homolog 3 Splice isoform l of ADAMTS-16 IPIO0386697 SEYElysayevo-R (SEQ ID 0.64 0.87 NC 0.93 NC Oecursor
Immunity/Defense Pregnancy-specific beta-l-glycoprotein 8 IPIO0334256 LQLSETNR (SEQID NO:556) 2.79 it 0.88 NC precursor Unknown 141 kDa protein IPI00478948. TQLDDR (SEQID NO:557) 15 kDa protein IPIOO413387 IVEPYVTFGFPNPK (SEQ ID NO:558)
Antigen MLAA-20 C1 or f40 protein
NC 1.15
Hypothetical protein DKFZp434A2017 IPIO0295380. EADVVAR (SEQID NO:567) Hypothetical protein DKFZp666G229 Hypothetical protein DKFZp686A06175 Hypothetical protein IPIOO185662 APAKPPGSGLDLADALDDQDDGR 0.32 DKFZD761H2O24 SEO ID NO:570 0.9 3. Inc.C NcC HypotheticalDKFZ781A0122 protein IPIOO470805 DAEEDMPQR (SEQ ID NO:571) 0.66 ancios Nc Hypothetical protein PSEC0200 IPIOO166392 SDDSVIQLLNPNR (SEQ ID NO:572) 1.54 in Inc. 1.11
KARCA1 protein pool NO:573WLCVVGGWDGSRR (SEQID 581 1.06 NC 0.57
Kelch/ankyrin repeat containing cyclin Al IPIOO449308 SygowDosRR SEQ ID 5.65 t interacting protein
F.G. 5U U.S. Patent Aug. 18, 2009 Sheet 26 of 97 US 7,575,876 B2
iTRAO Ratios Protein Name IP it Peptide Sequences AD PD DLB Earphy IPIO0107749 LHLVSR (SEQ ID NO:575) PREDICTED: hypothetical protein IPI00397059 YGEEIK (SEQ ID NO:576) 0.66 1.04 NC 0.84 NC XP 374046 PREDICTED: similar to melanoma IPI00455972 VLEAILR (SEQID NO:577) 0.64 1.06 NC 1.08 NC antigen, family A, 10 Similar to expressed TFASPNASGSGNTGAR (SEQ ID sequence AI5934.42 IPIO02 17781 NO:578) 0.45 1.18 NC 0.86 NC "T" III.iii. ProteinProteins unique to Pd.PD and identified by 2 or more peptides Neuronal Activities/Signal Transduction Amyloid-like protein 1 precursor IPIO0020012 SLEFAALOADPPQAER (SEQ ID 1.1 N C C DDTPMTLPK (SEQ ID NO:580) DELAPAGTGVSR (SEQID NO:581) EWAMADNQSK (SEQID NO:582) FLHQER (SEQID No.583) GGLQPPDSK (SEQ ID NO:584) GSTEQDAASPEK (SEQ ID NO:585) GSTEQDAASPEKEK (SEQID NO:586) LVETHATR (SEQID NO:587) MDQCESSTR (SEQID NO:588) MNPLEQYER (SEQ ID NO:589) QMYPELQIAR (SEQID NO:590) vEQATQAIPMER (SEQID No.591) VIALINDQR (SEQID NO:592) vLEYCR (SEQID No.593) VLLALR (SEQID NO:594) WEPDPQR (SEQID NO:595) - Cell growth regulator with EF hand domain|IPI00337548 ELPGETLESK (SEQ ID NO:596) 0.77 1.67 | 1.12 NC
ID NO:597) SESAEAEGDAporristoldGEAGGQAEAEGDAPGPR (SEQ ID | | | | GEAGGQAEAR (SEQ ID NO:600) HVEPGEPLAPSPQEPQAVGR (SEQ ID
F.G. 5V
U.S. Patent Aug. 18, 2009 Sheet 36 of 97 US 7,575,876 B2
Protein Name Pi Peptide Sequences iTRAQ Ratios immunoglobulin NO:864) receptor precursor t DLB LFAEEK (SEQID NO:865) Unknown 132 kDa protein IPIOO477893 ELGQMNLTER (SEQ ID NO:866) 1.48 0.49 1.01 NC EVEEEMEK (SEQID NO:867) FGEIYEK (SEQ ID NO:868) | | | FKNEVNTLEEEFLALK (SEQ ID NO:869 KMNSEFHSAAK (SEQID No.870) LEDLGELHRAAR (SEQ ID NO:871) LLIEER (SEQ ID NO:872) NMLERGEGER (SEQ ID NO:873) QMENMVSVLQNELSETKK (SEQ ID NO:874)
SGDVPGVEHVLAPGDTGVDKR (SEQ ID NO:876) SYMER (SEQID NO:877) Hypothetical protein DKFZp566O224 IPIOO383815
LALDIEIATYR (SEQ ID NO:881) PREDICTED: G2 protein AASGPK (SEQ ID NO:882) GFLNFMNTVLVAFTK (SEQ ID NO:883)
IV. Proteins unique to PD and identified by single peptide Neuronal Activities/Signal Transduction
Singeceptor Poissis VELDTR (SEQID NO:885) O.97 O.72 | 1.62 PREDICTED: ETPPLEDLAANQSEDPR (SEQ ID kia A337 protein P0000228 Nosso) 0.97 NC 0.64 t PREDICTED: similar to 28 kDa heat- and acid-stable phosphoprotein- IPIO0376589 ADLAQLAIIR (SEQ ID NO:887) 1.53 (PDGF-associated protein)
FIG. 5FF U.S. Patent Aug. 18, 2009 Sheet 37 Of 97 US 7,575,876 B2
AD D L B Sticulon '''" PI00335276 GPLPAAPPVAPER (SEQID No. 888) 098 N C 5 GSSGSVVVDLLYWR (SEQ ID NO:889) activating protein..., 10 P1001693.07 GSWGSGK (SEQID No.890) 1 NC 5 9 9 t tl 0. 48 FIF Cell Cycle Death collagen‘EE", isoform 3 PI00375409 GEAGLDGAK (SEQID No.891) 1.26 f 0.63 Cell Structure/Motility/Transport/Traffic ATP-binding cassette, sub-family IPI00293460, QNTADILQDLTGR (SEQID NO:892) 1.1 | NC 1.57 | f | 1.05 NC A member 1 Hypothetical protein DKFZ434P097 IPIO001 1232 LSAEILRLEK (SEQ ID NO:893) 1 NCO 5 5 0. 8 9 Hypothetical protein FLJ32842 DPQSTELPR (SEQ ID NO:894) C 0. 9 7
Putative 4 repeat voltage-gated ion SLQLEELLAR (SEQ ID NO:895) 0.64 26 O.91 N C channel Ribonuclease 4 OCCUISO YCNLMMQR (SEQID NO:896) 0.88 NC 1.68 ft | 1.08 NC Ribosomal protein L3-like VAWAQARLEK (SEQ ID NO:897) 1 NC 0.55 0.89 NC SAA 1 protein IPIOO452748 FFGHGAEDSLADQAANEWGR (SEQ ID NO:898) f 0.78 0.91 NC Splice isoform 1 of transcription factor IPIOOO13929 AADGSLDTQPK (SEQ ID NO:899) 0.84 N C 0.72 1.74 1. E2-alpha Metabolism Metabotropic glutamate receptor 3 IPI00478.165 ELIAAASR (SEQID NO:900) 0.86 NC 0.51 2. 4 3 precursor Extracellular matrix/Cell Adhesion Laminin gamma-l IPIO029828lNTIEETGNLAEQAR (SEQ ID NO:901) 0.93 NC 1.37 0. 6 chain precursor Mammalian ependymin related IPI002591.02 AGGSHSDPGR (SEQ ID NO:902) NC 0.57 155 protein 1 PREDICTED: odz, odd Oz/ten-m IPIO0398.020 NTMAMK (SEQID NO:903) 0.72 0.99 NC homolog 3 o integrinSEESE", alpha-7 PIOO220748ELEPPEQQEPGER (SEQID No.904) NC 0.64 Cft FIG. 5GG U.S. Patent Aug. 18, 2009 Sheet 38 of 97 US 7,575,876 B2
Protein Name Peptide Sequences AD PD DLB precursor Immunity/Defense | | | | | | Ig kappa chain V-1 region HK102 IPIOO478600 Estesovsk (SEQ ID 1.1 NC 1.53 0.87 NC precursor - Unknown 97 kDa protein IPI00472544 ELEERR (sEQID No.906) 0 || 0 |27s | 0 || 0 DJ977L-1 IPIOO478622 LQEAAEIVK (SEQID NO:907) 1.55 ft 0.61 0.94 NC HRPE773 YFSTTEDYDHEITGLR (SEQ ID 0.99 NC 0.44 1.78 NO:908) 1.08 Ncoss 1.12 NC Hyphal P'" PI00442326 MDRPSLVR (SEQID No.910) 1.13 0.93 NC Hypothetical protein AEAGGGWEGSASYK (SEQ ID 0.84 NC 1.69 | f | 0.91 NC FLJ46550 NO:91) OTTHUMPOOOOOO2 1593 IPIO0374531. TIEELAR (SEQID NO:912) O 0 1 2.75 O O V. Proteins unique to DLB and identified by 2 or more peptides N euronal Activities/Signal Transduction AALGESGEQADGPK (SEQ ID 110 kDa protein IPOO473056 NO:913) O.9 NC 0.93 NC 0.63 DLLGQQPHSEPGAAAFGELQNQMPG PSK (SEQ ID NO:914) EEQSLPAGAQEALSDGLQLEVQPSEE EAR (SEQ ID NO:915) HLPFLEALSQAPASDVLAR (SEQID NO:916) RPEASSPARPSKHSVGSER (SEQ ID NO:917) VPAMDFYR (SEQID NO:918) YEVSPVALQR (SEQID NO:919) Brain abundant, membrane attached IPIOO299024 AEPPKAPEQEQAAPGPAAGGEAPK signal protein 1 (SEQ ID NO:920) 0.39 NC 34 EADVVAR (SEQID NO:921) EKPDQDAEGK (SEQID No.922)
ESEPQAAAEPAEAK (SEQID No.923) Cocaine- and amphetamine pooooo, ALDIYSAVDDASHEK (SEQID 0.89 NC 107 NC 1.74 regulated transcript NO:924) protein precursor F ELIEALQEVLK (SEQ ID NO:925)
F.G. 5HH
U.S. Patent Aug. 18, 2009 Sheet 41 of 97 US 7,575,876 B2
P h Peptide Sequences iTRAQ Ratios DLB NO:981) F QFLQK (SEQ ID NO:982) SANSNPAMAPR (SEQ ID NO:983) sLAAAAGK (SEQID No.984) Splice isoform 1 of basigin precursor 0. 9 8 sC .21 f 0.67 Splice isoform 1 of receptor-type tyrosine-protein LSATLGGLLQDHGSR (SEQID 0.92 NC 0. 9 NC 0.54 phosphatase N2 NO:987) LYQEVHR (SEQ ID NO:988) SQTYSK (SEQ ID NO:989) VALQK (SEQ ID NO:990) H VPAMDFYR (SEQID NO:991) YEVSPVALQR (SEQID NO:992) Splice isoform 3 of DMIGRCFVLSQDLAIR (SEQ ID integrin alpha-7 1.13 NC 0.84 C 1.51 t precursor NO:993) DSASRLVPEVMLSGER (SEQ ID NO:994) ELEPPEQQEPGER (SEQ ID NO:995) GAVVILR (SEQID NO:996) GLVRAEELSFVAGAPR (SEQ ID NO:997) VCGDAMFQLQENVK (SEQ ID NO:998) VDQILETRDMIGR (SEQID NO:999)
YEARQR (SEQ ID NO:1000) Cell cycle/death Fas apoptotic APGTEGQQQVHGEK (SEQ ID inhibitorv molecule 2 IPIO001 7.569 NO:1001) 0.88 NC 0.85 NC 0.56 EAPAVPSAPPSYEEATSGEGMK (SEQ ID NO:1002 LSVANK(SEQID No. 1003) Hypothetical protein FLJ16490 IPIO0465099 ICYVCK (SEQID NO:1004) 1.1 O.63 El EGQAVAVPSSK (SEQ ID NO:005) Latent transforming growth factor beta IPIOO4651.45 AOPGWGSPR (SEQ ID NO:1006) 17 NNCC O.82
FIG. 5KK U.S. Patent Aug. 18, 2009 Sheet 42 of 97 US 7,575,876 B2
Protein Name IPI # Peptide Sequences iTRAO Ratios D L B binding protein 2 EQDAPVAGLQPVER (SEQ ID NO:1007) RPGGSYPAAAAAK (SEQ ID NO:1008 STPLGQQQPAPR (SEQ ID NO:1009) | | | | | YEPAGGDANR (SEQID No:1010) || | | | | | Splice isoform 1 of SWI/SNF-related, matrix associated, IPIO02201 19 ANTPDSDITEK (SEQ ID NO:1011) 6 1 f t NC 0.71 actin-dependent regulator HGLNGILADEMGLGK (SEQ ID NO: 1012) NLFNLDR (SEQ ID NO: 1013) QELREVLK (SEQID NO:1014) HERSADEQSIYEKER (SEQ ID NO: 1015) VFAEDQDMQYASQSEVPNGK (SEQ ID NO: 1017) YoHLMTINANNR (SEQID No:1018) Cell Structure/Motility/Transport/Traffic LOQTQAQVDEVVDIMR (SEQ ID 12 kDa protein Points NO:1019) NC 0.92 NcC 0. 5 7 MMIILGVICAIILIIIIVFFSG (SEQ ID NO:1020) Apollipoprotein C-III IPIOOO21857 DALSSVQESQVAQQAR (SEQ ID NC 0.57 1 2 8 recursor NO:1021) DYwsTVK (sEQID No. 1022) GWVTDGFSSLK (SEQ ID NO: 023) PPP"'P100021856 ESLSSYWESAK (SEQID No. 1024) 1.13 NC C 5 7 t OCCUSO in STAAMSTYTGIFTDQVLSVLK (SEQ ID NO:1025) TAAQNLYEK (SEQID NO:1026) TYLPAvdEK (SEQID No. 1027)
Divalent cation tolerant protein IPIO0034319 LLLLPR (SEQ ID NO:1028) O. 8 2 0.96 N 2.07 ft CUTA TQSSLVPALTDFVR (SEQ ID NO: 1029 Golgi phosphoprotein IPIO0171411 DLSENNDQR (SEQID NO:1030) 2.08 tf 0.89 N C 0. 7 7
F.G. 5LL U.S. Patent Aug. 18, 2009 Sheet 43 of 97 US 7,575,876 B2
Protein Name Peptide Sequences AD 2
EETNEIQVVNEEPQR (SEQ ID NO:1033) GFGGAGELGQTPQVQAA (SEQ ID NO: 1035) GFGGAGELGQTPQVQAALSVSQEN (SEQ ID NO:1036)
MGLGNGRRSMK (SEQ ID NO:1039) NIDVFNVEDQK (SEQ ID NO:1040) NIDVFNVEDQKR (SEQID NO: 1041) NQTNLERKFSYDLSQCINQMK (SEQ ID NO:1042) QQLQALSEPQPR (SEQ ID NO: 1043) RDTINLLDQREK (SEQ ID NO:1044) Hemopexin precursor IPI00022488. DYFMPCPGR (SEQID NO:1045)
GHQDLDPDNEGELR (SEQ ID NO: 1053)
RPPRPGTNGwsRR (SEQID No. 1057) IssTQMTwoALFR (SEQID No. 1058) Latent transforming growth factor-beta- IPI00292150I AEEEELARPPR (SEQ ID NO:1060) binding protein 2
FIG. 5MM
U.S. Patent Aug. 18, 2009 Sheet 48 of 97 US 7,575,876 B2
Protein Name IPI # Peptide Sequences iT RAQ Ratios D L B SLDFTELDVAAEK (SEQ ID NO:1178) TSPVDEK (SEQ ID NO:1179) VANPLSTA (SEQID NO: 1180) VLSALQAVQGLLVAQGR (SEQ ID NO: 1181) Apolipoprotein Cl IPIO0021855 EFGNTLEDK (SEQ ID NO:1182)
EWFSETFQK (SEQ ID NO:1183) LKEFGNTLEDK (SEQ ID NO: 1184)
QSELSAK (SEQID NO: 1185) Coagulation factor VIPI004 19311 ASEFLGYWEPR (SEQ ID NO:1186) NNCC 0. 4. 0.0. 6458 EFNPLVIVGLSK (SEQID NO:1187) Enolase 2 IPIOO216171 AAVPSGASTGIYEALELR (SEQID NO: 1188) 18 1. 3 2 GNPTVEVDLYTAK (SEQ ID NO: 1189) YITGDQLGALYQDFVR (SEQID Hypothetical protein AAVPSGASTGIYEALELR (SEQ ID Disc2" (PIOO465248 No.5) 0.57 3 2 GNPTVEVDLFTSK (SEQID No. 1192) VVIGMDVAASEFFR (SEQ ID CCCC NO:1193) E. IPIOOO19038 AWVAWR (SEQ ID NO:1194) 0.96 NC 0.88 NC GISLANWMCLAK (SEQID No. 1195) STDYGIFQINSR (SEQ ID NO:1196) N acetylactosaminide beta-1,3-N- IPIO0009997 EAENOHNK (SEQ ID NO:1197) acetylglucosaminyltr ansferase EMLDQSNQWGGTALVVPAFEIR
(SEQID NO: 1198) EPGEFALLR(SEQ ID NO:1199) QYGFNR (SEQID No. 1200) svdovK (SEQID No.1202) NO:ASVLDAsGDYR 1203 SEQID | TALASGGVLDASGDYRVYR (SEQ ID NO:1204) vPTFDER (SEQID No. 1205)
FIG. 5RR U.S. Patent Aug. 18, 2009 Sheet 49 of 97 US 7,575,876 B2
Protein Name Peptide Sequences iTRAO Ratios D L WEGPLSVSVFAATK (SEQ ID NO:1206) YEAAVPDPR (SEQ ID NO.1207) YPNSPR (SEQ ID NO:1208) Probable endonuclease IPIO000 1952 DSDIIEDVMVK (SEQID NO:1209) 0.88 NC NC 1.75 KIAA0830 precursor ICQRAEGAER (SEQ ID NO: 1210) ILEVVNQIQDEER (SEQID No. 1211) WYVNLHSLMDR (SEQ ID NO:1212)
Rho-associatedbrotein kinase 1 IPIO0022542 DVEMEPVOQAEK (SEQ ID NO: 1213) 0.85 NC 0.88 NC 0.79 GNEGQMR (SEQID NO.1214) INEYQRKAEQENEK (SEQ ID NO:1215) KANTQDLR (SEQ ID NO:1216)
LSQLQK (SEQID NO:1217) NDQWAWETLR (SEQ ID NO:1218) NINTERTLK (SEQID NO:1219) QLEEANDLLR (SEQID NO: 1220)
SQGGDGYYGR (SEQID NO: 1221) VVKNTSGK (SEQ ID NO: 1222) Similar to peptide N glycanase homolog ELLOR (SEQID NO: 1223) 0. 9 1 NC 106 NC 0. 6 l FECGSVGLK (SEQ ID NO:1224) GEMGLQR (SEQ ID NO:1225) IRDLIAIERSSR (SEQ ID NO:1226) KVETDWHMVYLARK (SEQ ID NO: 1227) QLFLSENRR (SEQID NO: 1228) YVWDYTELQRTLSLK (SEQID NO:1229) Sortilin-related PI00022608DEQYLFLVR (SEQID NO:1230) 1.06 NC 1 l 4.
F GFLVVQGDPR (SEQ ID NO: 1231) IITENDHVLLFWKSLALK (SEQ ID NO:1232 LNFGLGNR (SEQID No.1233) TDLGDSPLAFEHVMTR (SEQ ID NO:1234
FIG. 5SS
U.S. Patent Aug. 18, 2009 Sheet 51 of 97 US 7,575,876 B2
Protein Name IPI # Peptide Sequences iTRAQ Ratios A D PD D L B LACGVIGIAQ (SEQ ID NO: 1261) VWGSIK (SEQID NO: 1262) Transcription elongation regulator IPI00247871 FKAIEK (SEQ ID NO: 1263) 2.33 0.94 NC 0. 7 l YLVLDCVPEERR (SEQ ID NO:1264) Extracellular Matrix/Cell Adhesion Hypothetical protein DLPETLNELHLDHNK (SEQ ID FLJ35635 IPI00385748|Nois) 0.92 NC 0.89 NC 0. 6 5 IQAIELEDLLR(SEQID No. 1266) NC 1.03 NC 0.61 isoform precursor SESAVNick (SEQID DESKEPIVEVR (SEQ ID No. 1268) FFLCQvAGDAK (sEQID No. 1269) GLGEISAASEFK (sEQID No. 1270) IFQK (SEQ ID NO.1271)
T-Cadherin O. 5 8 0. 8 5 GIFRINENTGSVSVTR (SEQID NO:1276) TLFVHAR (SEQ ID NO: 1277) TPHAEDMAELVIVGGK (SEQ ID NO:1278) VDCNAAGALR (SEQID NO:1279) VNSDGGLVALR (SEQID NO:1280) YEVSSPYFK (SEQ ID NO:1281) Immunity/Defense CD99L2 protein IPIOO434755 APAKPPGSGLDLADALDDQDDGR 1.11 1.63 O. 4 (SEQ ID NO: 1282) 2 APANTLGNDFDLADALDDR (SEQ ID NO: 1283) CC tC Polymeric ASVDSGSSEEQGGSSR (SEQ ID immunoglobulin IPIO0004573 reCetO OreCurSO NO:1284) LFAEEK (SEQID NO:1285) Unknown Hypothetical protein IPI003.01255 DLDDTK (SEQID NO:1286) O 9 8 NC 11 NC 1.29
FIG. 5UU U.S. Patent Aug. 18, 2009 Sheet 52 of 97 US 7,575,876 B2
Protein Name Peptide Sequences iTRAQ Ratios PD DLB FLJ90835 DLDDTKMQKSLSLLDAENR (SEQ ID NO:1287) IVMTPSR (SEQ ID NO.1288) | | | | | SLSLLDAENR (SEQ ID NO:1289) TDGRMR (SEQ ID NO: 1290) PREDICTED: FLJ46675 protein IPIO0165319 AGMLVSGLAGEK (SEQID NO:1291) 1.17 | NC 0. 89 C O. 6 2 ASTLTIGWRAQEMSEK (SEQ ID NO:1292) DLEFEEDQR (SEQ ID NO:1293) DVLEK (SEQID NO: 1294) EMFLMAAMGPPGGGR (SEQ ID NO: 295 FHALSLGOGQAPIAAR (SEQID NO:1296) FLILOTETMETTAHGLFRR (SEQ ID NO:1297) IDSYLREIEGSFPNK (SEQID NO: 1298 LVEDLGLFPGR (SEQ ID NO:1299) MFEKLINK (SEQ ID NO:1300) MMEDALR (SEQ ID NO: 1301) NCHLALR (SEQ ID NO: 1302) NMEGGQGLK (SEQID NO: 1303) QELLAQANK (SEQID NO: 1304) VELDALQQIWEIAR (SEQ ID NO: 1305) VGRNGGEAEEK (SEQ ID NO: 1306) VIGOPRGNMLLVGIGGSGR (SEQ ID NO: 1307) YIREMFLMAAMGPPGGGR (SEQID NO: 1308) PREDICTED: G2 protein IPIOO176482 AASGPK (SEQ ID NO: 1309) 0.9 C l 2 8 t O. 5 8
GFLNFMNTVLVAFTK (SEQID NO:1310) LLPISPTWPFTEVR (SEQID NO: 1311) PREDICTED: hypothetical protein IPI00456355, LLYEK (SEQID NO:1312) O3 NC 0.97 NC 0.63 XP 498788 LYEK (SEQID NO: 1313)
FIG. 5VV U.S. Patent Aug. 18, 2009 Sheet 53 of 97 US 7,575,876 B2
Protein Name IPI # Peptide Sequences iTRAQ Ratios D L B RSTYPLADSTER (SEQ ID NO:1314) SVSNLNYOR (SEQ ID NO: 1315) VI. Proteins unique to DLB and identified by single peptide Neuronal Activities/Signal Transduction Activating receptor pilrbeta IPIO0186781 VELDTR (SEQID NO: 1316) U. 9 7 () 7 2 1. 6 2 Hypothetical protein FLJ13782 IPIO0016576 ASDSQEDQEK (SEQ ID NO: 1317) 0. 8 5 0. 9 5 0. 6 5 Metabotropic glutamate receptor 3 IPI00478.165 ELIAAASR (SEQ ID NO:1318) O. 8 6 O 5 1. 2 4 3. t Orecursor PREDICTED: ETPPLEDLAANQSEDPR (SEQ ID KIAA 1337 protein IPIO0002283 NO: 1319) O 9 7 O 4. . tf PREDICTED: similar to 28 kDa heat- and acid-stable phosphoprotein IPIO0376589 ADLAQLAIIR (SEQID NO:1320) 0.84 NC 1.53 0.62 (PDGF-associated protein) Rho-GTPase activating protein 10 IPIO01693.07 GSWGSGK (SEQID NO:1321) 5. 9 9 Splice isoform 2 of O1 4489 IDTIAADESFTELDLGDR (SEQ ID NNN ephrin type-A NO:1322) 0.88 NC 0.86 NC 0.51 receptor 5 precursor CCCCC Splice isoform 2 of metabotropic glutamate receptor 8 precursor Cell Structure/Motility/Transport/Traffic Hypothetical protein FLJ32363 Poosazza LTQTSPR (SEQID NO:1324) 109 NC Laminingamma-l NTIEETGNLAEQAR (SEQID chain precursor IPIO0298281 NO:1325) 0.93 NC 37 to 6. MGAT3 protein IPIO0020406 VDLVLPEDTTEYFVR (SEQ ID NO:1326) Nconcoso SH3-domain GRB2 like 1 IPIOOO1969 AVTEVLAR (SEQID NO: 1327) Splice isoform 1 of transcription factor IPIO0013929. AADGSLDTQPK (SEQ ID NO: 1328) E2-alpha Splice isoform 2 of sodium/potassium/cal cium exchanger 2 IPIO0218809 VAQGYHQR (SEQID NO: 1329) precursor
FIG. 5WW U.S. Patent Aug. 18, 2009 Sheet 54 of 97 US 7,575,876 B2
Protein Name Peptide Sequences iTRAO Ratios D L B Metabolism | | | | | DNA-directed RNA polymerase I largest IPI00031960 SITNPR (SEQID NO:1330) 0.59 0.89 NC 1.32 t subunit EP"IP100181556 SLAAAFPR (SEQID No.1331) 0.89 NC 0.56 MGAT3 protein IPI00020406 Seyfer EYFVR (SEQ ID NC 102 Ncogs Neuroendocrine convertase 2 IPIO0029131 EELEEELDEAVER (SEQID NO:1333) NC 1.11 NC 1.65 OCCuSOt SCMH1 protein IPIO01871 10 TLNGAEMAPIR (SEQ ID NO:1334) 1.02 NC 0.64 Selenoprotein P Tecursor IPIO0029061 LPTDSELAPR (SEQID NO: 1335) NC NC 1.7 t Splice isoform 2 of glutaryl-CoA dehydrogenase, IPIO0218112 NOLIQK (SEQID NO:1336) NC mitochondrial OrecurSO Splice isoform 3 of reelin precursor IPIO0298066 ITIPLPNAALTR (SEQID NO:1337) 09 Ncogs. Extracellular Matrix/Cell Adhesion Propontin IPIO0292130 GATTTFSAVER (SEQ ID NO: 1338) | 1.02 NC 1.11 NC 0.66
kaatsopoten posio.A:NO:1339) Kiwi (SEQIP is is coast. AAQALNR (SEQ ID NO:1340) 1 NC 0.93 NC 0.44 ependymin related IPI00259102 AGGSHSDPGR (SEQID NO:1341) 1.14 NC 0.57 | | | 1.55 Mammalianprotein 1 roo 1.13 NC 1.56 ft ponsoom poigos NO:1343)Sigvav (SEQIP incosancism Splice isoform 2 of
collagen alpha 2(VI) |IPI00220613 GDPGDAGPR (SEQID NO:1344) 0.85 NC 0.93 NC 0.55 chain precursor Immunity/Defense Hypothetical protein IPI00384931 ADSSPVK (SEQ ID NO:1345) 1.03 NC 104 f Unknown | | | | | E.234L14? IPIOOl65044 ASLQHGQAAEK (SEQ ID NO: 1346) 0.42 l HGS RE408 IPIO0290826 GLAEAAGPR (SEQID NO: 1347) 0.55 HRPE773 IPIO0060800 STEPYDHEIGLR (SEQID 0.99 incoaa. 1.78 in
FIG. 5XX U.S. Patent Aug. 18, 2009 Sheet 55 of 97 US 7,575,876 B2
Protein Name Peptide Sequences iTRAQ Ratios
AD PD DLB
Hypothetical protein
FLJ10650 IPIO0018805 TLQSTPR (SEQID NO:1349)
KARCA1 protein IPI00168703 yoWDGSRR (SEQID is smooncos, Kelch/ankyrin repeat containing cyclin Al IPI00449308 WLCYYGGWDGSRR (SEQID 5.65 NC 0.76 interacting protein NO:1351) PREDICTED: similar to adrenoleukodystroph IPIO0397198 GFLSQRLFAR (SEQ ID NO: 1352) 1.17 NC 1.17 NC 1.56 protein (ALDP) protein coor is pions. SPSF" oncos Ncos
F.G. 5YY U.S. Patent Aug. 18, 2009 Sheet 56 of 97 US 7,575,876 B2
11 kDa protein IPIOO479928 12 kDa protein IPIOO4771.83
132 kDa protein IPIOO412845 132 kDa protein IPIOO477893 154 kDa protein PIOO472011 16 kDa protein IPIOO328348 19 kDa protein IPIOO18134 2,3-bisphosphoglycerate mutase IPIOO2 15979
IPIOO.477336 IPIOO479531
29 kDa protein IPIO080776 2'-phosphodiesterase
38 kDa protein
39S ribosomal protein L9, mitochondrial precursor IPIO03074.09 42 kDa protein 44 kDa protein IPIOO479267
45 kDa calcium-binding protein precursor 45 kDa protein
IPIOO479902 59 kDa protein IPIOO479340 59 kDa protein 61 kDa protein IPIO0334408 65 kDa protein 72 kDa type IV collagenase precursor
IPIOO413996
IPIOO414205 9 kDa protein Actin, alpha skeletal muscle IPIOOO21428 Actin, cytoplasmic 1 Actin, cytoplasmic 2
Adseverin IPIO0002606 AF5q31 protein IPOOOO4344
F.G. 6A U.S. Patent Aug. 18, 2009 Sheet 57 Of 97 US 7,575,876 B2
IPI Address Afamin precursor Agrin ALB protein
ALDOC protein IPIOO418262 Alpha 2,6-sialyltransferase IPIOO479942 Alph A3 type VI collagen isoform 3 precursor IPIOOO729.17 Alpha-l3 IPIO0061448 Alpha-l-acid glycoprotein 2 precursor IPIO0020091 Alpha-1-anti in precursor IPIOO305457 Alpha-1B-glycoprotein precursor IPIO0022.895 Alpha-2-antiplasmin precursor IPIO0029863 Alpha-2-glycoprotein l, zinc IPIOO166729
Alpha-2-HS-glycoprotein precursor Alpha-2-macroglobulin precursor Alpha-2-macroglobulin receptor-associated protein precursor AMBP protein precursor IPIO0022.426 Amyloid-like protein 1 precursor IPIO0020012 Angiotensinogen precursor
APOBEC-1 stimulating protein - IPIO02994.99 Apollipoprotein A-I precursor IPIO0021841 Apollipoprotein A-II precursor PIO0021854 Apollipoprotein A-IV precursor IPIO0304273 Apollipoprotein C-I precursor IPIO002 1855 Apollipoprotein C-II precursor IPIO0021856 Apollipoprotein C-III precursor IPIO0021857 Apolipoprotein D precursor IPIO00066.62 IPIO0021842 Apollipoprotein M IPIO0030739 Associated molecule with the SH3 domain of STAM IPIOO290975 ATP-binding cassette, sub-family A member 8 IPIOO4792.96 ATP-binding cassette, sub-family A, member 2 isoform b IPIOO414303 Baculoviral IAP repeat-containing protein l IPIO0011547 Basement membrane-specific heparan sulfate proteoglycan core protein DeCuISO IPIOOO24284 Beta galactosyltransferase IPIOO184094 Beta-2-glycoprotein I precursor IPIOO298828 Beta-2-microglobulin precursor IPIOOOO4656
FIG. 6B U.S. Patent Aug. 18, 2009 Sheet 58 Of 97 US 7,575,876 B2
Beta-galactosidase Binding lectin precursor Beta-globin gene from a thalassemia patient, complete cas Betaglycan IPIO0304865
Bone morphogenetic protein 3b precursor
Bone-derived growth factor IPIO0299.024 Brain abundant, membrane attached signal protein 1 Brain immunoglobulin receptor precursor IPIOO166048 Brain-derived neurotrophic factor BDNFl IPIO0336003 IPIO0384734 Clorf16 protein IPIOO448672 C4B1 IPIOO418163
Cadherin EGF LAG seven-pass G-type receptor 2 precursor IPIOOO15346 Cadherin-13 precursor IPIO0024046 Calcium binding protein IPIOO384644 Calmodulin IPIO0075248 CAP-binding protein complex interacting protein 1 isoform a Carboxypeptidase E precursor IPIO0031121 Cathepsin B precursor IPIO0295741 Cathepsin D precursor IPIO001 1229 CD59 glycoprotein precursor IPIO001. 1302 CD99L2 protein IPIOO434755 Cell growth regulator with EF hand domain l IPIO0337548
IPIOOOO8584 Ceruloplasmin precursor IPIO001 7601 Chitinase-3 like protein l precursor IPIO0002147
Cholecystokinins precursor IPIO0026174 Chordin-like 1 IPIOO150751 Chromogranin A IPIOO419463 Chromogranin A precursor IPIO0290315
Ciliary dynein heavy chain 9 IPIOO302453 Ciliary rootlet coiled-coil, rootletin IPIOO456492 Clusterin isoform 1 IPIOO40O826 Clusterin precursor IPIO029.1262 Coagulation factor V IPIOO41931 Cocaine- and amphetamine-regulated transcript protein precursor IPIO0002925 Cochlin precursor IPIO001.2386
F.G. 6C U.S. Patent Aug. 18, 2009 Sheet 59 of 97 US 7,575,876 B2
Protein Name IP Address Collagen alpha l(I) chain precursor IPIOO297646 Collagen alpha 1 (VI) chain precursor IPF0029, 136 Collagen alpha 2CD chain precursor IPIO0304,962 Collagen alpha 2CV) chain precursor PIOO293881 Complement Clr subcomponent precursor IPIO0296.65 Complement Cls subcomponent precursor IPIOOO17696 Complement C2 precursor IPIOO3O3963 Complement C3 precursor IPIOO164623 Complement C4 precursor IPIOOO32258 Complement C5 precursor IPIOOO32291 Complement Component 4B proprotein IPIOO453459 Complement component C6 precursor IPIOOOO9920 Complement component C7 precursor IPIOO296608 Complement component C8 beta chain precursor IPIOO294395 Complement component C8 gamma chain precursor PIOOO11261 Complement component C9 precursor IPIOOO22395 Complement factor I precursor IPIOO29 1867 Contactin 2 precursor IPIOOO24966 COP9 signalosome complex subunit 4 IPIOO171844 Corticosteroid-binding globulin precursor IPIOOO27482 C-type matriuretic peptide precursor IPIOOO12075 Cyclophilin PIOO419585 Cystatin C precursor IPIOOO32293 Cytochrome P450 27, mitochondrial precursor IPIO0025307 Cytokeratin type II IPIOOOO5859 Cytosolic malate dehydrogenase IPIO029 1005 Dl dopamine receptor-interacting protein calcyon IPIOOO24587 DA14 HS. IPIOO478414 DEAD (Asp-Glu-Ala-Asp) box polypeptide 51 IPIO0217541 Death-associated protein kinase 1 PIO0021250 Dickkopf related protein-3 precursor IPIO0002714 Dihydropyridine-sensitive L-type, calcium channel alpha-2/delta subunits precursor IPIOO479514 Divalent cation tolerant protein CUTA IPOOO34319
Endothelin B receptor-like protein-2 precursor
FIG. 6D U.S. Patent Aug. 18, 2009 Sheet 60 of 97 US 7,575,876 B2
Protein Name IP Address
Exostosin-like 2 Extracellular matrix protein 1
Extracellular superoxide dismutase Cu-Zn) precursor Far upstream element binding protein 2
Fibrinogen beta chain precursor FK506-binding protein 1A FLJ00120 IPIO0291811 FLJ00268 IPIOO418544 FLJ00271 protein IPIO0386204 FLJ00412 protein IPIO03296.68 FLJ35220 protein IPIO0384.512 Follistatin-like 4 IPIOO477747 Full-length cDNA 5-PRIME end of clone CSODM009YC13 of Fetal liver of Homo sapiens IPIO0328609 Full-length cDNA clone CSODC025YL05 of Neuroblastoma of Homo sapiens IPOO16525 Full-length cDNA clone CSODIO28YM15 of Placenta of Homo sapiens rosara IPIOO384174 Full-length cDNA clone CSODIO85YI08 of Placenta of Homo sapiens rose IPOO382428 Full-length cDNA clone CSODN001YP04 of Adult brain of Homo poss sapiens IPIOO383975 Furin precursor G protein coupled receptor 158 Galectin-3 binding protein precursor IPIOOO23673 Ganglioside GM2 activator precursor GARS protein GBP protein isoform a
FIG. 6E U.S. Patent Aug. 18, 2009 Sheet 61 of 97 US 7,575,876 B2
Protein Name IP Address GM2 activator protein IPIOO418376 Golgi autoantigen, golgin subfamily B member 1 IPIO0004671 Golgi phosphoprotein 2 IPIOO 171411 Golgin-67 isoform c PIO0377 137 Growth/differentiation factor ll precursor
Haptoglobin precursor
Hemoglobin gamma-G IPIOO464992
Heparin-binding EGF-like growth factor precursor HERC2 protein IPIO0005826
HLA class I histocompatibility antigen, alpha chain H precursor IPIO0004672 HLA class I histocompatibility antigen, B-27 alpha chain precursor HLA class I histocompatibility antigen, B-35 alpha chain precursor HLA class I histocompatibility antigen, B-54 alpha chain precursor
HLA class I histocompatibility antigen, Cw- 15 alpha chain precursor HLA class I histocompatibility antigen, Cw-2 alpha chain precursor HLA class I histocompatibility antigen, Cw-7alpha chain precursor IPIOO 144014 HLA class I histocompatibility antigen, E alpha chain precursor IPIO00 10362 IPIOO472612 Hook homolog 3 IPIO0031768 HSAJ 1454 IPIOO419590 HU-K4 IPIOO478097 Hypothetical protein IPIOO4394.46 Hypothetical protein IPIO0026195 Hypothetical protein IPIOO154742 Hypothetical protein IPIOO165652 PIO0384355 Hypothetical protein IPIO0385332 Hvpothetical protein IPIO0386158
IPIO0395435 IPIOO419424 IPIOO430820
FIG. 6F U.S. Patent Aug. 18, 2009 Sheet 62 of 97 US 7,575,876 B2
IP. Address IPIOO430839 IPIOO430842 Hypothetical protein Hypothetical protein IPIOO440577
Hypothetical protein Hypothetical protein Hypothetical protein Hypothetical protein Hypothetical protein IPIOO473141 Hypothetical protein IPIOO 170503 Hypothetical protein DKFZp451 B1418 IPIOO401676 Hypothetical protein DKFZp566O224 IPIO0383815 Hypothetical protein DKFZp686B0286 IPOO465248 Hypothetical protein DKFZp686C02220 IPIOO423461 Hypothetical protein DKFZp686C15213 IPIOO42605 Hypothetical protein DKFZp686E04229 IPIOO426062 Hypothetical protein DKFZp686I15212 IPIOO418153 Hypothetical protein DKFZp686.J1375 IPIO0375843 Hypothetical protein DKFZp686Kll 107 IPIOO464973 Hypothetical protein DKFZp686L19235 IPIOO426056 Hypothetical protein DKFZp686P15220 IPIOO423445 Hypothetical protein DKFZp761H2024 IPIO0185662 Hypothetical protein DKFZp761O0610 IPIOO328584 Hypothetical protein FLJ14473 IPO0386879 Hypothetical protein FLJ16025 IPIOO446856 Hypothetical protein FLJ16420 IPIOO442150
Hypothetical protein FLJ20421 Hypothetical protein FLJ23121 Hypothetical protein FLJ25359 IPIO0307317 Hypothetical protein FLJ25530 IPIO0167215 Hypothetical protein FLJ31726 IPIO0043516 Hypothetical protein FLJ33516 IPIO0383970 Hypothetical protein FLJ33620 IPIO02 16853 Hypothetical protein FLJ33674 IPIO0301 019 Hypothetical protein FLJ35588 IPIO0300S64
FIG. 6G U.S. Patent Aug. 18, 2009 Sheet 63 of 97 US 7,575,876 B2
Protein Name Hypothetical protein FLJ35635 IPIOO385748
Hypothetical protein PSEC0072 Hypothetical protein SGCE Hypothetical protein WUGSCH NH0436C12.1 Ig heavy chain V-I region HG3 precursor IPIO0217045
Ig kappa chain V-I region AG IPOO387022
Ig kappa chain V-I region Ni Ig kappa chain V-I region OU Ig kappa chain V-III region B6 IPIO0387113 Ig kappa chain V-III region WOL IPIOO3871 8
IGHG 1 protein IPIOO448938
Insulin-like growth factor binding protein 5 precursor IPIO0029236 Insulin-like growth factor binding protein 6 precursor IPIO0029235 Insulin-like growth factor binding protein 7 precursor IPIO0016915 Insulin-like growth factor binding protein complex acid labile chain OeCuSO IPIO0020996 Integral membrane protein 2B PIOO477987 Inter-alpha-trypsin inhibitor heavy chain H1 precursor IPIO02925.30
FIG. 6H U.S. Patent Aug. 18, 2009 Sheet 64 of 97 US 7,575,876 B2
IP. Address
Kappa 1 light chain variable region IPIO0382577 Keratin l IPIOO220327 Keratin 10 Keratin 10 IPIOO383111
Keratin 16 IPIOO2 17963 Keratin lb IPIOO376379 Keratin 6 irS4 IPIOO479579 Keratin 6C IPOO479403
Keratin 9 IPIOOO19359 Keratin, type I cytoskeletal 10 IPIOOOO9865 Keratin, type II cuticular HBl IPIO0182654 Keratin, type II cuticular HB4 IPIO0300052 Keratin, type II cytoskeletal 2 epidermal IPIO002 1304 Keratin, type II cytoskeletal 3 IPIOO2.90857 Keratin, type II cytoskeletal 4 IPIOO290.078 Keratin, type II cytoskeletal 5 IPIO0009867 Keratin, type II cytoskeletal 6D IPIO0386438 Keratin, type II cytoskeletal 6F IPIOO2963.50 KLAA0170 protein IPIOO29 1929 KIAA0284 protein IPIOO180625 KIAA0319 protein IPIO0006524 KIAA0387 protein IPIO0024289 KIAA0523 protein IPIOO305349 KIAA0584 protein IPIOO413264 KIAA0644 protein IPIO0006556 KLAA0661 protein IPIOO162563 KIAA0792 protein IPIO0006006 KLAA1009 protein IPIO0007122 KIAA 1061 protein IPIO0298956 KIAA 1291 protein IPIOO413206 KIAA 1318 protein IPIO0002353 KIAA 1373 protein IPIO0002208 KIAA 1417 protein IPIOO165979 KIAA1458 protein IPIO0020601 KIAA1503 protein IPIOO292777
FIG. 6 U.S. Patent Aug. 18, 2009 Sheet 65 of 97 US 7,575,876 B2
KIAA1529 protein KIAA1838 protein KIAA1877 protein IPIO0064125
KRT8 protein Kunitz-type protease inhibitor 2 precursor IPIO0011662
IPIO0020665 Latent transforming growth factor beta binding protein 2 Latent transforming growth factor-beta binding protein 4 IPIO0395.783 Latent transforming growth factor-beta-binding protein 2 precursor IPIOO292 150 IPIO0064742 Leucine-rich alpha-2-glycoprotein precursor IPIO00224.17 L-FILIP IPIOO297210 IPIOOO13303 IPIOO186736 IPIO0329124 IPIOO409640 IPIO0065458 IPIO0020986 Ly-6/neurotoxin-like protein 1 precursor IPIO0289058 Lymphocyte antig OCCuSO IPIO001.4964 Lysosomal alpha-glucosidase precursor IPIO029.3088 IPIOOO19038 IPIOO291641 IPIO0032288 IPIO002871.4 IPIO0027166 MHC class I antigen precursor PIOO478438 MIC2L1 isoform E3'-E4'-E3-E4 IPIOO152491 Mimecan precursor IPIO0025465 MOG protein IPIO0333125 Monocyte differentiation antigen CD14 precursor IPIO0029260 Multiple coagulation factor deficiency protein 2 precursor IPOO328680 Multiple EGF-like-domain protein 4 IPIOOO273 10 Multiple PDZ domain protein IPIOO163612 Myelin-associated glycoprotein precursor IPIO0026237 Myosin-reactive immunoglobulin heavy chain variable region IPIO0007893 Myosin-reactive immunoglobulin heavy chain variable region IPIO0384391
FIG. 6J U.S. Patent Aug. 18, 2009 Sheet 66 of 97 US 7,575,876 B2
-reactive immunoglobulin heavy chain variable region Myosin-reactive immunog chain variable region IPIO0384406
Myristoylated alanine-rich protein kinase C substrate N-acetylgalactosamine-4-O-sulfotransferase
Nebulin IPIO03 03335
Neural cell adhesion molecule 1, 140 kDa isoform precursor IPIOO435020 Neural cell adhesion molecule 2 IPIOO478109 Neural-cadherin precursor IPIO0290085 Neurexin 1-alpha precursor IPIOO442299 Neurexophilin 1 precursor IPIO0048230 Neurexophilin 4 IPIO0376343 Neuroblastoma Suppressor of tumorigenicity 1 precursor IPIOOO13299 Neuroblastoma-amplified protein IPIO0333.913 Neurocan core protein precursor PIOO 159927 Neuroendocrine convertase 1 precursor IPIO0301961 Neurofascin isoform 2 IPIOO477942 Neurofilament triplet H protein IPIO0021751 Neuroligin 2 precursor IPIOO 176424 IPIOO478767 Neuron specific protein family member 1 IPIO0002334 Neuronal pentraxin I precursor IPIOO220,562 Neuronal pentraxin receptor IPIO003 1289 IPIO0334238 IPIOO1641.59 Neuropeptide Y precursor IPIO000 1506 Neurosecretory protein VGF precursor IPIO028950
Nidogen-2 precursor IPIO0028908 Nociceptin precursor IPIO0013701 Nogo receptor-like 3 IPIO0328746 Notch h homologh l 2 IPIOO480098 NOV protein homolog precursor IPIOOOl 1140
FIG. 6K U.S. Patent Aug. 18, 2009 Sheet 67 of 97 US 7,575,876 B2
P60 IPI00179473
PBP family protein precursor PePtidylglycine alPha-amidating monooxygenase isoforma, PreProProtein Pericentrin 2 IPIOO412869 Periplakin IPIO0298057 PF6 IPIOO 174345 Phosphatidylcholine-sterol acyltransferase precursor IPIO0022331
Platelet-derived growth factor beta isoform 2, PreProProtein Plectin 10 IPIO0398778 Plectin 2 IPIO0398775 Plectin 3 IPIOO420096 Plectin 6 IPIOO186711 Plectin 8 IPIO0398777 PLXDC2 protein IPIO0073777 Potassium/sodium hyperpolarization-activated cyclic nucleotide-gated channel 1 IPIO0031506 Potassium/sodium hyperpolarization-activated cyclic nucleotide-gated
channel 3 IPIOO163724 Potassium/sodium hyperpolarization-activated cyclic nucleotide-gated channel 4 IPIO00231.64 PREDICTED: dynein, cytoplasmic, heavy polypeptide 2 IPIOO171494 PREDICTED: FLJ46675 protein IPIOO16539 PREDICTED: G2 protein IPIOO176482
FIG. 6L U.S. Patent Aug. 18, 2009 Sheet 68 of 97 US 7,575,876 B2
PREDICTED: hemicentin-2
PREDICTED: hypothetical protein XP-373647 PREDICTED: hypothetical protein XP 373957 PREDICTED: hypothetical protein XP 498788 PREDICTED: KIAA 1076 protein
PREDICTED: leucine rich repeat containing 4B PREDICTED: lunatic fringe homolog PREDICTED: MAX dimerization protein 5 PREDICTED: similar to ataxin-1 ubiquitin-like interacting PREDICTED: similar to bA92K2.2 (similar to ubiquitin) PREDICTED: similar to Beta-1,3-N-acetylglucosaminyltransferase Poso lunatic fringe (O-fucosy IPIOO454960 PREDICTED: similar to Centromeric protein E (CENP-E protein) PREDICTED: Similar to FKSG30 IPIOO455552 PREDICTED: similar to hypothetical protein IPIOO 66.622 PREDICTED: similar to Keratin, type I cytoskeletal 18 (Cytokeratin 18) (K18) (CK 18) IPIOO455689 PREDICTED: Similar to KIAA1501 protein EPIO0399.193 PREDICTED: similar to KIAA1501 protein IPIOO455296 PREDICTED: similar to KIAA1693 protein IPIOO455450 PREDICTED: similar to Phosphatidylethanolamine-binding protein (PEBP) (Prostatic bindin IPIOO454722 PREDICTED: Similar to POTE2A IPIOO455547 PREDICTED: similar to pregnancy specific beta-l-glycoprotein 7 IPIOO455395 PREDICTED: similar to ribosomal protein L7 IPIO00 18680 PREDICTED: similar to ribosomal protein L7 PREDICTED: similar to ribosomal protein S27a IPIO0398132
PREDICTED: similar to RIKEN cDNA 4732495G21 gene PREDICTED: Similar to RIKEN cDNA 493.0583C14
PREDICTED: Similar to tripartite motif-containing IPIOO455440 PREDICTED: similar to Zgc:66168 protein Prepro-alpha2(I) collagen precursor
Prion protein Probable endonuclease KIAA0830 precursor
FIG. 6M U.S. Patent Aug. 18, 2009 Sheet 69 of 97 US 7,575,876 B2
Protein tyrosine Phosphatase, non-receptor type substrate 1 Precursor IPIO0332887
Protein tyrosine Phosphatase, receptor type, D isoform 2 Precursor IPIO0375547 phatase, receptor type, D isoform 3 Precursor IPIOO375548 Protein tyrosine Phosphatase, receptor type, N Polypeptide 2 isoform 2 Precursor IPIOO472249 Protein tyrosine Phosphatase, receptor type, sigma isoform 3 Precursor IPIOO293275
Quiescin
Ras GTPase-activating protein 2 IPIOOO15811 Ras GTPase-activating-like protein IQGAP1 IPIO0009342 Receptor-type tyrosine-protein phosphatase gamma precursor IPIO001 1651 Receptor-type tyrosine-protein phosphatase-like N precursor PIO0004440 Reticulocalbin 2 precursor IPIO0029628 Reticulon 4, isoform D IPIO0335276
FIG. 6N U.S. Patent Aug. 18, 2009 Sheet 70 of 97 US 7,575,876 B2
Protein Name IPI Address Secreted phoSphoprotein l (osteopontin, bone sialoprotein I, early T lymphocyte activat IPIO0306339 Secretogranin I precursor Secretogranin II precursor Secretogranin III precursor Selenium binding protein 1 Selenium-binding protein 1 Selenoprotein M precursor Semaphorin 7A precursor IPIOOO25257 Semaphorin sem2 Serine protease inhibitor Kazal-type 5 precursor IPIOO478816 Serine/threonine protein phosphatase 2B catalytic subunit, alpha isoform IPIO017945 protein kinase PLK2 Serotransferrin precursor SERPINC1 protein IPIOO16542 SERPIND1 protein IPIOO292950 Serum albumin precursor PIO0022434 Serum amyloid A-4 protein precursor IPIO00 19399 Seven transmembrane helix receptor IPIO02971.88 SH3 and multiple ankyrin repeat domains 2 isoform 1 IPIOO220490 Similar to peptide N-glycanase homolog IPIOO165496 SIN3B long isoform IPIOO464980 Single chain Fv IPIOO007899 Single-chain Fv IPIOO470653 SNC73 protein Sodium channel beta-3 subunit precursor IPIO0020747 Somatostatin precursor IPIO0000130 Sortilin l, preproprotein IPIO0383,591 Sortilin-related receptor precursor IPIO0022608
SPARC-like 1 IPIOO3842.93 SPARC-like protein 1 precursor Spectrin beta chain, brain 4 IPIO0219168
F.G. 6O U.S. Patent Aug. 18, 2009 Sheet 71 Of 97 US 7,575,876 B2
Protein Nane IPI Address Splice Isoform 1 Of Calcyclin-binding protein PIO0395627 Splice Isoform 1 Of Cannabinoid receptor 1 IPIO0009396 IPIOO451624 IPIOO305064 IPIOOO1959 Splice Isoform 1 Of Complement factor H precursor IPIO0029739 Splice Isoform 1 Of Connective tissue growth factor precursor PIO0020977 Splice Isoform 1 Of Contactin l precursor IPIO0029751 Splice Isoform 1 Of EGF-containing fibulin-like extracellular matrix protein l precursor IPIO002.9658 Splice Isoform 1 Of Engulfment and cell motility protein 1 IPIO0219532 Splice Isoform 1 Of Fibrinogen alpha/alpha-E chain precursor IPIO002 1885 Splice Isoform 1 Of Fibrinogen gamma chain precursor IPIO002 1891 olice Isoform 1 Of Fibronectin precursor IPIO0022418 Splice Isoform 1 Of Fibulin-1 precursor IPIO0296534 Splice Isoform 1 Of GDNF family receptor alpha 2 precursor IPIO001 1732 Splice Isoform 1 Of HLA class I histocompatibility antigen, Cw-16 alpha chain precursor IPIOO4.72711 Splice Isoform 1 Of Insulin-like growth factor II precursor IPIO0001611 Splice Isoform 1 Of Inter-alpha-trypsin inhibitor heavy chain H4 OcCLSO PIOO294193 Splice Isoform 1 Of Latrophilin 1 precursor IPIOO 183445 Splice Isoform 1 Of Lysosome-associated membrane glycoprotein 2 OecurSO IPIOO009030 Splice Isoform 1 Of Macrophage colony stimulating factor-1 precursor IPIOOO15881 Splice Isoform 1 Of Matrix metalloproteinase-17 precursor IPIOO008533 Splice Isoform 1 Of Neural cell adhesion molecule 1, 120 kDa isoform precursor IPOO4. 1478 Splice Isoform 1 Of Neurexin 2-alpha precursor IPIO0007921 Splice Isoform 1 Of Neuroendocrine protein 7B2 precursor IPIOO008.944 Splice Isoform l Of Pantothenate kinase 2, mitochondrial precursor IPIO0171 176 Splice Isoform 1 Of Phosphatidylinositol 3,4,5-trisphosphatedependent Rac exchanger 1 IPIO0295.252 Splice Isoform 1 Of Phospholipid transfer protein precursor IPIO0022733 Splice Isoform 1 Of Platelet-derived growth factor, A chain precursor IPIO002 1833 Splice Isoform 1 Of Plectin l IPIO0014898 Splice Isoform 1 Of Proactivator polypeptide precursor IPIOOO12503 Splice Isoform 1 Of Receptor-type tyrosine-protein phosphatase N2 OreCuSOt IPIO0334666
F.G. 6P U.S. Patent Aug. 18, 2009 Sheet 72 Of 97 US 7,575,876 B2
IPI Address Splice Isoform 1 Of Receptor-type tyrosine-protein phosphatase Zeta OreCuSO IPIO029 1099 Splice Isoform 1 Of Reticulon 1 IPIO0003971 Splice Isoform 1 Of Reticulon 4 IPIOOO2 1766 Splice Isoform 1 Of Serine/threonine-protein kinase Haspin PIOO397836
Splice Isoform l Of Serologically defined colon cancer antigen l IPIO03.01618 Splice Isoform 1 Of Stromal cell-derived factor 1 precursor IPIOO413781 Splice Isoform 1 Of SWI/SNF-related, matrix associated, actin dependent regulator of ch IPIOO2201 19 Splice Isoform 1 Of Testican-3 precursor IPIOO478890 Splice Isoform 2 Of Acyl-CoA-binding protein IPIOO218836 Splice Isoform 2 Of Alpha-mannosidase IIx IPIOO220303 Splice Isoform 2 Of Amyloid-like protein 2 precursor IPIO0220977 Splice Isoform 2 Of Apoptotic protease activating factor 1 IPIOO217460 Splice Isoform 2 Of Collagen alpha 1 (XVIII) chain precursor IPIOO414694 Splice Isoform 2 Of Connective tissue growth factor precursor IPIOO220647 Splice Isoform 2 Of Ectonucleotide pyrophosphatase/phosphodiesterase pool Splice Isoform 2 Of Far upstream element binding protein 1
Splice Isoform 2 Of Latrophilin 1 precursor
Splice Isoform 2 Of Neuroligin 3 precursor Splice Isoform 2 Of Neuronal cell adhesion molecule precursor Splice Isoform 2 Of Osteopontin precursor Splice Isoform 2 Of Proactivator polypeptide precursor IPIO0219824 Splice Isoform 2 Of Receptor-type tyrosine-protein phosphatase N2 DeCISO IPIO0334667 Splice Isoform 2 Of Sex hormone-binding g OreCuSO IPIOO295.83 Splice Isoform 2 Of Signal-regulatory protein beta-2 precursor IPIO0218600 Splice Isoform 2 Of SPARC related modular calcium-binding protein 1 precurSOr IPIOO412898 Splice Isoform 3 Of A-kinase anchor protein 9 IPIOO220625 Splice Isoform 3 Of Amyloid beta A4 protein precursor PIO0219183
FIG. 6O U.S. Patent Aug. 18, 2009 Sheet 73 of 97 US 7,575,876 B2
Protein Name IP Address Splice Isoform 3 Of Amyloid-like protein 2 precursor IPIOO220978 Splice Isoform 3 Of Bullous pemphigoid antigen 1, isoforms 6/9/10 IPIOO47319 Splice Isoform 3 Of EGF-containing fibulin-like extracellular matrix protein 1 precursor IPIOO220814 Splice Isoform 3 Of Integrin alpha-7 precursor Splice Isoform 3 Of Myelin-oligodendrocyte glycoprotein precursor Splice Isoform 3 Of Neuronal cell adhesion molecule precursor Splice Isoform 3 Of Neurotrimin precursor Splice Isoform 3 Of Peptidyl-glycine alpha-amidating monooxygenase OeCuSO IPIO0219042 Splice Isoform 3 Of Proactivator polypeptide precursor IPIO021982.5 Splice Isoform 3 Of Receptor-type tyrosine-protein phosphatase S porn precursor IPIO0332272 Splice Isoform 3 Of SH3 and multiple ankyrin repeat domains protein 1 poss IPIOO220,165 Splice Isoform 3 Of WAP four-disulfide core domain protein 2 precursor rose IPIOO183629 Splice Isoform 4 Of EGF-containing fibulin-like extracellular matrix protein l precursor IPIOO220815 Splice Isoform 4 Of Fibulin-1 precursor IPIO0296537 Splice Isoform 4 Of Integrin beta-1 precursor IPIO0217562 Splice Isoform 4 Of Nesprin l Splice Isoform 4 Of Nuclear autoantigen Sp-100 Splice Isoform 4 Of Osteopontin precursor IPIO0385896
Splice Isoform 4 Of Seizure 6-like protein precursor Splice Isoform 5 Of Amyloid beta A4 protein precursor
Splice Isoform 6 Of Amyloid beta A4 protein precursor IPIO0219 186 Splice Isoform 7 Of Amyloid beta A4 protein precursor IPIO021987 Splice Isoform 7 Of Myelin-oligodendrocyte glycoprotein precursor IPIO0376382 Splice Isoform 8 Of Amyloid beta A4 protein precursor IPIOO412924 Splice Isoform 8 Of Myelin-oligodendrocyte glycoprotein precursor IPIOO473134 Splice Isoform 9 Of Amvloid beta A4 protein precursor IPIOO412681 Spondin 1 precursor SPUF protein precursor Sulfatase 2 isoform b precursor Superoxide dismutasel, Soluble Synaptotagmin-ll
TA p63 alpha
TBC1 domain family member 10 T-cell activation Rho GTPase activating protein IPIOO166033 Tenascin-R IPIO01 60552
F.G. 6R U.S. Patent Aug. 18, 2009 Sheet 74 of 97 US 7,575,876 B2
IPI Address Testican-1 precursor IPIO0005292 Testican-2 precursor IPIO0006128
Thyroxine-binding globulin precursor IPIO0292.946 Titin IPIOO179357 IPIO0375498 Titin, heart isoform N2-B IPIOO455 173 Toll-like receptor 7 precursor IPIO0009812 Transforming growth factor-beta induced protein IG-H3 precursor IPIO0018219 Transient receptor potential cation channel subfamily M member 7 IPIO0290032 IPIO0022432 Tripartite motif protein 26 Type I inner root sheath specific keratin 25 irs3 IPIO03281 03
Ubiquitin and ribosomal protein L40 precursor IPIOO456429 Ubiquitin and ribosomal protein S27a precUrsor IPIO0179330 Usher syndrome 1 C binding protein 1 IPIO0297.559
Villin 1 IPIOO218852 Vitamin D-binding protein precursor IPOO298853 Vitamin K-dependent protein Sprecursor IPIOO294.004 IPIO0298971 VPS10 domain-containing receptor SorCS2 precursor PIO0044600 VPS10 domain-containing receptor SorCS3 precursor IPIOOO 10381 WAP four-disulfide core domain protein 1 precursor
WUGSC:H DJ0747G 8.3 protein WW domain containing adaptor with coiled-coil IPIOO47.8665
FIG. 6S U.S. Patent Aug. 18, 2009 Sheet 75 Of 97 US 7,575,876 B2
IP. Address WW domain-containing adapter with a colled-coil region, isoform 1 IPIO0010241 Xylosyltransferase I IPIOO183487 Zona pellucida sperm-binding protein 2 precursor PIP000 16870
FIG. 6T U.S. Patent Aug. 18, 2009 Sheet 76 of 97 US 7,575,876 B2
Protein Name IP Address 101 kDa protein IPIOO478742 101 kDa protein IPIOO291.316 IPIOO382841 110 kDa protein IPIOO473056 130 kD Golgi-localized phosphoprotein IPIO0004962 IPIOO478089
28S ribosomal protein Sl8c, mitochondrial precursor IPIOOOO7049 344 kDa protein IPIOO479834 376 kDa protein IPIOO479143
66 kDa protein IPIOO479085 67 kDa protein IPIO0332849
IPIO0217272
FIG. 7A U.S. Patent Aug. 18, 2009 Sheet 77 of 97 US 7,575,876 B2
Alpha 3 type VI collAgen isoform 4 precursor Alpha-1,3-mannosyl-glycoprotein 2-beta-Nacetylglucosaminyltransferase poss IPIO0000138 Alpha-l-acid glycoprotein l precursor Alpha-fetoprotein precursor Alpha-ketoglutarate dehydrogenase complex dihydrolipoyl pool succinyltransferase IPIO0033034
Ankyrin repeat and SOCS box protein 2 IPIO0216028 Ankyrin repeat domain protein 9 IPIO0073421 Antigen MLAA-20 IPIOO4471.78 APG7L protein IPIOO47991 1 Apollipoprotein F precursor IPIOO4801 19 ARHGAP8 protein IPIOO472223 AspArtAte AminotransferAse 1 IPIO0219029 ATP-binding cassette, sub-family A (ABC1), member 1 IPIOO477917 ATP-binding cassette, sub-family A, member 1 PIOO293460 ATP-dependent RNA helicase DDX24 IPIOOOO6987 Atri.Al/embryonic AlkAli myosin light chAin AXL receptor tyrosine kinase, isoform 1 IPIOO296992 BA231F10.1 IPIOO472.399 Bactericidal/permeability-increasing protein-like 3 precursor IPIOO414328 Beta-1,4N-acetylgalactosaminyltransferase IPIO0025473
BMP and activin membrane-bound inhibitor homolog precursor
FIG. 7B U.S. Patent Aug. 18, 2009 Sheet 78 of 97 US 7,575,876 B2
Bromodomain protein CELTIX1 Bullous pemphigoid antigen 1 isoform 1 Bullous pemphigoid antigen l isoform leB precursor ClD protein ClorfA.0 protein C21orf258 protein C9orf86 protein Cadherin 11, type 2, isoform 1 preproprotein Cadherin-22 precursor IPIO000.0436 Calcitonin gene-related peptide I precursor CalCium binding protein Cab45 preCursor
Calmodulin 2 Calreticulin precursor Calsyntenin-3 precursor CANT1 protein Cappuccino protein homolog Cathepsin L precursor Cathepsin Sprecursor
Centrosome-associated protein 350 Cerebellin 3 precursor IPIOO402157 Chemokine (C-X-C motif) ligand 16 IPIO0004946 Chondroitin sulfate proteoglycan 5-III IPI00434467IPIOO434467 Chromosome 10 open reading frame 88 IPIO0296845IPO0296845 Chromosome 6 open reading frame 152 IPIOO3340IPIO0334013 13 Chromosome 9 open reading frame 140 IPIO0328702IPIOO328702 Chronic myelogenous leukemia tumor antigen 66 IPIOO306398IPI00306398 Coagulation factor X precursor IPIO0019576IPIOOO19576 Coagulation factor XII precursor IPIOOO19581IPI0001958 Cohesin subunit SA-l IPIO0025IPIOOO25 158 Coiled-coil domain containing protein 9 IPIOO177642IPI00177642 Collagen alpha 1 IPIO0019090IPIOOO19090 Collagen alpha 1 (III) chain precursor IPIO002IPI00021033 1033
FIG. 7C U.S. Patent Aug. 18, 2009 Sheet 79 of 97 US 7,575,876 B2
Protein Name Collagen alpha l(XV) chain precursor PIOO29544 Complement C1q tumor necrosis factor-related protein 4 precursor Component of oligomeric golgi complex 6 IPIO0398963 CSRV314 LIPI00432626 Cystatin M precursor IPIO0019954 Cytochrome P450 1A1 IPIOO2 18839 Cytokine-like protein C17 precursor EPIO0032876 DEAH (Asp-Glu-Ala-His) box polypeptide 29
Dermatopontin precursor IPIO0292130 Dermcidin precursor IPIO0027547 Dermokine-beta IPIOO454.602 Dipeptidyl peptidase-like protein 2 IPIOO464986 Dipeptidyl-peptidase II precursor IPIO0296141 DJ 1003J2.3.1 DJ 1042K10.2.2 IPOOO23854 DJ1 19A7.3 IPIO0003697 IPIOO181864 DJ68D18.1.2 IPIOO300020 DJ788L20.2 IPIOO478622 DKFZp434L142 protein IPIOO 65044 DKFZP564O243 protein IPIO0042.514 DNA cytosine methyltransferase 3alpha, isoform a IPIO0329216 DNA excision repair protein ERCC-6 DNA-directed RNA polymerase I largest subunit IPIO0031960 IPIO0306229 IPIO0004970 Dyskerin Endothelin 3, isoform 2 preproprotein Ephrin Al isoform b prEcursor Ephrin receptor EphB3
Epsilon globin ErythrocytEmEmbranB protein band 4.2 IPIO0028120 Estrogen receptor binding protein Exostosin-like 3
FIG. 7D U.S. Patent Aug. 18, 2009 Sheet 80 of 97 US 7,575,876 B2
IP Address IPIO02 18834
IPIO0292732 IPIO0396282 IPIOO29 1755 IPIOO41 1980 IPIO029 1731 IPIO0152731 FLJ00332 protein IPIO021681 1 FLJ1 1029 protein IPIOO305822 FLJ34512 protein IPIO0216820 Follistatin-related protein 1 precursor IPIO0029723 IPIO0021176 IPIOO 102670 IPIOO294.650 IPIOO384O16 Full-length cDNA clone CSODM007YO13 of Fetal liver of Homo sapiens IPIO00071.99
IPIO0004367 IPIOOOO1580 IPIOOO15343 IPO0099.521 IPIO0029810 IPIOO479258 Gamma-synuclein IPIOO297714 Gelsolin precursor IPIO0026314 Glutaminyl-peptide cyclotransferase precursor IPIOOOO3919 Glutaredoxin (thiol transferase) IPOO2 19025 Glyceraldehyde-3-phosphate dehydroGenase IPIOO2 1901.8 Glycerol kinase, isoform a PIOO419934 PIOO414717 GolGin-67 isoform a IPIO0016475 IPIOO428657 IPIOO418687 IPIOO2.98476 IPIO0026926 IPIOO470375 IPIOO220362
IPIO0045223 IPIO0332106 Hemoglobin beta EPIO02 1881 6 Hepatocellular carcinoma associated protein TB6 IPIO0293898
FIG. 7E U.S. Patent Aug. 18, 2009 Sheet 81 of 97 US 7,575,876 B2
Protein Name Heterogeneous nuclear ribonucleoprotein L isoform b HEXIM1 protein Hexokinase, type II HGF activator like protein HGS RE408 HIV TAT specific factor 1 IPIO0013788 HSPC009
Husli-like protein Hypothetical protein
Hypothetical protein Hypothetical protein Hypothetical protein Hypothetical protein Hypothetical protein IPIOO419345IPI004 19345 Hypothetical protein IPIOO430806IPI00430806 Hypothetical protein IPIO0470772IPIOO470.772 Hypothetical protein DKFZp434A2017 IPIOO295.380IPI00295380 Hypothetical protein DKFZp434COll IPIOO152946IPI00152946 Hypothetical protein DKFZP434J0113 IPIO0217802IPI00217802 Hypothetical protein DKFZp434K1421 IPI00030274IPIO0030274 Hypothetical protein DKFZp434P097 IPIO001 1232 Hypothetical protein DKFZp434P1219 IPIOO3961IPI00396169 69 Hypothetical protein DKFZp547D2210 IPI00217787IPIOO2 17787 Hypothetical protein DKFZp547N 1615 IPIOO028864IPI00028864 Hypothetical protein DKFZp586K2123 IPIOO41IPI00411596 1596
FIG. 7F U.S. Patent Aug. 18, 2009 Sheet 82 of 97 US 7,575,876 B2
Hypothetical protein DKFZp686L 13193 Hypothetical protein DKFZp686N18114 IPIOO 171323 Hypothetical protein DKFZp686O0186 IPIO0384977 Hypothetical protein DKFZp761D171 PIO0385612 Hypothetical protein DKFZp761F0118
IPIOO395775 IPIOO153050 IPIO0002373 IPIOOOO9673 IPIO0336000 IPIOO303852
IPIO003 0385 PIOO165528 IPIO0304069 IPIO0395.424 IPIOO442338
IPIOO165009 IPIO0329662 IPIOO456642
FIG 7G U.S. Patent Aug. 18, 2009 Sheet 83 of 97 US 7,575,876 B2
IPI Address IPIO0060969 IPIOO67196 IPIO0059639 IPIO0043428 IPIOO374273 IPIOOO65491 IPIO0065349 IPIOO480.036 IPIOO413989 IPIO0171044 IPIOO167575 IPIO0384796 IPIOO167490 IPIOO179405 IPIO0167233 PIOO419164 IPIOO446159 IPIOO44.5546 IPIOO418993 Hypothetical protein FLJ4416l IPIOO445366 Hypothetical protein FLJ44241 PIOO465179 Hypothetical protein FLJ44324 IPIOO445227 IPIOO444823 Hypothetical protein FLJ45264 IPIOO444644 Hypothetical protein FLJ45525 IPIO02995 71 Hypothetical protein FLJ45715 PIOO4442.59 Hypothetical protein FLJ45736 IPIOO444240 Hypothetical protein FLJ46550 IPIOO443682 IPIOO479983 IPIOO443445 IPIOO477479 IPIO0328520 IPIOO181556 Hypothetical protein FLJ90661 IPIOO168352 Hypothetical protein LOC113174 IPIO0304935 Hypothetical protein LOC122618 IPIO0060310 Hypothetical protein LOC90624 IPIO0329321 Hypothetical protein MGC26885 IPIO0216887 Hypothetical protein MGC29784 IPIOO166131
FIG. 7H U.S. Patent Aug. 18, 2009 Sheet 84 of 97 US 7,575,876 B2
Hypothetical protein PIK3CG Hypothetical protein PSEC0200 IPIOO166392 ypothetical protein PSEC0250
Ig heavy chain V-II region SESS precursor Ig heavy chain V-III region BUT Ig heavy chain V-III region GA Ig kappa chain V-I region BAN IPIO0385555 Ig kappa chain V-I region HK102 precursor IPIOO478600 Ig kappa chain V-III region VG precursor IPO.041.9453 Ig kappa chain V-III region VH precursor IPIO00241.38 IPIO0386133 Ig lambda chain V-III region SH IPIO0382436 Ig lambda chain V-IV region Hil IPIOO382440 IL-17RC IPIO0303074 Immunoglobulin-like domain protein MGC33530 precursor IPIOO290411 Import inner membrane translocase subunit TIM44, mitochondrial OCCSO - IPIO0306516 Importin 9 IPIOO1851.46 Inhibin beta A chain precursor IPIOOO28670 Inositol polyphosphate-5-phosphatase F IPIO0383,580 Insulin receptor tyrosine kinase substrate IPIO0179326 Insulinoma-glucagonoma protein 20 splice variant 2 tryp
Intercellular adhesion molecule-5 precursor Interleukin 17 receptor C, isoform 3 Interleukin-1 receptor-associated kinase-like 2 IPIO0304986 Isocitrate dehydrogenase (NAD) subunit alpha, mitochondrial precursor Isocitrate dehydrogenase NADP) cytoplasmic IPIO0027223 KARCA1 protein Kelch/ankyrin repeat containing cyclin A1 interacting protein Keratin b20 IPIOO43 1749
FIG. 7 U.S. Patent Aug. 18, 2009 Sheet 85 of 97 US 7,575,876 B2
IP Address IPIO0307649 IPIO032957 IPIOO005634 KIAA0443 protein IPIO0060549 KIAA0477 protein IPIO0337544 IPIOO479532 IPIO0384636 IPIOO 170935 IPIO0297288 IPIO0008085 KIAA1274 protein IPIOO297212 KIAA 1384 protein IPIOO418195 IPIO0001790 IPIOO177381 IPIO0288939 PIOO1551.99 KIAA 1840 protein IPIOO1 01923 KIAA 1946 IPIOO3961 66 KSS splice variantb IPIOO375393 Laminin alpha-1 chain precursor IPIOO375294 Laminin gamma-1 chain precursor IPIOO298.281 L sites. IPIOO 107831 Latent transforming growth factor beta binding protein l isoform LTBP 1L IPIOO410152 Latent transforming growth factor beta binding protein, isoform lL
IPIOO303112 IPIOO 100947 Line-1 repeat mRNA with 2 open reading frames IPIOO477474 Lipopolysaccharide-binding protein precursor IPIO0032311 IPIO0289271 IPIO0394.856 IPIOO428724 IPIOOO 10471 LTLL9335 IPIOO432.693 Lysosomal-associated multitransmembrane protein IPIOOO13827
FIG. 7J U.S. Patent Aug. 18, 2009 Sheet 86 of 97 US 7,575,876 B2
IP Address MAP-kinase activating death domain-containing protein isoform a IPIOO 107844 IPIO001 7297 IPIOOO 14939 IPIOO178639 IPIOOOOO868 IPIO0003448 IPIOO4781.65 IPIO0032292 IPIO001 6666 IPIO0020406 IPIO0177578 IPIO0012832 IPIOO 104128 IPIOO374770 IPIO00591.84 IPIOO375915 IPIOO295066 IPIOOOO6775 Mitotic kinesin-related protein IPIO0044751 IPIOOOO2968 IPIO0296004 MOP-4 IPIO0023647 Mosaic serine protease IPIOOO12505 Mothers against decapentaplegic homolog 4 IPIO0013404 MSFL2541 IPIO0399.139 Mu-crystallin homolog IPIOOOOO949 Muellerian inhibiting factor precursor IPIOO008577 Multi-functional protein MFP IPIOO479.309 Multiple inositol polyphosphate phosphatase IPIOO293748 Muscle-cadherin precursor IPIO0024048 Muscle-type acylphosphatase 2 IPIO0216461 Myelin associated glycoprotein isoforMb precursor IPIOO3752.53 Myelin P0 protein precursor IPIOO 106596 Myelin protein zero IPIO0334017 Myosin If P002 18638 Myosin-reactive immunoglobulin heavy chain variable region IPIO0384395 Myosin-reactive immunoglobulin kappa chain variable region IPIO0384401 N-acetyltransferase 5 isoform b IPIOO375482 NDST2 protein IPIOO 103042 Nebulin-related anchoring protein IPIOO478974
FIG. 7K U.S. Patent Aug. 18, 2009 Sheet 87 of 97 US 7,575,876 B2
IPI Address Nectin-like protein 3 IPIOO293836 NEFL protein IPIO0237671 IPIOO185362 IPIO0299699 IPIOO42851 1 PIOO293723 IPIOO470625 IPIOOO291.31 Neurogenic locus notch homolog protein 2 precursor IPIO0297655 IPIO0025789 IPIO001 1063 IPIO0005416 IPIOO477060
IPIO0072576 IPIO0386384 IPIO01 07749 IPO0304 189 IPIO0028937 IPIOOO29796 IPIO0020990 IPIO037453 IPIO0337642 IPIO0337350 IPIOO477417 IPIOO154774 IPIO0000144. IPIOO 169348 IPIO0329775 Peptidyl-prolyl cis-trans isomerase C IPIO0024129 IPIO0027350 IPIOOO631.87 IPIOO413674 IPIO0099522 IPIO0172666 IPIOO412264 IPIO0382460 IPIOO419595 IPIO0004573 IPIO00101.74 IPIO0012857
FIG. 7L U.S. Patent Aug. 18, 2009 Sheet 88 of 97 US 7,575,876 B2
IP Address Potassium/sodium hyperpolarization-activated cyclic nucleotide-gated channel 2 IPIO02 18946
PREDICTED: hypothetical protein XP 499305 IPIOO464965 PREDICTED: KIAA0367 protein IPIO0004557
PREDICTED: similar to 28 kDa heat- and acid-stable phosphoprotein (PDGF-associated prot IPIO0376589 PREDICTED: Similar to 40S ribosomal protein S16 IPIO03977O1
PREDICTED: similar to CCR4-NOT transcription complex, subunit 6like IPIOO4552.53 PREDICTED: Similar to CG3047-PA IPIO0376412
FIG.7M U.S. Patent Aug. 18, 2009 Sheet 89 of 97 US 7,575,876 B2
IPI Address PREDICTED: similar to Chloride intracellular channel protein 4 (Intracellular chloride IPIOO455949 PREDICTED: similar to DEAD/H (Asp-Glu-Ala-Asp/His) box polypeptide ll PIO0398943 PREDICTED: similar to Fatty acid-binding protein, epidermal (EFABP) (Psoriasis-associa IPIO0398985 PREDICTED: similar to germ and embryonic stem cell enriched protein STELLA IPIOO402063 PREDICTED: similar to Glutathione S-transferase Mu 5 (GSTM5-5) GST class-Mu 5 IPIOO454.856 PREDICTED: similar to golgin-67 isoform c IPIOO4723.63 PREDICTED: similar to heat shock 10kDa protein 1 (chaperonin 10) IPIOO455469 PREDICTED: similar to Histidine-rich glycoprotein precursor (Histidine proline rich gly IPIOO454879 PREDICTED: similar to hypothetical protein A830023L05 IPIOO455633 PREDICTED: similar to Hypothetical protein BC005730 IPIO0252950 PREDICTED: Similar to matrilin 2 precursor IPIO0145674 PREDICTED: similar to melanoma antigen, family A, 10 IPIOO455972 PREDICTED: similar to pre-mRNA splicing SR protein related (68.2 kD) (rsr-1) IPIOO402573 PREDICTED: similar to Ribosome biogenesis protein BMS1 homolog IPIO0253009 IPIO03981 17 IPIOO455997 IPIO0073442 IPIOO455390
IPIOO457064 IPIO0084684 IPIO0334256 IPIO00321.89 IPIO02 19468 IPIOO235 167 IPIOO220739 IPIOO472584 IPIO0020987 Proline-rich protein 4 precursor IPIO0027.019 Prostate tumor overexpressed gene 1 IPIO0010118
IPIO02 19446 IPIO0297.040 Protein C20orf'98 IPIOOO 7231 Protein FAM38A IPIO000.6093
FIG. 7N U.S. Patent Aug. 18, 2009 Sheet 90 Of 97 US 7,575,876 B2
Protein Name IPI Address Protein PRO1854 IPIO0006005 Protein tyrosine Phosphatase domain containing 1 Protein isoform 1 IPIO0376989 osine phosphatase, non-receptor type 14 IPIOO477830 IPOO41 1680 Protein-tyrosine sulfotransferase l IPIO00301 06 Protocadherin l isoform 2 Precursor IPIOO 176458 Protocadherin Fat 2 precursor IPIO030264 EPIO002959 PSMC3 protein IPIO0018398 PSST739 IPIO0394870 PTPL1-associated RhoGAP IPIO0152011 IPIO0010148 Putative 4 repeat voltage-gated ion channel IPIO0217996 Putative acyl-CoA thioester hydrolase CGI-16 IPIOO220710 Putative alpha-mannosidase Clorf22 IPIO0009.410 Putative secreted ligand IPIO0003834 IPIO0027806 Pyruvate kinase 3 isoform 1 IPIOO47986 QVSK201 IPIOO465325 RAS p21 protein activator 3 IPIO0383401 Ras-related protein Rap-2b IPIO0018364 Receptor-interacting serine/threonine-protein kinase 2 IPIO002 1917
Retbindin RGD, leucine-rich repeat, tropomodulin and proline-rich containing rose protein IPIOO456628 Rho/rac-interacting citron kinase Rho/rac guanine nucleotide exchange factor (GEF) 2 IPIOO412782 Rho-GTPase-activating protein 5 IPIO0013988 Ribonuclease 4 precursor IPIO0029699 Ribosomal protein L39 IPIO0219 162 Ribosomal protein L3 IPIO02.9335 Ribosomal protein L7 IPIO0030179 Ribosome biogenesis protein BMS1 homolog IPIO0006099 RNA-binding protein 5 IPIO0005036 IPIOO4141 17 SAA 1 protein IPIOO452748 Salvador homolog l protein IPIO0301 738 Sarco/endoplasmic reticulum Ca2+-ATPase isoform d IPIO0218442
FIG. 7O U.S. Patent Aug. 18, 2009 Sheet 91. Of 97 US 7,575,876 B2
IPIIP Address IPI00432771IPIOO432771 IPI00187110IPIOO 1871 10 IPIO0029061IPIOOO29061 SEMA3B protein IPIOO44.8569IPI0044.8569 Serine protease HTRA1 precursor IPIO0003176 Serine protease inhibitor, Kazal type, 5 IPIO0299453 Serine/threonine-protein kinase Hl IPI00007810IPIOOOO7810 Seven transmembrane helix receptor IPI00376212IPIOO376212 SH3 domain-binding glutamic acid-rich-like protein IPIOOO253.18IPI00025318 SH3-domain GRB2-like 1 IPIOOOIPI00019169 19169 Sia-alpha-2,3-Gal-beta-1,4-GlcNAc-R:alpha 2.8-sialyltransferase IPIO0026285IPOOO26285 Similar to ecotropic viral integration site 5; Neuroblastoma stage 4S gene IPIOOO60473
SNC66 protein Sodium/potassium-transporting ATPase alpha-2 chain precursor Sortilin precursor IPIO0217882
Splice Isoform 1 Of ADAMTS-2 precursor
Splice Isoform 1 Of Calcium/calmodulin-dependent protein kinase type II pool alpha chain IPIOOO98624
Splice Isoform 1 Of COP9 signalosome complex subunit 1 Splice Isoform 1 Of Cullin homolog 4B Splice Isoform 1 Of Cyclic-AMP-dependent transcription factor ATF-6 beta IPIOO004084
FIG. 7P U.S. Patent Aug. 18, 2009 Sheet 92 of 97 US 7,575,876 B2
Protein Name IPI Address Splice Isoform 1 Of Double-stranded RNA-specific adenosine deaminase IPO0394665 Splice Isoform 1 Of Dynein intermediate chain l, cytosolic IPIOOO22461 Splice Isoform 1 Of Ecto-ADP-ribosyltransferase 3 precursor IPIOOO13682 Splice Isoform 1 Of Endothelin-3 precursor IPIO0025365 Splice Isoform 1 Of ERC protein l IPIO0216719 Splice Isoform 1 Of Gamma-tubulin complex component 6 IPIO0045491 Splice Isoform 1 Of Glutaryl-CoA dehydrogenase, mitochondrial OcCuSO IPIO0024317 PIO0337307 un-- y IPIO003 1545 Splice Isoform 1 Of IQ calmodulin-binding motif containing protein 1 IPIOOO14255 Splice Isoform 1 Of Lysosomal trafficking regulator IPIOOO 17094 Splice Isoform 1 Of Neuroligin, 1 precursor Splice Isoform 1 Of Neuropilin-1 precursor IPIO0299594 Splice Isoform 1 Of Osteopontin precursor IPIO0021000 Splice Isoform 1 Of p130Cas-associated protein IPIOO479643 Splice Isoform 1 Of Partitioning defective-6 homolog alpha IPIO0027217 Splice Isoform 1 Of Pleckstrin homology domain containing family C member 1 IPIO0000856 Splice Isoform 1 Of Polycystic kidney and hepatic disease 1 precursor Splice Isoform 1 Of Protachykinin l precursor Splice Isoform 1 Of Protein C2 lorf70 Splice Isoform 1 Of Putative polypeptide N acetylgalactosaminyltransferase-like protein IPIOO166613 Splice Isoform 1 Of Receptor-type tyrosine-protein phosphatase delta precursor IPIOOO 1642 Splice Isoform l Of Rotavirus 'X' associated non-structural protein IPIO00991.31 protein kinase RIPK4 IPIO002571.4 Splice Isoform 1 Of SET binding factor 1 IPIO0029.446 Splice Isoform 1 Of Short transient receptor potential channel 6 IPIO0031683 Splice Isoform 1 Of Sodium/potassium-transporting ATPase alpha-1 chain OneCuSO PIO0006482
Splice Isoform 1 Of Trans-Golgi network integral membrane protein 2 OCCuSO IPIOOO 12545 Splice Isoform l Of Tubby-like protein 4 IPIOOO24994 Splice Isoform 1 Of Ubiquitin carboxyl-terminal hydrolase 33 IPIO0236901
FIG. 7Q U.S. Patent Aug. 18, 2009 Sheet 93 of 97 US 7,575,876 B2
IP Address Splice Isoform 1. Of Ubiquitin-conjugating enzyme E2 variant 1 IPIOOO19599 plice Isoform 1 Of UPF0338 protein NG5 IPIO0043810 Splice Isoform 1 Of Vacuolar protein sorting IPIOOOO1985 Splice Isoform 1 Of Voltage-dependent N-type calcium channel alphal B subunit IPIO0025477 IPIOO41 0663 IPO0216421
IPIO0216422 IPIOO 1861 14 IPIOOO 12366 IPIOO220791 IPIOO 106911 IPIOO186903 IPIO0216520 IPIOOO 19906 IPIOOO 14021 Splice Isoform 2 Of Bromodomain adjacent to Zinc finger domain protein 1A PIOO383565 Splice Isoform 2 Of Cadherin-1 1 precursor IPIOO293539 Splice Isoform 2 Of Calcium/calmodulin-dependent protein kinase type II alpha chain IPIOO2.15715 Splice Isoform 2 Of Canalicular multispecific organic anion transporter 2 IPIO025 1066 IPIOO41 0682 PIOO220613 IPIOO220701 IPIOO2 18999 IPIOO334426
IPIOO40963
protein l precurso IPIOO220813 Splice Isoform 2 Of Endothelin-3 precursor IPIOO220210 Splice Isoform 2 Of Ephrin type-A receptor 5 precursor IPIO0215945 Splice Isoform 2 Of Glutaryl-CoA dehydrogenase, mitochondrial precursor IPIO0218112 Splice Isoform 2 Of HLA class I histocompatibility antigen, Cw-l6alpha chain precursor IPIOO472035 Splice Isoform 2 Of ICOS ligand precursor IPIOO414888 Splice Isoform 2 Of Insulin receptor precursor IPOO220325 Splice Isoform 2 Of Integrin alpha-7 precursor IPIOO220748
FIG 7R U.S. Patent Aug. 18, 2009 Sheet 94 of 97 US 7,575,876 B2
Splice Isoform 2 Of Interleukin-12 receptor beta-2 chain precursor IP I0043.8856 Splice Isoform 2 Of Interleukin-18 binding protein precursor IPIOO220525 Splice Isoform 2 Of MAGUK p55 subfamily member 2 I PIO021827 Splice Isoform 2 Of MAM domain-containing pool glycosylphosphatidylinositol anchor protein 1 IPIOO410349
Spliplice Isoform 2 Of Mitochondrial- - dicarboxylate- - carrier Splice Isoform 2 Of Myosin Va IPIOO 100956 Splice Isoform 2 Of Myosin VIIa IPIO0215753 Splice Isoform 2 Of N-acetylmuramoyl-L-alanine amidase precursor IPIOO394992 Splice Isoform 2 Of Neural cell adhesion molecule 1, 120 kDa isoform OcCuSO IPIO0220737 Splice Isoform 2 Of NTF2-related export protein 2 IPIO0221003 Splice Isoform 2 Of Oral-facial-digital syndrome 1 protein IPIO022.1364 Splice Isoform 2 Of Pleckstrin homology domain containing family C member 1 IPIOO383500 Splice Isoform 2 Of Protachykinin l precursor IPO0219086 Splice Isoform 2 Of Protein-L-isoaspartate Splice Isoform 2 Of Putative polypeptide N- posts acetylgalactosaminyltransferase-like protein IPIOO456715 Splice Isoform 2 Of Roundabout homolog 2 precursor IPIOO420043
carrier-associated membrane protein 1 IPIO0067352 Splice Isoform 2 Of Sentrin-specific protease 6 IPIO0332748 Splice Isoform 2 Of Sodium/potassium/calcium exchanger 2 precursor IPIO0218.809 Splice Isoform 2 Of Solute carrier family 26 member 6 IPIO02 18923 Splice Isoform 2 Of Stromal cell-derived factor 1 precursor IPIO0216304 Splice Isoform 2 Of T-cell surface glycoprotein E2 precursor IPIOO22011 7 Splice Isoform 2 Of Trans-Golgi network integral membrane protein 2 precursor IPIOO297.543 Splice Isoform 2 Of Ubiquilin 1 PIO0071 180 Splice Isoform 2 Of UDP-N-acetylglucosamine-peptide N acetylglucosaminyltransferase ll IPIO0219856 Splice Isoform 2 Of Voltage-dependent N-type calcium channel alphal B Subunit IPIOO220431.
Splice Isoform 3 Of Calcium/calmodulin-dependent protein kinase type II
beta chain IPIO0219 165
FIG. 7S U.S. Patent Aug. 18, 2009 Sheet 95 of 97 US 7,575,876 B2
IP Address Splice Isoform 3 Of Collagen alpha 2CVI) chain precursor IPIO0073.454 Splice Isoform 3 Of Dachshund homolog 2 IPIOO402353 Splice Isoform 3 Of Latrophilin 3 precursor IPIOO410312 Splice Isoform 3 Of Myosin XVIIIA Splice Isoform 3 Of Osteopontin precursor IPIO02 18875 Splice Isoform 3 Of Receptor-type tyrosine-protein phosphatase delta roo OeCuSO IPIOO29860 Splice Isoform 3 Of Reelin precursor IPIOO298066 Splice Isoform 3 Of Seizure 6-like protein precursor IPIOO220333 Splice Isoform 3 Of Signal-regulatory protein beta-2 precursor IPIO02 18601 precursor
Splice Isoform 4 Of Peptidyl-glycine alpha-amidating monooxygenase OeCuSO IPIO02 19043 Splice Isoform 4 Of Receptor-type tyrosine-protein phosphatase S precursor IPIO0332273 Splice Isoform 5 Of Chordin precursor IPIO022 163 Splice Isoform 5 Of Neuronal cell adhesion molecule precursor PIO0333781 Splice Isoform 5 Of Receptor-type tyrosine-protein phosphatase S OeCuSO IPIO0299590 Splice Isoform 6 Of Fibronectin precursor IPO0339226 Splice Isoform 6 Of Myelin-oligodendrocyte glycoprotein precursor IPIO02 19666 Splice Isoform 7 Of Calcium/calmodulin-dependent protein kinase type II beta chain IPIO0183066 SRPK2 protein
SWI/SNF-related matrix-associated actin-dependent regulator of chromatin al isoform b IPIO0376861 S IPIO0012902
FIG 7T U.S. Patent Aug. 18, 2009 Sheet 96 of 97 US 7,575,876 B2
IPI Address IPIO0216138
IPIO0298.447 IPIO0386630 Testis-specific BRDT protein IPIOO41 0355 IPIO0027774 p IPIO0022970 Transcription elongation regulator l IPIOO247871 Transcription factor SLl PIO0385907 Transcription factor SOX-7 IPIO0027779 Transcription initiation factor IIE, alpha subunit IPIO00 19977 Transcriptional activator SRCAP IPIO0009101 Transmembrane 7 superfamily member 1 IPIOOO1901 7 Transmembrane protein 16B IPIO0033553 TRAP/Mediator complex component TRAP25 IPIO0063213 TRIF-related adapter molecule IPIO032928 IPIOO465028 IPIO0298237 IPIOO450071 IPIOO47.9615 IPIOO169276 TSLCl-like 2 IPIOO176427 Tumor necrosis factor receptor superfamily member 19L precursor IPIO006.4377 Tyrosine phosphatase zeta polypeptide 2 HTPZP2 IPIOO472466 Tyrosine-protein kinase CSK IPIOOO 13212 Ubiquitin carboxyl-terminal hydrolase 24 PIOO398505 Ubiquitin carboxyl-terminal hydrolase isozyme Ll IPIOOO18352 Ubiquitin-like protein flubi and ribosomal protein S30 precUrsor IPIO00 19770 UDP-Gal:betaGlcNAc beta 1,4-galactosyltransferase 1, membrane-bound form IPIO0215767 IPIO0020512 IPIO0216914 IPIOO 1071 13 IPIO0009329 Vesicular integral-membrane protein VIP36 precursor IPIO0009950 Vitamin Kepoxide reductase complex, subunit 1-like 1 IPIOO166079 IPIOOO1 1072
IPIO0396243 IPIOO 102997
IPIO0383520 IPIO0288918
IPIOO399361
FIG. 7U U.S. Patent Aug. 18, 2009 Sheet 97 of 97 US 7,575,876 B2
PI Address Zinc finger protein 577 IPIOOO 13397
Zinc finger protein 95 homolog IPIO0032.316 ZNF627 protein PIOOO29023
FIG. 7V US 7,575,876 B2 1. 2 BOMARKERS FOR NEURODEGENERATIVE Furthermore, markers are most useful if they can detect at an DISORDERS early or even preclinical stages of diseases. In searching for biochemical markers in body fluids, including plasma, urine, CROSS-REFERENCE and cerebrospinal fluid (CSF), only limited success has been achieved despite decades of research. It has been felt recently This application claims the benefit of U.S. Provisional that this is largely due to the heterogeneity of all neurodegen Application No. 60/731,339, filed Oct. 27, 2005, which appli erative diseases, i.e. several markers may be needed to detect cation is incorporated herein by reference in its entirety. subpopulations of patients (Olsson et al., Clin Chem, 2005. 51(2): p. 336-45). GOVERNMENT RIGHTS 10 The development of genomics, proteomics, and metabolo mics has greatly enhanced the ability to discover multiple This invention was made with government Support under markers that are not only useful for diagnosis of AD, PD and federal grant nos. R01AG025327 and R01ES012703 DLB but also shed more lights on their pathogenesis. How awarded by National Institutes of Health. The United States ever, these studies are limited, as none has taken other neu Government may have certain rights in this invention. 15 rodegenerative diseases into consideration, and in addition, very few studies have been performed using cases with patho SEQUENCE LISTING logical verification. The present invention addresses this need. The present specification incorporates herein by reference, each in its entirety, the sequence information on the Compact Relevant Literature Disks (CDs) labeled Copy 1 and Copy 2. The CDs are for Bacskai et al., J. Cereb Blood Flow Metab, 2002. 22(9): p. matted on IBM-PC, with operating system compatibility with 1035-41: Klunk et al., J Neuropathol Exp Neurol, 2002. MS-Windows. The files on each of the CDs are as follows: 61 (9): p. 797-805; Small et al., J Mol Neurosci, 2002. 19(3): p. 323-7: Jankovic et al., ArchNeurol, 2000.57(3): p. 369-72: Copy 1—Seqlist.txt 214 KB created Jul. 28, 2006; 25 Hughes et al., Brain, 2002. 125(Pt 4): p. 861-70; Litvan et al., and Arch Neurol, 1998. 55(7): p. 969-78; Rajput et al., Can J Neurol Sci, 1991. 18(3): p. 275-8: Hughes et al., J Neurol Copy 2 Seqlist.txt 214 KB created Jul. 28, 2006. Neurosurg Psychiatry, 1992. 55(3): p. 181-4; McKeith et al., Semin ClinNeuropsychiatry, 2003.8(1): p. 46-57: Love et al., BACKGROUND OF THE INVENTION 30 Histopathology, 2004. 44(4): p. 309-17; Olsson et al., Clin Chem, 2005. 51(2): p. 336-45; Zhanget al., Neurobiol Aging, Neurodegenerative disorders, e.g. Alzheimer's disease 2005. 26(2): p. 207-27; and Zhang et al., J Alzheimers Dis, (AD), Parkinson's disease (PD), and dementia with Lewy 2005. 7(2): p. 125-33. body (DLB) diseases, are diagnosed primarily by clinical SUMMARY OF THE INVENTION presentations, limited laboratory investigations and, more 35 recently, structural and functional neuroimaging analysis (Bacskai et al., J. Cereb Blood Flow Metab, 2002. 22(9): p. The present invention provides methods for diagnosing 1035-41: Klunk et al., J Neuropathol Exp Neurol, 2002. neurodegenerative disease, such as Alzheimer's Disease, Par 61 (9): p. 797-805; and Small et al., J Mol Neurosci, 2002. kinson's Disease, and dementia with Lewy body disease by 19(3): p. 323-7). However the diagnosis based on these 40 detecting a pattern of gene product (e.g., protein) expression approaches is unsatisfactory. As determined by pathological in a cerebrospinal fluid sample and comparing the pattern of examination, diagnostic accuracy of various neurodegenera gene product expression from the sample to a library of gene tive diseases varies between 50% to 85% depending on the product expression pattern known to be indicative of the disease involved, the experience of physicians and the stages presence or absence of a neurodegenerative disease. Also of the diseases (Jankovic et al., Arch Neurol, 2000. 57(3): p. 45 provided are kits, systems and devices for practicing the 369-72; Hughes et al., Brain, 2002. 125(Pt 4): p. 861-70; Subject methods. Litvan et al., Arch Neurol, 1998.55(7): p. 969-78; Rajput et The present invention provides a method for detecting al., CanJ NeurolSci, 1991. 18(3): p. 275-8; Hughes et al., J presence or absence of a neurodegenerative disease in a Sub Neurol Neurosurg Psychiatry, 1992. 55(3): p. 181-4; and ject by detecting a pattern of gene product expression present McKeith et al., Semin Clin Neuropsychiatry, 2003. 8(1): p. 50 in a cerebrospinal fluid sample obtained from a subject; and 46-57). The fact that the diagnosis cannot be made with comparing the pattern of gene product expression from the reasonable certainty until the latter stages of the diseases cerebrospinal fluid sample to a library of gene product expres possibly underlies the current state of clinical management, sion pattern known to be indicative of the presence or absence i.e. none of the available therapies, particularly those aimed at of a neurodegenerative disease, wherein the comparing indi preventing disease's progression, is effective; this could sim 55 cates the presence or absence of a neurodegenerative disease. ply be due to the fact that most neurons are already degener In some embodiments, the gene product is a polypeptide. ated by the time diagnosis is made. It is also noteworthy that In some embodiments, the detecting is by mass spectrometry. it is common for patients with various neurodegenerative In other embodiments, the detecting is by immunoassay. In diseases to go undetected using current approaches (Love et certain embodiments, the immunoassay is enzyme linked al. Histopathology, 2004. 44(4): p. 309-17). 60 immunosorbent assay (ELISA). In other embodiments, the Biomarkers are biological characteristics used to indicate detecting by a Luminex XMAP system. In certain embodi or to measure disease risk, presence, and progression. Ideally, ments, the neurodegenerative disease is Alzheimer's disease, an optimal biomarker should be precise, reliable, inexpen Parkinson's disease, or dementia with Lewy body disease. sive, as well as reflect the pathophysiological mechanisms of The present invention also provides a method for monitor neurodegenerative diseases. Presently, no established diag 65 ing progression of a neurodegenerative disease in a subject by nostic biomarkers can confirm AD, PD or DLB or monitor detecting a first pattern of expression of gene products present their progression with high sensitivity at high specificity. in a cerebrospinal fluid sample obtained from a Subject at a US 7,575,876 B2 3 4 first time point, wherein said first pattern is indicative of a These and other objects, advantages, and features of the neurodegenerative disease; detecting a second pattern of invention will become apparent to those persons skilled in the expression of gene products present in a cerebrospinal fluid art upon reading the details of the invention as more fully sample obtained from a subject at a second time point; and described below. comparing the first and second patterns of expression of gene 5 products from the cerebrospinal fluid samples, wherein the BRIEF DESCRIPTION OF THE DRAWINGS comparing provides for monitoring of the progression of the The invention is best understood from the following neurodegenerative disease from the first time point to the detailed description when read in conjunction with the second time point. 10 accompanying drawings. It is emphasized that, according to In some embodiments, the gene product is a polypeptide. common practice, the various features of the drawings are not In some embodiments, the detecting is by mass spectrometry. to-scale. On the contrary, the dimensions of the various fea In other embodiments, the detecting is by immunoassay. In tures are arbitrarily expanded or reduced for clarity. Included certain embodiments, the immunoassay is enzyme linked in the drawings are the following figures: 15 FIG. 1 shows a pie chart depicting the 1,540 proteins char immunosorbent assay (ELISA). In other embodiments, the acterized by nano-LC-MALDI-TOF-TOF. A complete list of detecting by a Luminex XMAP system. In certain embodi the identified proteins is provided in FIGS. 6A-6T and FIGS. ments, the neurodegenerative disease is Alzheimer's disease, 7A-V. Parkinson's disease, or dementia with Lewy body disease. FIG. 2 shows confirmation of B-Fibrinogen with Western The present invention also provides a method of providing blot in pooled and individual samples. Proteomic data a differential diagnosis between Alzheimer's disease (AD), showed that B fibrinogen increased significantly over controls Parkinson's disease (PD), and dementia with Lewy body with pooled samples. Panel A: with standard Western blot disease (DLB) in a subject by detecting a pattern of gene protocol, pooled samples were analyzed with an antibody product expression present in a cerebrospinal fluid sample against B fibrinogen (1:2000) both based on total loading 25 amount (10 ug, i.e. similar to proteomic analysis) or CSF obtained from a subject; and comparing the pattern of gene volume (10 ul). Panel B: with identical approach in Panel A. product expression from the cerebrospinal fluid sample to a but B fibrinogen was analyzed again in individual samples. library of gene product expression patterns known to be AD: Alzheimer's disease; PD: Parkinson's disease; DLB: indicative of the presence or absence of AD, PD and DLB, dementia with Lewy body disease, CT: age-matched controls. wherein the comparing providing a differential diagnosis 30 FIG. 3 shows composite markers for AD vs. other neuro between AD, PD, and DLB. degenerative diseases. Panel A is a scatter plot showing the association between standardized B fibrinogen and standard In some embodiments, the gene product is a polypeptide. ized VitD BP with AD, PD, and DLB cases and healthy In some embodiments, the detecting is by mass spectrometry. controls as well. Line in plot represents the composite marker In other embodiments, the detecting is by immunoassay. In 35 defined from logistic regression. The actual line represented certain embodiments, the immunoassay is enzyme linked here gives the classification rule for 95% specificity. Panel B immunosorbent assay (ELISA). In other embodiments, the shows ROC curves for VitD BPB fibrinogen and the compos detecting by a Luminex XMAP system. ite marker (CM). The following statistics are obtained from The present invention also provides a system, including a CM: AUC (area under curve)=0.99; Sensitivity at 95%–1.00 40 with p-value for VitD BP=0.0635 and p-value for B-fibrino computing environment; an input device, connected to the gen=0.0207. computing environment, to receive data from a user, wherein FIG. 4 shows composite markers for PD vs. other neuro the data received includes a pattern of gene product expres degenerative diseases. Panel A is a scatter plot showing the sion from a cerebrospinal fluid sample obtained from a sub association between standardized Chromogramin B and stan ject; an output device, connected to the computing environ 45 dardized ApoH with AD, PD and DLB cases and healthy ment, to provide information to the user; and a computer controls as well. Line in plot represents the composite marker readable storage medium having stored thereon at least one defined from the logistic regression. The actual line repre algorithm to provide for comparing the pattern of gene prod sented here gives the classification rule for 95% specificity. uct expression from the cerebrospinal fluid sample to a library Panel B: ROC curves for chromogranin B, ApoH and the of gene product expression pattern known to be indicative of 50 composite marker (CM). The following statistics are obtained the presence or absence of a neurodegenerative disease. In from CM: AUC (area under curve)=0.92; Sensitivity at 95%–0.78 with p-value for chromogranin B=0.0056 and Some embodiments, the computing environment includes a p-value for ApoH=0.0068. local computer local to the user and a remote computer at a FIGS. 5A-5YY is a table showing the proteins that have site remote to the user, wherein the local computer and the 55 changes in expression levels unique to AD, PD, or DLB. The remote computer are connected through a network, and table is presented in six sections (I) proteins unique to AD and wherein the computer readable storage medium is provided identified by two or more peptides; (II) proteins unique to AD on the remote computer. and identified by a single peptide; (III) proteins unique to PD The present invention also provides a computer readable and identified by two or more peptides; (IV) proteins unique medium including a program stored thereon, wherein the 60 to PD and identified by a single peptide: (V) proteins unique program provides for execution of one or more algorithms to to DLP and identified by two or more peptides; and (VI) provide for comparing a pattern of gene product expression proteins unique to DLB and identified by a single peptide. The identified proteins have also been grouped based on from a cerebrospinal fluid sample obtained from a subject to function within each category. Exemplary functional group a library of gene product expression pattern known to be 65 ings include neuronal activities/signal transduction, cell indicative of the presence or absence of a neurodegenerative structure/motility/transport/traffic, and extracellular matrix/ disease. cell adhesion, immunity/defense. The assignment of function