From Cancer Genomics to Cancer Treatment : from hope to reality Yusuke Nakamura Human Genome Center Institute of Medical Science The University of Tokyo International HapMap Consortium October 27, 2005 Construction of public database for genetic variations in human Country Genotyping Center %Genome Chromosome Platform Japan RIKEN 24.3% 5, 11, 14, 15, 16, 17, 19 Third Wave Invader Wellcome Trust Sanger UK 23.7% 1, 6, 10, 13, 20 Illumina BeadArray Institute McGill Univ. / Genome Canada 10.1% 2, 4p Illumina BeadArray QuebecWe Innovation (The Centre SNP Research Center in RIKEN) Sequenom MassExtend, China Chinesecontributed HapMap Consortium the largest9.5% 3, SNP8p, 21 data (24.3%) contributed the largest SNP data (24.3%)Illumina BeadArray Illumina among the16.1% SNP 8q,typing 9, 18q, 22, centers X Illumina BeadArray Broad Institute of Harvard and in this paper Sequenom MassExtend, 9.7% 4q, 7q, 18p, Y, mtDNA MIT Illumina BeadArray USA Baylor College of Medicine 4.6% 12 ParAllele MIP PerkinElmer AcycloPrime- UCSF / Washington Univ. 2.0% 7p FP High-denstity Perlegen Sciences All oligonucleotide array Biobank Japan Sample Collection at February 29, 2008 Hyperlipidemia 42,354 Prostate cancer 5,839 Leukemia 1,597 Diabetes 39,982 Periodontitis 5,652 Esophageal cancer 1,461 Cataract 19,070 Pollinosis 5,572 Cervical caner 1,405 Cerebral infarction 16,012 Glaucoma 5,257 Hepatitis B 1,391 Arrhythmia 15,440 Lung cancer 4,845 Uterine corpus cancer 1,189 Stable angina pectoris 14,855 Unstable angina pectoris 4,161 Nephrotic syndrome 1,038 MyocardialWe Infarction have 12,956 genotyped Rheumatoid arthritis more than 4,155 Ovarian15,000 cancer patients 976 BronchialWe asthma have 8,657genotyped Atopic dermatitis more than2,967 Tuberculosis15,000 patients 894 Cardiac failure 7,438 COPDat 250K-550K 2,797SNPs Keloid 814 Breast cancer 7,349 Cerebral aneurysm 2,735 ALS 788 Colorectal cancer 6,957 Arteriosclerotic obliterans 2,609 ILD 780 Gastric cancerA 6,869 total Liver cirrhosisof 6-billion data-points.2,494 Drug-induced hypersensitivity 598 Urinary stone 6,605 Liver cancer 2,452 Pancreatic cancer 531 Osteoporosis 6,412 Hyperthyroidism 2,320 CCC 503 Myoma uteri 5,988 Epilepsy 2,250 Febrile seizures 475 Hepatitis C 5,962 Endometriosis 1,827 Total 295,278 ICIC askedasked 235,841 235,841 patientspatients TotalTotal Cases Cases 295,278 295,278 cases cases ICIC obtainedobtained 201,805 201,805 patientspatients WithdrawnWithdrawn 172 172 individuals individuals （（85.6%)85.6%) Genes isolated through genome-wide association studies in RIKEN and University of Tokyo Myocardial Infarction LTA Nature Genetics 2002 LGALS2 Nature 2004 PSMA6 Nature Genetics 2006 Rheumatoid Arthritis PADI4 Nature Genetics 2003 SLC22A4 Nature Genetics 2003 FcRH3 Nature Genetics 2005 Diabetic nephropathySLC12A3 Diabetes 2003 ELMO1 Diabetes 2005 (Diabetes) KCNQ1 Nature Genetics 2008 IgA nephropathy SEL-L,-E Am J Hum Genet 2002 Osteoarthritis Asporin Nature Genetics 2005 Calmodulin 1 Human Mol. Gen 2005 GDF5 Nature Genetics 2007 DVWA Nature Genetics 2008 Disc herniation CILP Nature Genetics 2005 Brain Infarction PRKCH1 Nature Genetics 2007 Kawasaki disease ITPKC Nature Genetics 2008 Crohn disease TNSF15 Human Mol. Gen 2005 Colon cancer multipel genes Nature Genetics 2008 Lung fibrosis TERT JMG 2008 Hapmap Nature 2003 Nature 2005 Nature 2007 From molecular targets to antianti-cancer-cancer drugs SMC Antibody Dominant-negative Target Peptide Molecules peptide Molecules vaccine Anti-sense DNA Cell Gene siRNA Therapy Therapy From molecular targets to antianti-cancer-cancer drugs SMC Antibody Dominant-negative Target Peptide Molecules peptide Molecules vaccine Anti-sense DNA Cell Gene siRNA Therapy Therapy IsolationIsolation ofof molecularmolecular targetstargets forfor cancercancer treatmenttreatment usingusing clinicalclinical materialsmaterials cDNA microarray consisting of 32,000 genes ComparisonComparison ofof expressionexpression ExpressionExpression profilesprofiles ofof profilesprofiles ofof cancercancer andand 3030 normalnormal humanhuman correspondingcorresponding normalnormal tissuestissues tissuestissues Selection of novel molecular targets DiagnosisDiagnosis TreatmentTreatment TumorTumor MarkerMarker SmallSmall molecularmolecular compoundcompound PredictionPrediction toto MonoclonalMonoclonal AntibodyAntibody chemosensitivitychemosensitivity PeptidePeptide VaccineVaccine siRNAsiRNA cDNAcDNA MicroarrayMicroarray systemsystem High-density spotting Cancer 20000 spots/glass Normal tissue tissue ＬＭＭ ＬＭＭ Cancer cells Normal ductal cells T7-based RNA Amplification T7-based RNA Amplification （2-rounds） （2-rounds） aRNA aRNA labeling labeling Cy5 Cy3 co-hybridization Number of clinical samples analyzed by cDNA microarray Tissue Number Tissue Number Lung 126 Bile duct 45 Breast 135 Uterus 44 Soft Tissue 101 ALL 25 AML 87 Kidney 25 Colon 78 Endometriosis 23 CML 84 Liver 20 Ovary 59 Pancreas 20 Malignant lymphoma 54 Melanoma 20 Prostate 54 Thyroid 20 Stomach 51 Neuroblastoma 16 Bladder 55 Testis 13 Eshophagus 45 TOTAL 1200 Criteria for selection of candidate targets for drug development (S) Small molecular compound (P) Peptide vaccine (R) siRNA (A) Antibody 1. Genes which were highly over-expressed in a large proportion of clinical cancer samples examined (S, P, R, A) 2. Genes which were expressed in none of important vital organs; ideally not expressed in any organs (S, P, R, A) 3. Genes whose expressions are essential for cell survival (S, R, A?) 4. Cytoplasmic membrane protein, Secreted protein (A) Development of anti-FZD10 antibody therapy for Synovial Sarcoma Chikako Fukukawa Satoshi Nagayama Toyomasa Katagiri Frizzled Homologue 10 (FZD10) RT-PCR C S SS S MFH LMS L M Synovial Sarcoma MPNST cell lines 123456789101112131514 16 17 18 19 20 21 22 23 24 25 26 27 28 FZD10 b2MG Northern Blotting SS CL Srug. Frizzled family (Wnt signal) Lung Kidney Brain Liver Heart Pancreas SS582 SS487 Placenta HS-SY-2 YaFuSS Bone marrow Bone Wnt? 9.5 7.5 SS cell 4.4 2.37 1.35 SS ; Synovial Sarcoma FZD10 Protein expression of FZD10 in colon cancer 03-24640 FZD10 expression in colon cancer 34 41 59 69 123 124 128 129 141 146 147 NTNTNTNTNTNTNTNTNTNPr MN Pr M FZD10 β2MG 149 150 151 153 154 155 201 NMPr NMPr NMPr NMPr NPPr NNTrSS FZD10 β2MG 04-26192 04-25950 Paraffin slides EDTA buffer (pH9.0) 125oC, 30sec α-FZD10 mAb 92-13 20μg/mL, 4oC, O/N Celler Immunization Monoclonal antibody recognizing a complex structure Cells (COS-7 etc.) transfection Ag pCAGGS-FZD10 (FL)-myc･His Immunization to mouse Monoclonal Antibody Hybrydoma Antibody stayed at tumor lesion at 5 days after injection InternalizationInternalization ofof anti-FDZ10anti-FZD10 antibodyantibody FZD10(+) FZD10(-) SYO-1 YaFuSS LoVo No Tx No 92-13 93-22 In vivo effect of Y90-anti-FZD10 Antibody Day 0 Day 5 Day 9 Day 40 Day 54 90Y-anti-FZD10 Day 0 Day 9 Day 33 Day 40 Day 54 90Y-anti-FZD10 Day 0 Day 9 90Y-CD20-Ab In vivo effect of Y90-anti-FZD10 Antibody Balb-c/nu (male) / SYO-1 tumor 90Y - DTPA - antibody 100uCi Intraveneous injection Single injection on Day0 7 Non-Labeled 92-13 (n=5) Non-treated (n=5) 6 ) 3 5 4 3 Tumor Volume(cm 2 90Y - 92-13 (n=11) 1 0 0 5 10 15 20 25 30 35 40 Days Injection 11 / 11 4 / 11 DevelopmentDevelopment of cancercancer peptidepeptide vaccine andand ConstructionConstruction ofof TRTR networknetwork inin JapanJapan Types of Cancer Vaccines • Antigen/adjuvant vaccines • Whole cell cancer vaccines • Dendritic cell (DC) vaccines • Idiotype vaccines RecentRecent advancesadvances forfor cancercancer vaccinevaccine • 1991 Discovery of Tumor specific antigen (T. Boon, Science) • 1995 Clinical Trial against melanoma (Int J Cancer) • 1998 IL-2 + Peptides (Rosenberg, Nature Med) DC + Peptide (Nestle, Nature Med) • 2004 Less than 3% response rate for advanced cancer (Rosenberg, Nature Med) • 2006 33%33% reductionreduction ofof recurrencerecurrence forfor lunglung cancercancer afterafter surgerysurgery (GSK,(GSK, ASCO)ASCO) 50%50% reductionreduction ofof recurrencerecurrence forfor breastbreast cancercancer afterafter surgerysurgery (Peoples,(Peoples, SanSan AntonioAntonio Int.Int. BreastBreast CancerCancer Meeting)Meeting) Expectation to Cancer Vaccine Treatment 100 1998 Promising Results on 2015 Approval melanonma of 15-20 Vac. 2007 Effect on reduction 0 of recurrence 1991 Tumor antigen 2003 Rosenberg report -100 1990 2000 2010 2020 AppropriateCancer Vaccine Cancer Treatment Vaccine atTreatment present 10-cm tumor = 1011-12 cells CTL CTL CTL CTL＜＜＜＜＜Cancer＜＜＜＜＜CancerCancer CellsCells 10000 cells Peripheral Lym 1010 cells CTL=106-7 cells CTL Vaccine Treatment at an earlier stage of cancer Canc er Canc CTL er CTL Canc Canc er er CTL Canc CTL Canc Canc er CTL er er Canc CTL er CTL Canc er CTL CTL CTL Canc er Canc er CTL CancerFurther Vaccine enhancement Treatment of CTL at activity present CTL CTL CTL CTL CTL＜＜＜＜＜Cancer＜＜＜＜＜Cancer Cells 10000 cells CTL FurtherTo overcom enhancement the present of CTL status activity CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL １．Improvement of CTL inductionCTL CTL CTL CTL CTL ２．BestCTL selection of antigen CTL CTL CTL CTL UnevenCTL precursor T cell population CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL ３．Regulation of Treg CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL DevelopmentDevelopment of cancer vaccinevaccine Development of cancer vaccines using novel tumortumor-specific-specific oncoantigensoncoantigens identifiedidentified through genome-widegenome-wide cDNAcDNA microarraymicroarray analysisanalysis andand subsequent“Oncoantigen” functional analysis is defined