Yusuke Nakamura Human Genome Center Institute of Medical Science - - PowerPoint PPT Presentation

From Cancer Genomics to Cancer Treatment : from hope to reality

Yusuke Nakamura

Human Genome Center Institute of Medical Science The University of Tokyo

October 27, 2005

International HapMap Consortium International HapMap Consortium

Construction of public database for genetic variations in human 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 UK Wellcome Trust Sanger Institute 23.7% 1, 6, 10, 13, 20 Illumina BeadArray Canada McGill Univ. / Genome Quebec Innovation Centre 10.1% 2, 4p Illumina BeadArray China Chinese HapMap Consortium 9.5% 3, 8p, 21 Sequenom MassExtend, Illumina BeadArray USA Illumina 16.1% 8q, 9, 18q, 22, X Illumina BeadArray Broad Institute of Harvard and MIT 9.7% 4q, 7q, 18p, Y, mtDNA Sequenom MassExtend, Illumina BeadArray Baylor College of Medicine 4.6% 12 ParAllele MIP UCSF / Washington Univ. 2.0% 7p PerkinElmer AcycloPrime- FP Perlegen Sciences All High-denstity

• ligonucleotide array

We (The SNP Research Center in RIKEN) contributed the largest SNP data (24.3%) among the SNP typing centers in this paper We (The SNP Research Center in RIKEN) contributed the largest SNP data (24.3%) among the SNP typing centers in this paper

Biobank Japan Sample Collection at February 29, 2008 Biobank Japan Sample Collection at February 29, 2008

IC asked 235,841 patients IC obtained 201,805 patients （ 85.6%) IC asked 235,841 patients IC obtained 201,805 patients （ 85.6%) Total Cases 295,278 cases Withdrawn 172 individuals Total Cases 295,278 cases Withdrawn 172 individuals

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 Myocardial Infarction 12,956 Rheumatoid arthritis 4,155 Ovarian cancer 976 Bronchial asthma 8,657 Atopic dermatitis 2,967 Tuberculosis 894 Cardiac failure 7,438 COPD 2,797 Keloid 814 Breast cancer 7,349 Cerebral aneurysm 2,735 ALS 788 Colorectal cancer 6,957 Arteriosclerotic obliterans 2,609 ILD 780 Gastric cancer 6,869 Liver cirrhosis 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

We have genotyped more than 15,000 patients at 250K-550K SNPs A total of 6-billion data-points. We have genotyped more than 15,000 patients at 250K-550K SNPs A total of 6-billion data-points.

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 anti-cancer drugs From molecular targets to anti From molecular targets to anti-

• cancer drugs

cancer drugs

Target Molecules Target Molecules SMC

Antibody Peptide vaccine Anti-sense DNA siRNA

Dominant-negative peptide

Gene Therapy Cell Therapy

From molecular targets to anti-cancer drugs From molecular targets to anti From molecular targets to anti-

• cancer drugs

cancer drugs

Target Molecules Target Molecules SMC

Antibody Peptide vaccine Anti-sense DNA siRNA

Dominant-negative peptide

Gene Therapy Cell Therapy

Diagnosis Tumor Marker Prediction to chemosensitivity Diagnosis Tumor Marker Prediction to chemosensitivity cDNA microarray consisting of 32,000 genes

Isolation of molecular targets for cancer treatment using clinical materials Isolation of molecular targets for cancer treatment using clinical materials

Comparison of expression profiles of cancer and corresponding normal tissues Comparison of expression profiles of cancer and corresponding normal tissues Expression profiles of 30 normal human tissues Expression profiles of 30 normal human tissues

Selection of novel molecular targets

Treatment Small molecular compound Monoclonal Antibody Peptide Vaccine siRNA Treatment Small molecular compound Monoclonal Antibody Peptide Vaccine siRNA

N

• r

m a l d u c t a l c e l l s N

• r

m a l t i s s u e a R N A a R N A l a b e l i n g C y 5 C y 5 C y 3 C y 3 C a n c e r t i s s u e c

• h

y b r i d i z a t i

• n

l a b e l i n g

cDNA Microarray system cDNA Microarray system

C a n c e r c e l l s

T 7

• b

a s e d R N A A m p l i f i c a t i

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a s e d R N A A m p l i f i c a t i

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Ｌ ＭＭ Ｌ ＭＭ H i g h

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Number of clinical samples analyzed by cDNA microarray 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

• 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
• rgans; 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)
• 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
• rgans; 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)

Criteria for selection of candidate targets for drug development (S) Small molecular compound (P) Peptide vaccine (R) siRNA (A) Antibody

Development of anti-FZD10 antibody therapy for Synovial Sarcoma Development of anti-FZD10 antibody therapy for Synovial Sarcoma

Chikako Fukukawa Satoshi Nagayama Toyomasa Katagiri Chikako Fukukawa Satoshi Nagayama Toyomasa Katagiri

Frizzled Homologue 10 (FZD10) Frizzled Homologue 10 (FZD10)

RT-PCR

Northern Blotting

Frizzled family (Wnt signal)

SS cell SS ; Synovial Sarcoma

Wnt? M S C MFH LMS L S SS cell lines MPNST Synovial Sarcoma 1 2 3 4 5 6 7 8 9 10 1112 13 15 14 16 17 18 19 20 21 22 23 24 25 26 27 28

b2MG FZD10

Heart Brain Lung Liver Kidney Bone marrow Pancreas Placenta HS-SY-2 YaFuSS

9.5 7.5 4.4 2.37 1.35

SS487 SS582 SS CL Srug.

β2MG FZD10

N T M Pr 34 41 59 69 123 124 128 129 141 146 147 149 150 151 153 154 155 201 N T N T N T N T N T N T N T N T N M Pr N Pr N M Pr N M Pr N M Pr N M Pr N Pr N N T SS

β2MG FZD10

FZD10 expression in colon cancer

Protein expression of FZD10 in colon cancer

03-24640 04-25950 04-26192

Paraffin slides EDTA buffer (pH9.0) 125oC, 30sec α-FZD10 mAb 92-13 20μg/mL, 4oC, O/N

FZD10

Celler Immunization

Cells(COS-7 etc.) transfection Immunization to mouse Hybrydoma Monoclonal Antibody

Monoclonal antibody recognizing a complex structure

pCAGGS-FZD10 (FL)-myc･His

Ag

Antibody stayed at tumor lesion at 5 days after injection

Internalization of anti-FDZ10 antibody Internalization of anti-FZD10 antibody

SYO-1 YaFuSS LoVo FZD10(+) FZD10(-) No Tx 92-13 93-22

90Y-CD20-Ab

Day 0 Day 9

In vivo effect of Y90-anti-FZD10 Antibody

90Y-anti-FZD10

Day 0 Day 5 Day 9 Day 33 Day 40 Day 54 Day 40 Day 54 Day 9 Day 0

90Y-anti-FZD10

5 10 15 20 25 30 35 40 Days 1 2 3 4 5 6 7 Tumor Volume (cm3)

Non-treated (n=5) Non-Labeled 92-13 (n=5)

Balb-c/nu (male) / SYO-1 tumor

90Y - DTPA - antibody 100uCi

Intraveneous injection Single injection on Day0

Injection

90Y - 92-13 (n=11)

11 / 11 4 / 11

In vivo effect of Y90-anti-FZD10 Antibody

Development of cancer peptide vaccine and Construction of TR network in Japan Development of cancer peptide vaccine Development of cancer peptide vaccine and and Construction of TR network in Japan Construction of TR network in Japan

Types of Cancer Vaccines Types of Cancer Vaccines

• Antigen/adjuvant vaccines
• Whole cell cancer vaccines
• Dendritic cell (DC) vaccines
• Idiotype vaccines
• Antigen/adjuvant vaccines
• Whole cell cancer vaccines
• Dendritic cell (DC) vaccines
• Idiotype vaccines

Recent advances for cancer vaccine Recent advances for cancer vaccine

• 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% reduction of recurrence for lung cancer after surgery 33% reduction of recurrence for lung cancer after surgery (GSK, ASCO) (GSK, ASCO) 50% reduction of recurrence for breast cancer after surgery 50% reduction of recurrence for breast cancer after surgery (Peoples, San Antonio Int. Breast Cancer Meeting) (Peoples, San Antonio Int. Breast Cancer Meeting)

Expectation to Cancer Vaccine Treatment Expectation to Cancer Vaccine Treatment

1

• 1

1 9 9 2 2 1 2 2

1991 Tumor antigen 1998 Promising Results on melanonma 2003 Rosenberg report 2007 Effect on reduction

• f recurrence

2015 Approval

• f 15-20 Vac.

CTL CTL CTL CTL CTL CTL

10000 cells

Cancer Vaccine Treatment at present Cancer Vaccine Treatment at present

CTL＜＜＜＜＜Cancer Cells CTL＜＜＜＜＜Cancer Cells

Cancer Cells Cancer Cells

Appropriate Cancer Vaccine Treatment Appropriate Cancer Vaccine Treatment

10-cm tumor = 1011-12 cells

Peripheral Lym 1010 cells CTL=106-7 cells

CTL CTL CTL CTL CTL CTL

Canc er Canc er Canc er Canc er Canc er Canc er Canc er Canc er Canc er

CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL

Canc er Canc er

Vaccine Treatment at an earlier stage of cancer

CTL CTL CTL CTL CTL CTL

10000 cells

Cancer Vaccine Treatment at present Cancer Vaccine Treatment at present

CTL＜＜＜＜＜Cancer Cells CTL＜＜＜＜＜Cancer Cells

CTL CTL CTL CTL

Further enhancement of CTL activity

CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL CTL

To overcom the present status Further enhancement of CTL activity

１ ． I m p r

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e m e n t

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C T L i n d u c t i

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a n t i g e n U n e v e n p r e c u r s

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T c e l l p

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T r e g

Development of cancer vaccines using novel tumor-specific oncoantigens identified through genome-wide cDNA microarray analysis and subsequent functional analysis Development of cancer vaccines Development of cancer vaccines using novel tumor using novel tumor-

• specific

specific oncoantigens

• ncoantigens identified

identified through genome through genome-

• wide cDNA microarray analysis and

wide cDNA microarray analysis and subsequent functional analysis subsequent functional analysis

Development of cancer vaccine Development of cancer vaccine Development of cancer vaccine

Development of cancer vaccines targeting tumor angiogenesis Development of cancer vaccines Development of cancer vaccines targeting tumor targeting tumor angiogenesis angiogenesis “Oncoantigen” is defined as a protein that functions as an

• ncogene product and can

enhance an immunogenic reaction. “Oncoantigen” is defined as a protein that functions as an

• ncogene product and can

enhance an immunogenic reaction.

Expression pattern of MPHOSPH1 （Bladder Cancer) Expression pattern of MPHOSPH1 （Bladder Cancer)

Growth suppressive effect of MPHOSPH1 siRNA Growth suppressive effect of MPHOSPH1 siRNA

MPHOSPH1 is defined as an “oncoantigen” Low chance of loss of antigen expression in cancer cells MPHOSPH1 is defined as an “oncoantigen” Low chance of loss of antigen expression in cancer cells From more than 50 oncoantigens identified, we have screened 9- or 10- amino-acid peptides (HLA-A02 or HLA-A24 restricted) that can stimulate CTL (cytotoxic T-lymphocytes) , and identified ~60 peptides covering most of cancer types. From more than 50 oncoantigens identified, we have screened 9- or 10- amino-acid peptides (HLA-A02 or HLA-A24 restricted) that can stimulate CTL (cytotoxic T-lymphocytes) , and identified ~60 peptides covering most of cancer types.

Since their expressions are restricted in cancer and testis (or fetal tissues or placenta), it is very unlikely that activated CTLs cause adverse reactions like autoimmune diseases. Since their expressions are restricted in cancer and testis (or fetal tissues or placenta), it is very unlikely that activated CTLs cause adverse reactions like autoimmune diseases.

Organizer: Takuya Tsunoda, Yusuke Nakamura

Cancer Peptide Vaccine Translational Network (Captivation Network)

Organ Num of Hospitals Esophagus 7 (2) Stomach 4 Colon 6 (1) Liver 3 (1) Bile duct 2 Pancreas 5 (1) Kidney 2 (2) Bladder 1 Lung 2 Breast 1 (2) Head and Neck 0 (2) Total 33 (11)

24 Universities 48 protocols for 11 organs (2008.08) 24 Universities 48 protocols for 11 organs (2008.08)

Colon Cancer Phase II Clinical Research Adjuvant setting Advanced cancer 16 Hospitals Colon Cancer Phase II Clinical Research Adjuvant setting Advanced cancer 16 Hospitals

Early Phase Development

• f Cancer Vaccines

Early Phase Development

• f Cancer Vaccines
• Study objective:

Identification of a pharmacologically effective dose or optimal biologic dose rather than MTD

• Study objective:

Identification of a pharmacologically effective dose or optimal biologic dose rather than MTD

• Rationale

Biologically active doses may occur well below the MTD Probably not feasible to examine MTD Immunological endpoints

• Rationale

Biologically active doses may occur well below the MTD Probably not feasible to examine MTD Immunological endpoints

The Aims of Captivation Network The Aims of Captivation Network

１．Evaluation of Safety １．Evaluation of Safety ２．Evaluation of immunological responses ２．Evaluation of immunological responses ３．Searching the most appropriate condition that can induce the maximum immunological response ３．Searching the most appropriate condition that can induce the maximum immunological response ４．Clinical Effect ４．Clinical Effect

The number of the patients enrolled The number of the patients enrolled

(Aug 31, 2008)

（ Total 212 cases)

Month

Cases

2006 2007 2008

5 1 1 5 2 2 5 3 3 5 4 4 5 5 A u g

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5 1 1 5 2 2 5 3 3 5 4 4 5 5 A u g

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a n F e b

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Phase 1 Phase 2

Cancer Vaccine Monotherapy for Advanced esophageal cancer （Phase I） Cancer Vaccine Monotherapy for Advanced Cancer Vaccine Monotherapy for Advanced esophageal cancer esophageal cancer （ （Phase I Phase I） ）

Yamanashi University, First Department of Surgery

• Drs. Koji Kono

Yoshiki Mizukami Hideki Fujii

• URLC10, TTK, KOC1 (each 1mg)
• IFA as an adjuvant
• HLA-A*2402-restricted
• Weekly s.c. injection
• URLC10, TTK, KOC1 (each 1mg)
• IFA as an adjuvant
• HLA-A*2402-restricted
• Weekly s.c. injection

1 Vac 2 Vac 3 Vac 4 Vac 5 Vac Vaccination day 1 day 8 day 15 day 22 day29 day43

Evaluation

Before (07.3.7) After 2 months (07.7.9) After 1 course (07.5.2) S1 S1+2 S1 S1 S1+2 S1+2

Case 5 Lung Metastasis Case 5 Lung Metastasis Case 5 Lung Metastasis

Case 5 Liver Metastasis Case 5 Liver Metastasis

Before (07.3.7) After 2 months (07.7.9) After 1 course (07.5.2) About 1X1011-12 cells Total number of peripheral lymphocytes=~1 X 1010 cells

The number of caner cells >>> CTLs High level of Treg Loss of HLA or antigen The number of caner cells >>> CTLs High level of Treg Loss of HLA or antigen

Specific Spots ratio

Case 5 Specific T Case 5 Specific T-

• cell response

cell response

. 2 . 4 . 6 . 8 1 1 . 2 1 . 4 1 . 6 1 . 8 2 U R L C 1 T T K 5 6 7 K O C 1 B e f

• r

e A f t e r 2 i n j . A f t e r 1 c

• Specific Spots ratio:

target molecule + HLA transfectant/HLA alone transfectant

R/S:1 *p<0.05

* * * * *

Pre-vaccine Post-vaccine 76th day Post-vaccine 104th day

Complete regression was observed after 2 courses of the treatment. A total number of tumor cells 108 Complete regression was observed after 2 courses of the treatment. A total number of tumor cells 108

Case 7 Case 7

Clinical development of cancer vaccine

• --patient population

Clinical development of cancer vaccine

• --patient population
• Metastatic disease
• Advantages

Shorter and faster enrollment (short evaluation period) Small sample size Allows tumor response evaluation

• Metastatic disease
• Advantages

Shorter and faster enrollment (short evaluation period) Small sample size Allows tumor response evaluation

• Minimal or no evidence
• f diseases
• Disadvantages

Longer trial period Large sample size Trial endpoints may be challenging

• Minimal or no evidence
• f diseases
• Disadvantages

Longer trial period Large sample size Trial endpoints may be challenging

Clinical development of cancer vaccine

• --patient population

Clinical development of cancer vaccine

• --patient population
• Metastatic disease
• Disadvantages

Often multiple prior cancer treatment Incompetent immune status (from prior therapies) Inadequate evaluation due to rapid disease progression

• Metastatic disease
• Disadvantages

Often multiple prior cancer treatment Incompetent immune status (from prior therapies) Inadequate evaluation due to rapid disease progression

• Minimal or no evidence
• f diseases
• Advantages

Fewer prior therapies Better in immune status Adequate time for evaluation before disease progression Optimal for the proposed MOA of cancer vaccines

• Minimal or no evidence
• f diseases
• Advantages

Fewer prior therapies Better in immune status Adequate time for evaluation before disease progression Optimal for the proposed MOA of cancer vaccines