09 26 2006 Cancer As Genetic Diseases

EXOGENOUS Chemical X-ray U.V. ENDOGENOUS Oxygen Replication Error Chemical Alterations

MALIGNANT PHENOTYPE Oncogenes Tumor Suppressor Gene

Growth Apoptosis Differentiation Invasion Metastasis

Francis Peyton Rous, 1879-1970.

Incidence of breast cancer for 1983-7 in women aged 20—54 for Asian-Americans living in America and in the countries of origin Community, ethnicity

Age-standardized

Cumulative(%)

San-Francisco-Oakland-LA, Hawaii White

91.8

3.45

Chinese

53.7

2.03

Japanese

69.0

2.59

Filipino

72.5

2.72

China, Shanghai/Tianjin

27.5/27.4

1.03/1.03

Hong Kong

37.7

1.41

Singapore

41.7

1.56

Japan, Miyagi/Osaka

40.9/28.9

1.53/1.08

Philippines, Manila/Rizal

54.7/45.7

2.06/1.72

>44,000twins Site of Cancer No of concordant No. of discordant RR(95%CI) Concordance Lung MZ DZ Prostate MZ DZ Cervix MZ DZ

affected pairs

pairs

15 24

233 436

7.7(4.4-13.6) 6.7(4.3-10.5)

0.11 0.10

40 20

299 584

12.3(8.4-18.1) 3.1(1.9-4.9)

0.21 0.06

1 3

107 201

2.9(0.4-21.4) 4.5(1.4-14.4)

0.02 0.03

RR:The relative risk of cancer for persons whose twins had a particular type of cancer, as compared with those whose twins did not Concordance: the proportion of all persons with cancer whose twins had cancer at the same site

N Engl J Med 2000;343:78

Li-Fraumeni Syndrome (cased by germline mutation of p53 or hCHK2) Early-onset Rare tumor Bilateral pattern

Three generations of familial retinoblastoma. Red denotes an affected individual, a square indicates a male, a circle a female.

Retinoblastoma (Rb)

Familial adenomatous polyposis (FAP) caused by mutant APC

Figure 7.22 The Biology of Cancer (© Garland Science 2007)

Identification of a disease susceptibility gene

Linkage analysis and Recombination

Meiosis

Crossing-over (Recombination)

Linkage analysis using microsatellite markers

M++ Others Cases

A

0

Non-cases

0

D

LOD score

Association

E-cadherin germline mutations in familial gastric cancer

Nature 1998;392:402-5

Nature 1998;392:402-5

Nature 1998;392:402-5

Science 1997;278:1043

Cancer Cell 2002;2:103-112

Cancer results from a series of molecular alterations Mismatched Repair Gene Mutation 5q

DNA

12p

18q

17p LOH and DCC Mutation

LOH and APC Mutation

Alteration in DNA Methylation

K-ras Mutation

LOH and p53 Mutation

Science 1997;278:1043

Am J Pathol 1998;153:333

Cell 2004;116:235

Given the fact that Familial cancer only accounts for a very small fraction (<10%) to total cancer, what is the importance to localize cancer susceptibility genes by linkage analysis?

Alfred G. Knudson, Jr.

Retinoblastoma Sporadic form

Inherited form

Unilateral

Bilateral

One cancer per person

3 primary cancers

No family history

Family history

Rare, one in 30,000

Recurrence after surgical intervention

Two-hit model of tumor suppressor gene

The two-mutation model of retinoblastoma development.

EXOGENOUS Chemical X-ray U.V. ENDOGENOUS Oxygen Replication Error Chemical Alterations

Two-hit model

Oncogenes Tumor Suppressor Gene

Susceptibility

MALIGNANT PHENOTYPE Growth Apoptosis Differentiation Invasion Metastasis

RR is the cancer risk in a carrier versus a noncarrier of the allele

Nature 2001;411:336; Nature Genet 2002;31:33

Averaged across all ages, the risk of breast cancer to the sister, mother or daughter of a case is increased about twofold.

RR is the cancer risk in a carrier versus a noncarrier of the allele

Nature 2001;411:336; Nature Genet 2002;31:33

familial breast cancers that cannot been explained by BRCA1 or BRCA2 are caused by the combined effect of a large number of codominant alleles, each of which is associated with a small increase in risk

Common Disease-Common Variant Hypothesis Mutation Familial syndrome (high penetrance)

Linkage analysis

Gene Polymorphism Sporadic disease (low penetrance)

Association study

Double Strand Break Checkpoint/Repair

Why is there no evidence available to suggest the contribution of the nonhomologous end joining (NHEJ) pathway, which has been known to be critical for mammalian cells to repair DSB, in breast cancer development or any cancer predisposition disorder?

Knockout mice of NHEJ genes

Ku70, Ku80, DNA-PKcs

LigaseIV, XRCC4

B-cell/T-cell tumor Growth retardation Embryotic lethality

Ataxia telangiectasia (AT, ATM)

Cancer predisposition

Nijmegen breakage syndrome (NBS, NBS1) NBS-like (Ligase IV)

Developmental delay Immunodeficiency

Nijmegen breakage syndrome

Germ-line mutated NBS1 results in NBS

Fu et al., Cancer Res., 2003

The SNPs genotyped for the genes of NHEJ 30 SNPs (6 SNPs for each gene; all SNPs were “LD”-type markers) No polymorphism was identified in 18 SNPs in initial screening (192 Cs+192 Cn) 12 SNPs (3 for Ku70, 2 for Ku80, 1 for DNA-PKcs, 3 for LigaseIV, 3 for XRCC4) were in data analysis

• Cancer risk associated with “lowpenetrance alleles” • Joint effect of genes involved in the NHEJ pathway • Gene (NHEJ)-estrogen interaction

Gene

Case(%)

Controls(%) OR(95%CI) aOR(95%CI)

Ku70 low-risk genotype 122(26.8) 271(36.9) high-risk genotype 334(73.2) 464(63.1)   XRCC4 low risk genotype 173(35.9) 325(44.0) high-risk genotype 309(64.1) 414(56.0)

1.00

1.00

1.60 1.68 (1.24-2.07) (1.28-2.20)

1.00

1.00

1.40 1.45 (1.11-1.78) (1.13-1.85) Fu et al., Cancer Res.,

The joint effect of susceptible genotypes of the genes in the NHEJ is significantly associated with breast cancer risk - Combination of individual effects NHEJ pathway: Ku70, Ku80 DNA-PKcs, LigaseIV, XRCC4 No. of susceptible genotypes 0 1 2 3 4 5

aOR for trend (95%CI) 1.46 (1.19-1.80)

p value 0.0003

The women harboring a higher number of susceptible genotypes of the NHEJ genes show a higher risk of developing breast cancer, and this risk is strongly modified by the history of pregnancy No. of susceptible NHEJ genotypes Pregnancy

aOR(95%CI)

<=1

Yes

1.00(ref.)

>1

Yes

2.42(1.61-3.65)

<=1

No

2.75(0.92-8.26)

>1

No

5.29(2.67-10.5) Fu et al., Cancer Res., 2003

• High-penetrance tumor suppressor genes may display their tumorigenic contribution via the presence of low-penetrance allele. • The joint effect of low-penetrance alleles of the genes participating in a same functional pathway is associated with cancer development. • Tumorigenic effect of low-penetrance alleles depends on their interaction with environmental exposure

The genotype of BRCA1 modifies breast cancer risk conferred by a high number of high-risk genotypes of NHEJ genes No. high-risk Years before genotype FFTP BRCA1 genotype Wt/Wt 0-2 <12 0-2 >=12 3-5 <12 3-5 >=12 Wt/Vt, Vt/Vt 0-2 <12 0-2 >=12 3-5 <12 3-5 >=12

aOR(95%CI) 1.00(ref.) 1.06(0.60-1.87) 0.77(0.45-1.32) 1.07(0.55-2.09) 1.00(ref.) 1.43(0.92-2.22) 1.30(0.85-1.99) 2.55(1.50-4.34)

In vitro end-joining assay shows higher end-joining capacity in BRCA1 competent cell (MCF7), as compared to BRCA1-null cell (HCC1937) MCF7

linearized & de-P plasmids P label HCC1937 Cell extract

End joining

autoradiography

Bau et al., 2004

30

BRCA1 status is associated with precise EJ capacity but is not associated with overall EJ capacity

Relative End-Joining Effciency (% of uncut group)

MCF-7

25

(BRCA1+)

HCC1937

(BRCA1-)

20

15

P<0.05

Overall EJ Precise EJ

10

5

0

Bau et al., 2004

BRCA1 deficient cell (HCC1937) regains EJ capacity by reintroducing wt-BRCA1, and BRCA1 proficient cell (MCF7) loses EJ capacity by siRNA inhibition of BRCA1 12

16

10

BRCA1 0 4 8 12 16 ug

Precise End-Joining Efficiency (% of uncut control)

Precise End-Joining Efficiency (% of uncut control)

HCC1937

8

6

4

2

MCF7

14

siRNA

12

- 20nM

10 8 6 4 2

0

0

4

8

12

BRCA1 plasmid, g

16

0

0

5

10

15

BRCA1 siRNA, nM

20

Cancer as a Genetic Disease High-penetrance effect of High-penetrance gene Low-penetrance effect of High-penetrance gene Low-penetrance effect of Low-penetrance gene

Isolation of EMSY and Interaction Studies

EMSY Relocalizes in Response to DNA Damage

EMSY Amplification in Breast Cancer

JNCI 2005;97:1302

Nat Med 2004;10:127

Mutation Diseasecausing Familial High-penetrance

Polymorphism Epigenetic mechanisms Diseasemodifying Sporadic Low-penetrance and modified by exposure