Assessment of Biological Plausibility in Cancer

1. Assessment of Biological Plausibility in Cancer Paolo Boffetta IARC, Lyon, France

2. Characteristics of cancer • Pathology • Uncontrolled growth • Ability to spread • Morphological and functional changes • cellular, tissue and organ level • Clinics • Local • compression, infiltration, bleeding, etc. • Systemic • adsorption, excretion, metabolism, etc.

3. Underlying mechanisms • Six general mechanisms • self-sufficiency in growth signals • insensitivity to growth-inhibitory signals • evasion of programmed cell death • limitless replicative potential • sustained angiogenesis • ability to invade tissue and metastasis • Additional considerations • redundancy • multiplicity of pathways Hanahan and Weinberg, 2000

4. Heterogeneity of cancer • Types of cancer • epithelial neoplasms • carcinoma • mesenchimal neoplasms • sarcoma • lymphatic and hematopoietic neoplasms • lymphoma, leukemia • Individual cases of cancer

5. Criteria for assessing biological plausibility of genetic variants • Gene is known to be relevant to one or more steps of the carcinogenic process • Variant is known to affect one or more steps in the carcinogenic process • Consistency with experimental results

6. 1. Gene is known to be relevant to one or more steps of the carcinogenic process • Direct involvement • DNA repair; detoxification • Indirect involvement • tobacco dependence • Complexity of carcinogenic pathways • imperfect knowledge

7. Early stages in carcinogenesis Exogenous carcinogen Endogenous carcinogen Dependence Exposure Activation Detoxification DNA damage DNA repair DNA mutation

8. The power of nicotine addiction

9. Evidence for a genetic role in nicotine addiction Family and twin studies Concordance rates higher in monozygotic than dizygotic twins Estimated heritability rates: 60% - initiation of smoking 70% - maintenance of dependent smoking Smoking is a complex genetic trait with gene-gene and gene- environmental interactions and phenotypic and genotypic heterogeneity

10.  80% NICOTINECOTININE CYP2A6 Nicotine-dependent individuals adjust smoking behavior to maintain nicotine levels Nicotine intake (smoking) Nicotine removal (metabolism)

11. CYP2A6 gene duplications increase nicotine clearance Duplication (*1x2) clearance Inactive alleles (*4)  clearance Nicotine-d2 Clearance (ml/min/kg) Hukkanen et al., 2005

12. Smoking cessation by CYP2A6 genotype(Assessment in a Bupropion treatment trial) p=0.03 Ahluwalia et al. 2002

13. p53 polymorphism and cervical cancer • Interaction between HPV oncogenes and p53 in cervical carcinogenesis • Arg/Pro polymorphism at codon 72 of TP53 • Arg form more vulnerable to binding and degradation by HPV-E6 • Meta-analysis of association studies • LSIL: 18 studies, RR 0.9 (95% CI 0.7-1.1) • HSIL: 22 studies, RR 1.0 (95% CI 0.9-1.1) • Inv Ca: 24 studies, RR 1.1 (95% CI 0.9-1.3) • SqCC: 22 studies, RR 1.5 (95% CI 1.2-1.9) • AC: 4 studies, RR 1.7 (95% CI 1.0-2.7) Koushik et al. 2004

14. 2. Variant is known to affect one or more steps in the carcinogenic process • Lack of functional data for many variants • Function often depends on interaction with other genes and non-genetic factors

15. N-acetylation in bladder cancer • N-acetylation of arylamines and hydrazines • drugs (isoniazid, caffeine) • carcinogens (benzidine, 4-aminobiphenyl, heterocyclic amines) • several NAT2 polymorphisms • the most common result in decreased activity

16. Results of studies of NAT2 slow polymorphism and bladder cancer Garcia-Closas et al., 2005

17. Results of studies of interaction between NAT2 slow polymorphism and tobacco smoking in bladder carcinogenesis Garcia-Closas et al., 2005

18. Gene-environment interaction in lung carcinogenesisITC intake and GST polymorphims • Green cruciferous vegetables • Shown protective against lung cancer • Rich in isothiocyanates (ITCs) • Experimental studies show ITCs inhibit lung carcinogenesis • However, a definite causal association between ITC and decreased risk of any cancer cannot be drawn, due to confounding effect of other nutrients (IARC handbook, 2004) • Glutathione S-transferase (GSTs) • Promote elimination of ITCs • GSTM1 or T1 null: does not have enzyme activity increased ITC levels • The distribution of GSTM1 and T1 genotypes is unlikely to be associated with other dietary sources. • If ITC is the responsible chemopreventive agents, then the protective effect of cruciferous vegetables should be modified by GSTs genotypes

19. Effects of cruciferous vegetables on lung cancer, by GST M1 and T1 polymorphisms Adjusted by age, gender, country, education, and tobacco pack-years. – Brennan et al., 2005

20. 3. Consistency with experimental results • Same gene • response on TP53 LOH mice to suspected human carcinogens • Comparable gene/pathway • Issues • target organ, histologic type • magnitude of effect (background incidence) • limitations of database

21. Performance of TP53 LOH mice by IARC classification French et al., 2004

22. Biological plausibility in the assessment of cancer genetic associations • Support to results of human studies • Generation of specific hypotheses • gene-gene and gene-environment interactions • Contribution to understanding of human carcinogenesis • BRCA1 and BRCA2 as DNA repair genes

23. Conclusions • Imperfect knowledge of carcinogenesis • lack of biological support to ‘real’ associations • generation of a posteriori plausibility • Criteria • strength of underlying biological data • human vs. animal evidence • parsimony

24. CHEK2 and cancer • Cell cycle control gene (located on chromosome 22), the product of which can bind to and activate the p53 protein and other cell cycle proteins • Risk of 1100delC in breast cancer confirmed, although increase in risk is moderate • I157T polymorphism associated with a moderate risk of 6 cancer types from 13 studied • Testing the hypothesis of a role of I157T polymorphism on cancer risk • Application to a multi-site association study from Central Europe Nature Genetics, 2002, 31: 55-59 Am.J.Hum.Genet., 2004; 75: 1131-35

25. CHEK2 I157T variant • rs17879961 • Results in a replacement of A by G at position 26 in exon 6 • Evolutionary highly conserved among different species • Alteration of the binding site in the protein in a way that, in yeast, prevents the homologous protein from activating the yeast homologue of the p53 protein

26. CHEK2 I157T and lung cancer risk

27. CHEK2 I157T and head and neck cancer risk

28. CHEK2 I157T and kidney cancer risk

29. CHEK2 and cancer risk • Opposite effect on different organs • Role of CHEK2 • Quality of functional data • Support from experimental studies