Purpose Ataxia–Telangiectasia Mutated ( ATM ) has been implicated in the risk of several cancers, but establishing a causal relationship is often challenging. Although ATM single-nucleotide polymorphisms have been linked to melanoma, few functional alleles have been identified. Therefore, ATM impact on melanoma predisposition is unclear. Methods From 22 American, Australian, and European sites, we collected 2,104 familial, multiple primary (MPM), and sporadic melanoma cases who underwent ATM genotyping via panel, exome, or genome sequencing, and compared the allele frequency (AF) of selected ATM variants classified as loss-of-function (LOF) and variants of uncertain significance (VUS) between this cohort and the gnomAD non-Finnish European (NFE) data set. Results LOF variants were more represented in our study cohort than in gnomAD NFE, both in all (AF = 0.005 and 0.002, OR = 2.6, 95% CI = 1.56–4.11, p < 0.01), and familial + MPM cases (AF = 0.0054 and 0.002, OR = 2.97, p < 0.01). Similarly, VUS were enriched in all (AF = 0.046 and 0.033, OR = 1.41, 95% CI = 1.6–5.09, p < 0.01) and familial + MPM cases (AF = 0.053 and 0.033, OR = 1.63, p < 0.01). In a case–control comparison of two centers that provided 1,446 controls, LOF and VUS were enriched in familial + MPM cases ( p = 0.027, p = 0.018). Conclusion This study, describing the largest multicenter melanoma cohort investigated for ATM germline variants, supports the role of ATM as a melanoma predisposition gene, with LOF variants suggesting a moderate-risk.

Background: The chromosome 9p21.3 region has been implicated in the pathogenesis of multiple cancers. Methods: We systematically examined up to 203 tagging SNPs of 22 genes on 9p21.3 (19.9–32.8 Mb) in eight case–control studies: thyroid cancer, endometrial cancer (EC), renal cell carcinoma, colorectal cancer (CRC), colorectal adenoma (CA), oesophageal squamous cell carcinoma (ESCC), gastric cardia adenocarcinoma and osteosarcoma (OS). We used logistic regression to perform single SNP analyses for each study separately, adjusting for study-specific covariates. We combined SNP results across studies by fixed-effect meta-analyses and a newly developed subset-based statistical approach (ASSET). Gene-based P -values were obtained by the minP method using the Adaptive Rank Truncated Product program. We adjusted for multiple comparisons by Bonferroni correction. Results: Rs3731239 in cyclin-dependent kinase inhibitors 2A ( CDKN2A) was significantly associated with ESCC ( P =7 × 10 −6 ). The CDKN2A -ESCC association was further supported by gene-based analyses ( P gene =0.0001). In the meta-analyses by ASSET, four SNPs (rs3731239 in CDKN2A , rs615552 and rs573687 in CDKN2B and rs564398 in CDKN2BAS ) showed significant associations with ESCC and EC ( P <2.46 × 10 −4 ). One SNP in MTAP ( methylthioadenosine phosphorylase) (rs7023329) that was previously associated with melanoma and nevi in multiple genome-wide association studies was associated with CRC, CA and OS by ASSET ( P =0.007). Conclusion: Our data indicate that genetic variants in CDKN2A , and possibly nearby genes, may be associated with ESCC and several other tumours, further highlighting the importance of 9p21.3 genetic variants in carcinogenesis.

Abstract Background: E-cadherin is a tumor suppressor gene involved in cell-cell adhesion, epithelial-to-mesenchymal transitions (EMT) and invasion. Loss of E-cadherin expression is strongly associated with lobular breast cancers, which exhibit single cell patterns of infiltration and are often estrogen receptor positive. We sought to determine if relative risk estimates for 19 established breast cancer susceptibility loci were modified by E-cadherin breast tumor tissue expression. Methods: Case-control analyses included up to 1885 invasive breast cancer cases and 2366 age and site matched controls aged 20–74 years from the Polish Breast Cancer Study (PBCS), a population based case-control study conducted in Poland from 2000–2003. Genotyping of the 19 single nucleotide polymorphisms (SNPs) was performed using TaqMan® assays. Tissue expression of E-cadherin was assessed using immunohistochemical (IHC) staining of tissue microarrays and IHC results were scored as the product of percent positive tumor cells × intensity. Tumors having a score of <10 were classified as E-cadherin low and those with a score ≥10 as E-cadherin high. Polytomous logistic regression models adjusted for age and study site were used to estimate odds ratios (OR) and 95% confidence intervals (95% CI) for each breast cancer subtype, defined by E-cadherin expression levels, compared to controls. Case-only data from the PBCS (N = 797) and the Study of Epidemiology and Risk Factors in Cancer Heredity, SEARCH (N = 2155) was used in logistic regression models to test for heterogeneity of SNPs by E-cadherin expression. Results: Three SNPs suggested significant heterogeneity by E-cadherin expression in the PBCS: rs2046210 at 6q25.1(ESR1) [per-allele ORs (95% CI); 1.53 (1.19–1.98) for E-cadherin low tumors and 0.99 (0.86–1.14) for E-cadherin high tumors, P-heterogeneity = 0.002]; rs1045485 at 10p14 (CASP8) [per-allele ORs (95% CI); 0.62 (0.41–0.93) for E-cadherin low tumors and 0.98 (0.81–1.18) for E-cadherin high tumors, P-heterogeneity = 0.04]; and rs11249433 at 1p11.2 (NOTCH2/FCGR1B) [per-allele ORs (95% CI); 1.29 (1.02–1.64) for E-cadherin low tumors and 1.01 (0.88–1.15) for E-cadherin high tumors, P-heterogeneity = 0.06]. Combined case-only analysis of PBCS and SEARCH for these three SNPs showed significant heterogeneity by E-cadherin expression for rs11249433 [Interaction OR (95% CI); 1.19 (1.05–1.36), P-heterogeneity = 0.007] and rs1045485 [Interaction OR (95% CI); 0.69 (0.53–0.90), P-heterogeneity = 0.007]. The association with rs2046210 [Interaction OR (95% CI); 1.12 (0.61–2.03), P-heterogeneity = 0.73] did not remain significant in combined analyses. Conclusion: Our findings provide evidence that associations for breast cancer susceptibility loci vary by E-cadherin tumor tissue expression, which has not been described previously. Specifically, our results suggest that the genetic markers rs11249433 and rs1045485 may preferentially modify risk for tumors with low or absent E-cadherin expression in two independent data sets. Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr P3-08-02.

Abstract Background: CDKN2A is a major susceptibility gene for familial melanoma, but its incomplete penetrance suggests that other factors may modify the effect of this gene in melanoma-prone families. Epigenetic changes involving reduced levels of global DNA methylation in blood have been associated with genomic instability and cancer risk. In addition, genetic background has the potential to influence epigenetic processes through cis- or trans-regulatory variation. Therefore, we hypothesize that the CDKN2A carrier mutation state may be an important factor influencing global DNA methylation in predisposed individuals and that aberrant methylation may be an indicator of modified risk in individuals from melanoma-prone families segregating CDKN2A germline mutations. Methods: We measured the overall level of genomic DNA methylation using bisulfite pyrosequencing at four CpG sites (CpG1, CpG2, CpG3 and CpG4) of the Long Interspersed Nucleotide Element-1 (LINE-1) sequences in peripheral blood mononuclear cells (PBMCs) from 42 CDKN2A mutation positive cases (20 males and 22 females) and 73 controls (29 CDKN2A carrier controls [9 males and 20 females] and 44 non-carrier controls [19 males and 25 females]) in 26 families with CDKN2A mutations. LINE-1 methylation was reported as the average (of all 4 CpGs) and as site-specific percentage methylation. Odds' ratios (OR) and P-values were obtained by comparing melanoma cases to controls using unconditional logistic regression with adjustment for family correlation (using the generalized estimating equations [GEE] approach), age, sex, number of moles and with melanoma status as the outcome variable. Percentage LINE-1 methylation was categorized using tertiles with the highest tertile as the reference. Results: Male gender was significantly associated with higher average LINE-1 methylation (P=<0.001) and methylation at all CpG sites (P=<0.03). In contrast, age, solar injury, dysplastic nevi (DN) and number of moles were not associated with average LINE-1 methylation. Although mean levels of LINE-1 methylation were similar in melanoma cases, carrier controls, and non-carrier controls, after adjusting for family correlation, age, sex and number of moles, low average LINE-1 methylation (tertile 1) was significantly associated with increased risk of melanoma (OR=3.87, 95% confidence interval [CI]: 1.11–13.5, P=0.033, compared to all controls; OR=1.83, 95% CI: 1.09–11.7, P=0.035, compared to carrier controls; and OR=3.43, 95% CI: 1.17–10.0, P= 0.024, compared to non-carrier controls). This association was observed for most CpG sites with the most significant association seen for site 3 and risk of melanoma compared to all controls (OR=5.18, 95% CI: 1.54–17.4, P=0.008). We observed no significant association between average or CpG site-specific LINE-1 methylation and CDKN2A mutation status among controls although the comparison was limited by small sample size. Conclusions: Melanoma cases from families segregating CDKN2A germline mutations have significantly lower LINE-1 global methylation in their PBMCs compared to controls. Changes in LINE-1 methylation in PBMCs from CDKN2A mutation positive cases may reflect a potential genetic background influence on global DNA methylation in these individuals. Further studies are needed to clarify these preliminary observations. Citation Information: Cancer Prev Res 2011;4(10 Suppl):A84.

BACKGROUND:Carrying the cyclin-dependent kinase inhibitor 2A (CDKN2A) germline mutations is associated with a high risk for melanoma. Penetrance of CDKN2A mutations is modified by pigmentation characteristics, nevus phenotypes, and some variants of the melanocortin-1 receptor gene (MC1R), which is known to have a role in the pigmentation process. However, investigation of the associations of both MC1R variants and host phenotypes with melanoma risk has been limited. METHODS:We included 815 CDKN2A mutation carriers (473 affected, and 342 unaffected, with melanoma) from 186 families from 15 centers in Europe, North America, and Australia who participated in the Melanoma Genetics Consortium. In this family-based study, we assessed the associations of the four most frequent MC1R variants (V60L, V92M, R151C, and R160W) and the number of variants (1, ≥2 variants), alone or jointly with the host phenotypes (hair color, propensity to sunburn, and number of nevi), with melanoma risk in CDKN2A mutation carriers. These associations were estimated and tested using generalized estimating equations. All statistical tests were two-sided. RESULTS:Carrying any one of the four most frequent MC1R variants (V60L, V92M, R151C, R160W) in CDKN2A mutation carriers was associated with a statistically significantly increased risk for melanoma across all continents (1.24 × 10(-6) ≤ P ≤ .0007). A consistent pattern of increase in melanoma risk was also associated with increase in number of MC1R variants. The risk of melanoma associated with at least two MC1R variants was 2.6-fold higher than the risk associated with only one variant (odds ratio = 5.83 [95% confidence interval = 3.60 to 9.46] vs 2.25 [95% confidence interval = 1.44 to 3.52]; P(trend) = 1.86 × 10(-8)). The joint analysis of MC1R variants and host phenotypes showed statistically significant associations of melanoma risk, together with MC1R variants (.0001 ≤ P ≤ .04), hair color (.006 ≤ P ≤ .06), and number of nevi (6.9 × 10(-6) ≤ P ≤ .02). CONCLUSION:Results show that MC1R variants, hair color, and number of nevi were jointly associated with melanoma risk in CDKN2A mutation carriers. This joint association may have important consequences for risk assessments in familial settings.

Previous prospective studies have found an association between prolactin (PRL) levels and increased risk of breast cancer. Using data from a population-based breast cancer case–control study conducted in two cities in Poland (2000–2003), we examined the association of PRL levels with breast cancer risk factors among controls and with tumour characteristics among the cases. We analysed PRL serum levels among 773 controls without breast cancer matched on age and residence to 776 invasive breast cancer cases with available pretreatment serum. Tumours were centrally reviewed and prepared as tissue microarrays for immunohistochemical analysis. Breast cancer risk factors, assessed by interview, were related to serum PRL levels among controls using analysis of variance. Mean serum PRL levels by tumour characteristics are reported. These associations also were evaluated using polytomous logistic regression. Prolactin levels were associated with nulliparity in premenopausal (P=0.05) but not in postmenopausal women. Associations in postmenopausal women included an inverse association with increasing body mass index (P=0.0008) and direct association with use of recent/current hormone therapy (P=0.0006). In case-only analyses, higher PRL levels were more strongly associated with lobular compared with ductal carcinoma among postmenopausal women (P=0.02). Levels were not different by tumour size, grade, node involvement or oestrogen receptor, progesterone receptor, or human epidermal growth factor receptor 2 status. Our analysis demonstrates that PRL levels are higher among premenopausal nulliparous as compared with parous women. Among postmenopausal women, levels were higher among hormone users and lower among obese women. These results may have value in understanding the mechanisms underlying several breast cancer risk factor associations.