Background: The overall worldwide incidence of cutaneous malignant melanoma (CMM) is increasing faster than that of any other cancer. In the lack of early detection, CMM shows high fatality. Taken together, these data show the urgent need for new approaches to risk prediction and targeted prevention. Approximately 10-13% of CMM cases are clustered in families, whereas the remaining are sporadic CMMs. Germline mutations (i.e., p16INK4a, p14ARF, CDK4) or variants (i.e., MC1R, hTERT) determining increased CMM risks have been identified in both familial and sporadic cases. Genetic determinants, however, account only for very small proportions of CMMs.
Specific Aims: We hypothesize that epigenetic mutations (epimutations), which can arise in the germline due to DNA methylation and cause soma-wide altered expression, might contribute to the risk of familial and sporadic CMM unaccounted for by genetic mutations. Aim 1: Identify germline epimutations in the p16INK4a, p14ARF, CDK4 genes in familial and sporadic cases that do not carry high-penetrance genetic mutations in the same loci Aim 2: Identify germline epimutations in MCR1 and hTERT and correlate them with: i) objective and instrumental measures of individual pigmentary phenotypes (for MC1R); ii) telomere length (for hTERT); iii) CMM risk (MC1R, hTERT). Aim 3: Determine whether carriers of epimutations have greater risks from established genetic, host, and environmental determinants of CMM.
Study Population and Approach: We will have the unique opportunity to pursue these specific aims in two related investigations of familial CMM (family-based study of 55 families with 2 to 5 CMM patients, including 84 CMM cases and 203 first-degree relatives) and sporadic CMM (casecontrol study of 183 CMM cases and 179 frequency-matched controls) in Northeastern Italy. The two studies have extensive genetic data and information of host-related and environmental risk factors. To analyze DNA methylation, we will bisulfite-sequence the candidate loci by highprecision Pyrosequencing analysis.
Study Team: Dr. Bollati, a junior investigator with a demonstrated productivity in cancer epigenetics, will lead a multidisciplinary team with extensive expertise in the relevant areas of epigenetics and DNA methylation, melanoma etiology, molecular epidemiology, and biostatistics.
Strengths and Expected Results a) Feasibility of the proposed aims and cost-effective design thanks to existing studies of familial and sporadic CMM. b) Family-based studies have been traditionally used to detect uncommon high-penetrance mutations, whereas case-control studies are more efficient for more common, low-penetrance mutations. No information is available on the penetrance of potential CMM-related epimutations. The use of both study designs will provide us with a unique opportunity to detect epimutations linked with CMM risk across the risk and frequency spectra. c) The two study populations have very low frequency of CMM-related mutations. d) The loci proposed for analysis have functions that have been shown to be causally associated with CMM risk and represent prime candidates for CMM epimutations. e) Extensive experience for DNA methylation analysis in the high precision, quantitative bisulfite- PCR Pyrosequencing technique that we will employ in this proposed study. f) Potential to uncover epigenetic determinants of CMM risk and new approaches to individualized risk prediction and prevention.