Our study has identified 41 methylation marks associated with familial prostate cancer, and 9 of these marks (near VTRNA2-1) also have nominally significant associations with aggressive prostate cancer risk in the general population. Note that we would not expect all 41 methylation marks to be associated with risk in the population, e.g. BRCA1 is usually not detected by genome-wide association studies. Also, the magnitude of risk in the population-based and familial settings could differ greatly, due to rare mQTLs or epimutations causing changes in methylation that are many times the population standard deviation.

Nine of the 41 heritable methylation marks associated with prostate cancer risk are in the imprinted VTRNA2-1 region, with a loss of imprinting in this region consistent with a Mendelian pattern of inheritance; see also [3]. Imprinted regions are often associated with tissue growth, and a loss of imprinting can be linked to tumorigenesis, as is well-described for the H19/IGF2 region. VTRNA2-1 has tumour suppressor gene properties, as it regulates cell growth via inhibition of protein kinase RNA-activated (PKR). Down regulation of VTRNA2-1 in a variety of tumours and cancer cell lines has been well documented and associated with promoter CpG hypermethylation. As we found previously using the same datasets, VTRNA2-1 methylation marks are associated with aggressive prostate cancer in the population [5] and are largely independent of the underlying genetic sequence [6]. We and others have described the VTRNA2-1 locus as a metastable epiallele because the loss of imprinting in this region occurs systemically, can be modulated by the periconceptional environment and persists through adulthood [10].

Seven of the heritable methylation marks associated with prostate cancer risk are located at peptidase M20 domain containing 1 (PM20D1), a known methylation and expression quantitative trait locus associated with risk of Alzheimer’s disease. These and the other annotated and unannotated CpGs identified in this study require further research to understand the biological explanation for their association with heritable prostate cancer risk. Non-genetic causes of heritability could not be investigated in the current study, but it is possible that familial environmental and lifestyle factors play a role in determining DNA methylation at these loci.

Despite having a modest sample size, we were able to identify many heritable, cancer-associated methylation marks. Excluding non-heritable methylation marks before testing for association with cancer excludes many marks that cannot cause familial cancer, while retaining any that can. Our method of excluding non-heritable marks before testing for association with disease is therefore a very powerful way of enriching the candidate set of methylation marks for those that could cause familial disease. Further discussion of the methodology can be found in Joo et al., [3].

Heritable methylation marks can mimic the effects of genetic variants, so identifying them is similar in many ways to finding genetic loci that are associated with cancer. As for genetic loci, these marks can implicate new biological mechanisms and therefore shed light on the processes of prostate cancer initiation and progression. These methylation marks could also be used in risk-prediction algorithms to give more precise estimates of a person’s risk of prostate cancer, and so provide more tailored screening.

Our study has several strengths, including its use of an innovative method to identify heritable methylation marks, its method of enriching the candidate set of methylation marks for those that could cause familial disease, and its use of a cohort study to further investigate the findings from the family-based phase. The main weakness of the study was that some of the marks from the family-based phase could not be investigated in the population-based phase, due to the use of different arrays in the two phases. Our modest sample size is also a weakness, though this is offset by the enrichment step described above.

In summary, our study has identified 41 heritable methylation marks that are associated with prostate cancer risk in the context of multiple-case families, with 9 of these marks near VTRNA2-1 likely to be associated with aggressive prostate cancer in the general population.