Environmental Exposure and DNA Damage

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Peripheral blood DNA methylation may be associated with breast cancer, but studies of candidate genes and global and genome-wide DNA methylation have been inconsistent. We performed an epigenome-wide study using Infinium HumanMethylation450 BeadChips with prospectively collected blood DNA samples from the Sister Study (1552 cases, 1224 subcohort). We identified 9601 CpG markers associated with invasive breast cancer (false discovery rate = q < 0.01), with 510 meeting a strict Bonferroni correction threshold (10-7). A total of 2095 of these CpGs replicated in the independent EPIC-Italy dataset, including 144 meeting the Bonferroni threshold. Most dmCpGs showed lower methylation in invasive cases. In case-only analysis, methylation was statistically significantly associated with time to diagnosis for 892 (42.6%) of the dmCpGs. Analyses based on genetic association suggest that methylation differences are likely a consequence rather than a cause of breast cancer. Pathway analysis shows enrichment of breast cancer-related gene pathways, and dmCpGs are overrepresented in known breast cancer susceptibility genes. Our findings suggest that the DNA methylation profile of blood starts to change in response to invasive breast cancer years before the tumor is clinically detected. Cannabis use is highly prevalent and is associated with adverse and beneficial effects. DNA methylation (DNAm) is candidate marker for cannabis exposure, yet no blood-based epigenome-wide association studies (EWAS) exist. We conducted an EWAS of lifetime cannabis use using blood-based DNAm data from this same case-cohort study of breast cancer. We identified and replicated an association with lifetime cannabis use at one CpG site in the gene CEMIP (p = 3.3 10-8 ). We found no overlap between published blood-based cis-meQTLs of this CpG and reported lifetime cannabis use-associated single nucleotide polymorphism (SNPs; p < .05), suggesting that the observed DNAm difference was driven by cannabis exposure. We also developed a multi-CpG classifier of lifetime cannabis use using penalized regression of top EWAS CpGs. The resulting 50-CpG classifier produced an area under the curve (AUC) = 0.74 (p = 2.00 10-5 ) in the discovery sample and AUC = 0.54 p = 2.87 10-2 ) in the replication sample. Our EWAS findings provide evidence that blood-based DNAm is associated with lifetime cannabis use. Our work makes wide use of Illumina BeadChips. Several studies have reported that many probes on these arrays have poor reliability which is measured by intra-class correlation coefficients (ICC). We describe the characteristics of ICC across the genome, within and between studies, and across different array platforms. We find that with raw data only 22.5% of the CpGs on 450 K array have 'acceptable' ICCs (>0.5). Data preprocessing steps, such as background correction and dye bias correction, can reduce technical noise and improve the percentage to 38.5%. Similar to previous studies, we found that ICC is associated with CpG methylation level such that 83% of CpGs with intermediate methylation (0.1< beta-value <0.9) have acceptable ICCs, whereas only 21% of CpGs with low or high methylation (beta-value <0.1 or >0.9) have acceptable ICCs. ICC is also correlated with CpG methylation variance; after mutual adjustment for beta-value and variance, only variance remains correlated. Many CpGs with poor ICCs (<0.5) are located in biologically important regulatory regions, including gene promoters and CpG islands. Poor ICC at these sites appears to be a consequence of low biologic variation among individuals rather than increased technical measurement variation. ICCs quality classifications are highly concordant across different array platforms and across different studies. CpGs with acceptable ICC have higher study power and are more commonly reported in published epigenome-wide studies. We find that ICC can be reliably estimated with 30 pairs of duplicate samples.