Genetic Susceptibility And The Environment In Cancer Risk

National Institute Of Environmental Health Sciences

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Abstract

Isolated orofacial clefts are among the most common congenital birth defects. Although the underlying biological mechanisms remain largely unknown, clefts are thought to be complex disorders influenced by genetic, environmental, and potentially epigenetic factors. In blood samples from 2- to 3-day-old infants (n=747) collected in a nationwide population-based study of orofacial clefts in Norway, we measured DNA methylation profiles for more than 450,000 CpGs and then conducted epigenome-wide association analyses (EWAS). We tested methylation profile difference at each CpG between controls (n=436) and each of the cleft subtypes (92 cleft lip only, CLO; 84 cleft palate only, CPO; 132 cleft lip and palate, CLP). We also compared controls to various combinations of case groups and compared case subtypes to each other. Finally, using the EWAS results, we searched for larger differentially methylated regions (DMRs) associated with orofacial clefts. In EWAS comparing controls to individual cleft subtypes, we found no significant associations at a Bonferroni P value threshold of 10-7. After pooling case groups, we found two significantly differentially methylated CpGs: cg09696939 near gene BICC1 is associated with CLO+CLP (P=9.5810-8); cg26985354 in gene CLASRP is associated with CPO+CLP (P=7.3810-8). In DMR analysis, we identified a total of 56 significant regions when comparing controls to individual cleft subtypes (10 for CLO, 6 for CPO, 41 for CLP). Only one DMR is shared among the three cleft groups. In combined case group analysis, we found 26 DMRs for CLP+CLO, 31 for CLP+CPO, and 37 when all subtypes are combined. Finally, in case-case comparisons of subtypes, we identified 10 DMRs when comparing CLP to CPO, 9 in CLP compared to CLO, and 13 in CLP compared to CPO. We identified two individual CpGs and multiple DMRs that differ between controls and cleft case subtypes. Although we find some evidence for the possible role of DNA methylation in etiology of orofacial clefts, our study does not support previous reports of widespread differences in blood DNA methylation between babies with and without facial clefts. Peripheral blood DNA methylation may be associated with breast cancer, but studies of candidate genes, 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 sub-cohort). Differentially methylated CpGs were identified using case-cohort proportional hazard models and replicated using deposited data from EPIC-Italy (n=329). Correlation between methylation and time-to-diagnosis was examined using robust linear regression. Causal/consequential relationships of methylation to breast cancer was examined by Mendelian randomization using OncoArray 500K SNP data. All statistical tests were two-sided. We identified 9601 CpG markers associated with invasive breast cancer (false discovery rate FDR q<0.01), with 510 meeting a strict Bonferroni correction threshold (10-7). 2095 of these CpGs replicated in the independent EPIC-Italy dataset, including 144 meeting the Bonferroni threshold. Sister Study women who developed ductal carcinoma in situ had methylation similar to non-cases. Most (1501; 71.6%) differentially methylated CpGs (dmCpGs) showed lower methylation in invasive cases. In case-only analysis methylation was statistically significantly associated (FDR q<0.05) 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 DNA methylation profile of blood starts to change in response to invasive breast cancer years before the tumor is clinically detected.