Pirie, Ailith

Abstract Background: While numerous susceptibility loci for epithelial ovarian cancer (EOC) have been identified, few associations have been reported with overall survival. In the absence of common prognostic genetic markers, we hypothesize that rare coding variants may be associated with overall EOC survival and assessed their contribution in two exome-based genotyping projects of the Ovarian Cancer Association Consortium (OCAC). Methods: The primary patient set (Set 1) included 14 independent EOC studies (4,293 patients) and 227,892 variants, and a secondary patient set (Set 2) included six additional EOC studies (1,744 patients) and 114,620 variants. Because power to detect rare variants individually is reduced, gene-level tests were conducted. Sets were analyzed separately at individual variants and by gene, and then combined with meta-analyses (73,203 variants and 13,163 genes overlapped). Results: No individual variant reached genome-wide statistical significance. A SNP previously implicated to be associated with EOC risk and, to a lesser extent, survival, rs8170, showed the strongest evidence of association with survival and similar effect size estimates across sets (Pmeta = 1.1E−6, HRSet1 = 1.17, HRSet2 = 1.14). Rare variants in ATG2B, an autophagy gene important for apoptosis, were significantly associated with survival after multiple testing correction (Pmeta = 1.1E−6; Pcorrected = 0.01). Conclusions: Common variant rs8170 and rare variants in ATG2B may be associated with EOC overall survival, although further study is needed. Impact: This study represents the first exome-wide association study of EOC survival to include rare variant analyses, and suggests that complementary single variant and gene-level analyses in large studies are needed to identify rare variants that warrant follow-up study. Cancer Epidemiol Biomarkers Prev; 25(3); 446–54. ©2016 AACR.

Abstract Many germline genetic variants have been found to be associated with susceptibility to epithelial ovarian cancer (EOC), but currently few associations have been identified for EOC outcomes such as overall survival. In the absence of evidence for common germline markers, it is possible that rare variants not captured in GWAS arrays may be associated with overall survival. We hypothesize that rare variants, either individually or combined across a gene, may be associated with overall survival in EOC. We utilized data from two genotyping projects of the Ovarian Cancer Association Consortium (OCAC) that used commercial exome-based arrays. The primary sample, genotyped on the Affymetrix Axiom Exome Array at 227,892 standard and custom variants, consisted of 14 independent EOC studies (4293 cases; 2257 deaths). The secondary sample, genotyped on the Illumina Infinium HumanExome BeadChip at 114,620 variants, consisted of six additional EOC studies (1744 cases; 1027 deaths). Both sets were analyzed using Cox proportional hazards regression to determine the association of each variant with overall survival time. Meta-analysis was conducted across samples at 73,203 overlapping variants. Gene-level tests of over 18,000 genes were also conducted using burden tests and Sequencing Kernel Association Tests (SKAT) adapted for Cox proportional hazards models. Gene-level tests were conducted separately for the primary and secondary cases and then were meta-analyzed. Models were adjusted for age, site, and three principle components. Six individual variants had meta-analysis p-values < 1.0E-4, five of which were rare (MAF < 0.003). Common variant rs8170 (MAF = 0.2) in BABAM1 encoding BRISC and BRCA1 A complex member 1 - previously reported to be associated with EOC risk_showed evidence of association with overall survival (Pmeta = 1.5E-5) and consistent estimates of effect size across both samples (HR1 = 1.18, HR2 = 1.19). In imputed genotype data from over 18,000 subjects from OCAC genotyped on the COGS array, this association reaches genome-wide significance (HR = 1.15, P = 4.7E-9). Five genes had gene-level meta-analysis p-values < 1.0E-4, including top ranked ATG2B (p = 1.1E-6), an autophagy gene important for apoptosis; zinc finger protein PEG3 (p = 1.8E-5), playing a role in autophagy and down-regulated in EOC; helicase-like transcription factor HLTF (p = 1.9E-5), involved in post-replication DNA repair; mitochondrial fission regulator 1-like gene FAM54B (p = 3.1E-5); and T-cell surface glycoprotein CD1E (p = 8.8E-5). With over 6000 EOC cases in the primary and secondary samples, this is the largest genome-wide assessment of rare variant genetic associations with EOC overall survival. Variant rs8170 in BABAM1, a necessary component of the BRCA1 complex, is associated with overall survival. Single variant and gene-level analyses provide complementary approaches, suggesting additional candidates that warrant follow-up study. Citation Format: Stacey J. Winham, Brooke L. Fridley, Melissa C. Larson, Zachary Fogarty, Andrew Berchuck, Yian Ann Chen, Hui-Yi Lin, Georgia Chenevix-Trench, Jenny Permuth-Wey, Thomas A. Sellers, Ailith Pirie, Ellen L. Goode, Ovarian Cancer Association Consortium. Investigation of exome variants associated with overall survival in ovarian cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4636. doi:10.1158/1538-7445.AM2015-4636

Abstract Background: Common genetic variants identified by genome-wide association studies and follow-up genotyping initiatives only explain a small proportion of the heritability of epithelial ovarian cancer (EOC). We hypothesized that some of the missing heritability may be explained by rare (minor allele frequency (MAF) <0.5%) and low-frequency (MAF 0.5-5%) protein-coding variants that are not well captured by genotyping arrays that focus on common variants. Methods: To examine the association between uncommon variants and EOC susceptibility, we genotyped 7,060 EOC cases and 6,712 controls from the Ovarian Cancer Association Consortium using the Affymetrix Axiom Exome Genotyping Array. This platform contains 300,785 putative functional coding variants selected from exome sequences of 12,000+ individuals representing multiple ethnicities and complex traits. After performing quality control (QC), we used unconditional logistic regression treating the number of minor alleles carried as an ordinal variable (log-additive model) to evaluate SNP-EOC risk associations. Genotype cluster plots were visually inspected for top-ranking variants to determine assay success, and allele frequencies of samples from the 1000 Genomes Project and other cohorts were evaluated for QC purposes. Results: A total of 13,318 samples (97%) passed QC, and 12,779 eligible samples (6,293 invasive serous and endometrioid cases and 6,486 controls) were included in the analyses. 280,737 (93%) of the 300,785 attempted markers were successfully genotyped and had call rates >90%. After removing 31,436 monomorphic markers, the 249,301 remaining markers were included in association analyses. We confirmed variants at previously reported EOC susceptibility loci (9p22, 3q25, 17cen-q21.3, and 8q24), with p-values between 10-5 and 10-11. Several correlated low-frequency non-synonymous variants (MAF=5%) residing in a novel EOC susceptibility gene at 3q25.31 were risk-associated. The most significant variant at 3q25.31 had an OR=1.36 and p = 5.58x10-8. Also noteworthy is a rare variant at 15q15.1-q21.1 (MAF=0.5%) that introduces a stop codon in a gene belonging to the tyrosine kinase family (OR=0.39, CI=0.26-0.58, p = 4.52x10-6). We also identified a more common synonymous coding variant in a phosphatase at 6q21.3 that was associated with increased risk (OR=1.22, CI=1.13-1.33, p = 1.84x10-6, MAF=9.6%). Population stratification and gene-based analyses are underway to further examine the impact of the newly-identified variants on EOC risk. A meta-analysis to include data from an additional 2104 EOC cases and 2516 controls genotyped with Illumina's HumanExome Beadchip rare variant array is also underway. Functional analysis of the most promising variants will follow. Conclusions: Preliminary data from this large-scale study reveals novel rare and low-frequency variants that may influence susceptibility to epithelial ovarian cancer. Citation Format: Jennifer Permuth-Wey, Y. Ann Chen, Zhihua Chen, Andrew Berchuck, Georgia Chenevix-Trench, Jennifer Doherty, Simon Gayther, Ellen L. Goode, Edwin Iversen, Alvaro N.A. Monteiro, Leigh Pearce, Paul D.P. Pharoah, Catherine M. Phelan, Ailith Pirie, Susan Ramus, Mary Ann Rossing, Joellen M. Schildkraut, Thomas A. Sellers, on behalf of the Ovarian Cancer Association Consortium. Exome genotyping array identifies rare and low-frequency variants that may be associated with ovarian cancer risk. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 946. doi:10.1158/1538-7445.AM2014-946