Permuth‐Wey, Jenny

Telomeres cap chromosome ends, protecting them from degradation, double‐strand breaks, and end‐to‐end fusions. Telomeres are maintained by telomerase, a reverse transcriptase encoded by TERT, and an RNA template encoded by TERC. Loci in the TERT and adjoining CLPTM1L region are associated with risk of multiple cancers. We therefore investigated associations between variants in 22 telomere structure and maintenance gene regions and colorectal, breast, prostate, ovarian, and lung cancer risk. We performed subset‐based meta‐analyses of 204,993 directly‐measured and imputed SNPs among 61,851 cancer cases and 74,457 controls of European descent. Independent associations for SNP minor alleles were identified using sequential conditional analysis (with gene‐level p value cutoffs ≤3.08 × 10−5). Of the thirteen independent SNPs observed to be associated with cancer risk, novel findings were observed for seven loci. Across the DCLRE1B region, rs974494 and rs12144215 were inversely associated with prostate and lung cancers, and colorectal, breast, and prostate cancers, respectively. Across the TERC region, rs75316749 was positively associated with colorectal, breast, ovarian, and lung cancers. Across the DCLRE1B region, rs974404 and rs12144215 were inversely associated with prostate and lung cancers, and colorectal, breast, and prostate cancers, respectively. Near POT1, rs116895242 was inversely associated with colorectal, ovarian, and lung cancers, and RTEL1 rs34978822 was inversely associated with prostate and lung cancers. The complex association patterns in telomere‐related genes across cancer types may provide insight into mechanisms through which telomere dysfunction in different tissues influences cancer risk.

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 DNA copy number variations (CNVs) are a significant and ubiquitous source of human genetic variation. However, the influence of CNVs on cancer susceptibility remains poorly understood. Out of 83 genome-wide association studies (GWAS) on cancer to date, only a few studies found significant associations on germline CNVs and cancer. Here we analyzed data from our ongoing two-stage GWAS of four North American case-control studies to test the hypothesis that CNVs in germline DNA from peripheral blood lymphocytes may serve as risk factors for epithelial ovarian cancer (EOC). To reduce disease heterogeneity, we focused on the subset of cases with serous histology. The analysis was therefore based on 942 serous ovarian cancer patients and 1,682 healthy controls who were genotyped using the Illumina 610K quad array. Subjects with extreme noise and genomic waviness in log R ratio (LR) were excluded prior to segmentation; principal component analysis was performed to adjust for batch effects. CNV segmentation was performed on LR data from 22 autosomes using circular binary segmentation embedded in the Copy Number Analysis Module from Golden Helix SNP Variation Suite version 7. Copy number segment covariates were discretized based on the thresholds that signify a transition between copy number states (deletion/no deletion; duplication/no duplication) after the segmentation. Unconditional logistic regression on a log-additive model was used to evaluate the association between copy number states and serous EOC risk after adjusting for study sites. By comparing single marker copy number states at 388,958 SNPs on 22 autosomes, we observed a total of 134 SNPs significantly associated with risk of serous EOC with a p value below 10−6. Associations with deletion polymorphisms were observed on chromosomes 7, 8, 14, and 18 when controlling for false discovery rate at 1%; no duplication polymorphisms were significant. We observed a large deletion at chromosome 14 that occurred in 8.9% of cases but in only 3.9% of controls, with a p value of 5.59×10−8 (Odds Ratio (OR): 2.56, 95% confidence interval (CI): 1.82-3.60). Another common deletion on chromosome 7 occurred in 8.8% of cases and only 4.3% of controls (p=3.83×10−6; OR: 2.23, 95% CI: 1.60-3.10). Two additional regions on chromosomes 8 and 18 with deletion events less than 5% were also identified. Women who harbored the deletion on chromosome 8 were at lower risk of developing serous ovarian cancer (p=4.83×10−8; OR: 0.04, 95% CI: 0.01-0.30). Women who carried the deletion on chromosome 18 had an increased risk for serous EOC (p=9.04×10−7; OR: 7.26, 95% CI: 2.94-17.97). Further validation of these four regions using independent data sets is currently underway. In summary, this is the largest reported genome-wide study of CNVs and serous EOC risk. These preliminary results suggested that germline CNVs may play an important role in ovarian cancer susceptibility. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1649. doi:1538-7445.AM2012-1649

Abstract The etiology of ovarian cancer is poorly understood but there is clearly a heritable component. Efforts to identify susceptibility alleles rarely consider interactions among alleles or their joint effects because it quickly becomes computationally intractable. In this study, we sought to identify multi-SNP effects jointly with a pathway-based analysis (GSEA-SNP) of 1,952 EOC cases and 2,042 frequency-matched controls genotyped with the Illumina 610K array. All subjects were self-reported non-Hispanic non-Jewish Caucasians, with SNP and sample call rates > 95%. Subjects with ambiguous gender, unresolved identical genotypes and < 80% European ancestry were excluded. SNPs with MAF < 1% were excluded. Missing genotypes were inferred using Mach based on the HapMap CEU population. SNPs not within introns were annotated to genes within 100 bp. We retrieved the following databases of gene sets (GSs) from a compiled database, MsigDB: human chromosome and cytogenetic band, chemical and genetic perturbations, canonical pathways, microRNA binding targets, transcription factor targets (TFT), cancer gene neighborhood (CGN), Cancer modules and Gene Ontology. The GSEA-SNP approach calculates and rank orders the trend statistic for association between each SNP and EOC risk. The Enrichment Score (ES) estimates the overrepresentation of top-ranked SNPs for each GS; the statistical significance was estimated using 10,000 permutations. The ES was normalized according to the size of GS to yield Normalized Enrichment Score (NES). The false discovery (FDR) rate was estimated using NES to adjust for multiple hypothesis testing. A total of 5181 gene-sets were included in the analysis. When controlling the FDR at 15%, 14 of the GSs were highly enriched with association signals, including two chromosomal regions, 8q13 (p = 0.0055, FDR = 0.15) and 6p24 (p = 0.0012, FDR = 0.08). BIOCARTA_LYM_PATHWAY, a pathway related to cell adhesion and diapedesis of lymphocytes, was also significant (p = 0.0002, FDR = 0.12). A TFT GS, composed of genes with promoter regions around transcription start sites containing the motif GGCNRNWCTTYS, was associated with risk (p = 0.0004, FDR = 0.08). Currently, no known transcription factors bind to this computationally predicted motif. Molecular function of double stranded RNA binding was also enriched (p = 0.005, FDR = 0.10). The remaining enriched sets were CGN sets, for which the neighborhoods around the cancer genes were originally defined using correlation of gene expression from 4 large data sets of various cancer types. The enriched CGN included: CD48 (p = 0.011), CD53 (p = 0.009), CD97 (p = 0.011), INPP5D (p = 0.010), PTPN6 (p = 0.008), VAV1 (p = 0.011), ITGAL (p = 0.012), PTPRC (p = 0.012), and STAT6 (p = 0.010). In summary, these analyses detected biologically plausible GSs related to etiology of EOC, highlighting SNPs in core enrichment groups that were not identified using individual SNP tests. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4729. doi:10.1158/1538-7445.AM2011-4729

Abstract Recent GWAS have identified and confirmed associations between several SNPs in the gene desert located on chromosome 8q24 with risk of cancers of the breast, prostate, glioma, bladder and colorectum. Evidence supporting an association with ovarian cancer is limited and inconsistent. Here we evaluate preliminary results in the 8q24 region from stage I of an ongoing invasive epithelial ovarian cancer GWAS among 1864 cases (1096 serous cases) and 1912 controls. All subjects were self-reported non-Hispanic non-Jewish Caucasian, and were genotyped with the Illumina 610quad Chip. Samples and SNPs with call rates less than 95% were excluded (9.6% and 9.9%, respectively) as well as 128 subjects with ambiguous gender, unresolved identical genotypes and less than 80% European ancestry were also excluded. Our most significant hit in the region was rs6651252 located at 8q24.21 with p = 0.000125 for all ovarian cancer, and 1.02*10−5 for serous cases only. This region is located approximately 849 kb downstream of the previous 8q24.21 findings on other cancers, and was not in linkage disequilibrium with any of the haplotype blocks reported previously. We further examined the block structure of this region with eight genotyped SNPs located within 129.61 to 129.64 MB (rs10088218, rs1516973, rs1516975, rs16903097, rs10098821, rs6651252, rs1561928, and rs1561927) and performed haplotype analysis to evaluate the association with ovarian cancer risk. One haplotype (AAGCAGGG, estimated frequency=0.11) was significantly associated with decreased ovarian cancer risk with an odds ratio of 0.67 (all cases, 95% CI: 0.78-0.91, p=0.0026, empirical p= 0.0020). This region contains no known genes, but it is located upstream and overlaps with a known copy number variation (CNV) reported by Redon et al. (2006). Previous published literature on array CGH and CNV results from TCGA (The Cancer Genome Atlas) also indicate that the 8q24.21 region is associated with frequent gains/amplification in ovarian tumors. Combined analysis of SNPs and CNV identified in this region is underway to elucidate the joint effect of these two types of genetic variation on ovarian cancer risk. Our result indicates a novel locus on 8q24.21 is associated with epithelial ovarian cancer susceptibility. Haplotype analysis reveals one specific haplotype that is associated with decreased ovarian cancer risk compared to the most common haplotype. Given previous 8q24 findings from other cancers and evidence from publicly available resources, fine-mapping and resequencing of this region is warranted. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4724.

Abstract Introduction: Tissue microarrays (TMA) enable rapid analysis of biomarkers in large-scale studies involving archival tumor specimens, however, their utility in heterogeneous tumors such as ovarian cancer is limited. Methods: In this study, immunohistochemical analysis was done on TMAs comprised of epithelial ovarian cancer (EOC) to estimate the prevalence of loss of expression of three mismatch repair proteins. TMAs were initially created using cores sampled from the center of donor tissue blocks from 59 EOC cases. Full sections were subsequently created and levels of expression were compared between tissues sampled from the central portion versus the periphery. Follow-up analyses were done by obtaining cores from the periphery of up to five additional donor blocks per case. A linear mixed model for each protein was used to investigate differences between results from the initial and follow-up blocks. Results: In the original TMAs created using centrally sampled cores, loss of mismatch repair expression was noted in 17 (29%) of the 59 cases. By comparison, analyses from peripherally sampled cores revealed loss of expression in only 6 of these 17 cases. For each protein, significant differences (P < 0.05) were detected between results from the initial donor block and the majority of the follow-up blocks. Conclusions: Our investigations, based on EOC, suggest that sampling variability in protein expression may result when TMAs are used. Thus, at least for EOC, it is important to preferentially sample from the periphery of tumor blocks where exposure to tissue fixatives is optimal. (Cancer Epidemiol Biomarkers Prev 2009;18(1):28–34)