Impact of insomnia on ovarian cancer risk and survival: a Mendelian randomization study.

BACKGROUND: Insomnia is the most common sleep disorder in patients with epithelial ovarian cancer (EOC). We investigated the causal association between genetically predicted insomnia and EOC risk and survival through a two-sample Mendelian randomization (MR) study. METHODS: Insomnia was proxied using genetic variants identified in a genome-wide association study (GWAS) meta-analysis of UK Biobank and 23andMe. Using genetic associations with EOC risk and overall survival from the Ovarian Cancer Association Consortium (OCAC) GWAS in 66,450 women (over 11,000 cases with clinical follow-up), we performed Iterative Mendelian Randomization and Pleiotropy (IMRP) analysis followed by a set of sensitivity analyses. Genetic associations with survival and response to treatment in ovarian cancer study of The Cancer Genome Atlas (TCGA) were estimated controlling for chemotherapy and clinical factors. FINDINGS: Insomnia was associated with higher risk of endometrioid EOC (OR = 1.60, 95% CI 1.05-2.45) and lower risk of high-grade serous EOC (HGSOC) and clear cell EOC (OR = 0.79 and 0.48, 95% CI 0.63-1.00 and 0.27-0.86, respectively). In survival analysis, insomnia was associated with shorter survival of invasive EOC (OR = 1.45, 95% CI 1.13-1.87) and HGSOC (OR = 1.4, 95% CI 1.04-1.89), which was attenuated after adjustment for body mass index and reproductive age. Insomnia was associated with reduced survival in TCGA HGSOC cases who received standard chemotherapy (OR = 2.48, 95% CI 1.13-5.42), but was attenuated after adjustment for clinical factors. INTERPRETATION: This study supports the impact of insomnia on EOC risk and survival, suggesting treatments targeting insomnia could be pivotal for prevention and improving patient survival. FUNDING: National Institutes of Health, National Cancer Institute. Full funding details are provided in acknowledgments.

Functional analysis of the 1p34.3 risk locus implicates GNL2 in high-grade serous ovarian cancer.

The high-grade serous ovarian cancer (HGSOC) risk locus at chromosome 1p34.3 resides within a frequently amplified genomic region signifying the presence of an oncogene. Here, we integrate in silico variant-to-function analysis with functional studies to characterize the oncogenic potential of candidate genes in the 1p34.3 locus. Fine mapping of genome-wide association statistics identified candidate causal SNPs local to H3K27ac-demarcated enhancer regions that exhibit allele-specific binding for CTCF in HGSOC and normal fallopian tube secretory epithelium cells (FTSECs). SNP risk associations colocalized with eQTL for six genes (DNALI1, GNL2, RSPO1, SNIP1, MEAF6, and LINC01137) that are more highly expressed in carriers of the risk allele, and three (DNALI1, GNL2, and RSPO1) were upregulated in HGSOC compared to normal ovarian surface epithelium cells and/or FTSECs. Increased expression of GNL2 and MEAF6 was associated with shorter survival in HGSOC with 1p34.3 amplifications. Despite its activation of ß-catenin signaling, RSPO1 overexpression exerted no effects on proliferation or colony formation in our study of ovarian cancer and FTSECs. Instead, GNL2, MEAF6, and SNIP1 silencing impaired in vitro ovarian cancer cell growth. Additionally, GNL2 silencing diminished xenograft tumor formation, whereas overexpression stimulated proliferation and colony formation in FTSECs. GNL2 influences 60S ribosomal subunit maturation and global protein synthesis in ovarian cancer and FTSECs, providing a potential mechanism of how GNL2 upregulation might promote ovarian cancer development and mediate genetic susceptibility of HGSOC.

Utilizing digital pathology to quantify stromal caveolin-1 expression in malignant and benign ovarian tumors: Associations with clinicopathological parameters and clinical outcomes.

Loss of stromal caveolin-1 (Cav-1) is a biomarker of a cancer-associated fibroblast (CAF) phenotype and is related to progression, metastasis, and poor outcomes in several cancers. The objective of this study was to evaluate the clinical significance of Cav-1 expression in invasive epithelial ovarian cancer (OvCa). Epithelial and stromal Cav-1 expression were quantified in serous OvCa and benign ovarian tissue in two, independent cohorts-one quantified expression using immunohistochemistry (IHC) and the other using multiplex immunofluorescence (IF) with digital image analysis designed to target CAF-specific expression. Cav-1 expression was significantly downregulated in OvCa stroma compared to non-neoplastic stroma using both the IHC (p = 0.002) and IF (p = 1.8x10-13) assays. OvCa stroma showed Cav-1 downregulation compared to tumor epithelium with IHC (p = 1.2x10-24). Conversely, Cav-1 expression was higher in OvCa stroma compared to tumor epithelium with IF (p = 0.002). There was moderate correlation between IHC and IF methods for stromal Cav-1 expression (r2 = 0.69, p = 0.006) whereas there was no correlation for epithelial expression (r2 = 0.006, p = 0.98). Irrespective of the staining assay, neither response to therapy or overall survival correlated with the expression level of Cav-1 in the stroma or tumor epithelium. Our findings demonstrate a loss of stromal Cav-1 expression in ovarian serous carcinomas. Studies are needed to replicate these findings and explore therapeutic implications, particularly for immunotherapy response.

Morphologic and molecular correlates of EZH2 as a predictor of platinum resistance in high-grade ovarian serous carcinoma.

BACKGROUND: Enhancer of zesta homologue 2 (EZH2) is an essential component of polycomb repressive complex 2 (PRC2) that contributes to tumor progression and chemo-resistance. The aim of this study was to comprehensively assess the prognostic value of EZH2 across the morphologic and molecular spectra of high-grade serous ovarian carcinoma (HGSOC) by utilizing both immunohistochemistry (IHC) and proteogenomic technologies. METHODS: IHC of EZH2 was performed using a tissue microarray of 79 HGSOC scored (+/-) for lymphovascular invasion (LVI), tumor-infiltrating lymphocytic aggregates ≥1 mm (TIL) and architectural growth patterns. The association of EZH2 H-score with response to therapy and overall survival was evaluated by tumor features. We also evaluated EZH2 transcriptional (RNA sequencing) and protein (mass spectrometry) expression from bulk tumor samples from 336 HGSOC from The Cancer Genome Atlas (TCGA). EZH2 expression and co-expression networks were compared by clinical outcomes. RESULTS: For HGSOC without TIL (58%), EZH2 expression was almost 2-fold higher in platinum resistant tumors (P = 0.01). Conversely, EZH2 was not associated with platinum resistance among TIL+ HGSOC (P = 0.41). EZH2 expression was associated with reduced survival for tumors with LVI (P = 0.04). Analysis of TCGA found higher EZH2 expression in immunoreactive and proliferative tumors (P = 6.7 × 10- 5) although protein levels were similar across molecular subtypes (P = 0.52). Both mRNA and protein levels of EZH2 were lower in platinum resistant tumors although they were not associated with survival. Co-expression analysis revealed EZH2 networks totaling 1049 mRNA and 448 proteins that were exclusive to platinum sensitive or resistant tumors. The EZH2 network in resistant HGSOC included CARM1 which was positively correlated with EZH2 at both mRNA (r = 0.33, p = 0.003) and protein (r = 0.14, P = 0.01) levels. Further, EZH2 co-expression with CARM1 corresponded to a decreased prognostic significance of EZH2 expression in resistant tumors. CONCLUSIONS: Our findings demonstrate that EZH2 expression varies based on its interactions with immunologic pathways and tumor microenvironment, impacting the prognostic interpretation. The association between high EZH2 expression and platinum resistance in TIL- HGSOC warrants further study of the implications for therapeutic strategies.

DNA Methylation in Ovarian Cancer Susceptibility.

Epigenetic alterations are somatically acquired over the lifetime and during neoplastic transformation but may also be inherited as widespread 'constitutional' alterations in normal tissues that can cause cancer predisposition. Epithelial ovarian cancer (EOC) has an established genetic susceptibility and mounting epidemiological evidence demonstrates that DNA methylation (DNAm) intermediates as well as independently contributes to risk. Targeted studies of known EOC susceptibility genes (CSGs) indicate rare, constitutional BRCA1 promoter methylation increases familial and sporadic EOC risk. Blood-based epigenome-wide association studies (EWAS) for EOC have detected a total of 2846 differentially methylated probes (DMPs) with 71 genes replicated across studies despite significant heterogeneity. While EWAS detect both symptomatic and etiologic DMPs, adjustments and analytic techniques may enrich risk associations, as evidenced by the detection of dysregulated methylation of BNC2-a known CSG identified by genome-wide associations studies (GWAS). Integrative genetic-epigenetic approaches have mapped methylation quantitative trait loci (meQTL) to EOC risk, revealing DNAm variations that are associated with nine GWAS loci and, further, one novel risk locus. Increasing efforts to mapping epigenome variation across populations and cell types will be key to decoding both the genomic and epigenomic causal pathways to EOC.

Genome-wide Analysis of Common Copy Number Variation and Epithelial Ovarian Cancer Risk.

BACKGROUND: Germline DNA copy number variation (CNV) is a ubiquitous source of genetic variation and remains largely unexplored in association with epithelial ovarian cancer (EOC) risk. METHODS: CNV was quantified in the DNA of approximately 3,500 cases and controls genotyped with the Illumina 610k and HumanOmni2.5M arrays. We performed a genome-wide association study of common (>1%) CNV regions (CNVRs) with EOC and high-grade serous (HGSOC) risk and, using The Cancer Genome Atlas (TCGA), performed in silico analyses of tumor-gene expression. RESULTS: Three CNVRs were associated (P < 0.01) with EOC risk: two large (∼100 kb) regions within the 610k set and one small (<5 kb) region with the higher resolution 2.5M data. Large CNVRs included a duplication at LILRA6 (OR = 2.57; P = 0.001) and a deletion at CYP2A7 (OR = 1.90; P = 0.007) that were strongly associated with HGSOC risk (OR = 3.02; P = 8.98 × 10-5). Somatic CYP2A7 alterations correlated with EGLN2 expression in tumors (P = 2.94 × 10-47). An intronic ERBB4/HER4 deletion was associated with reduced EOC risk (OR = 0.33; P = 9.5 × 10-2), and somatic deletions correlated with ERBB4 downregulation (P = 7.05 × 10-5). Five CNVRs were associated with HGSOC, including two reduced-risk deletions: one at 1p36.33 (OR = 0.28; P = 0.001) that correlated with lower CDKIIA expression in TCGA tumors (P = 2.7 × 10-7), and another at 8p21.2 (OR = 0.52; P = 0.002) that was present somatically where it correlated with lower GNRH1 expression (P = 5.9 × 10-5). CONCLUSIONS: Though CNV appears to not contribute largely to EOC susceptibility, a number of low-to-common frequency variants may influence the risk of EOC and tumor-gene expression. IMPACT: Further research on CNV and EOC susceptibility is warranted, particularly with CNVs estimated from high-density arrays.

MAFG is a potential therapeutic target to restore chemosensitivity in cisplatin-resistant cancer cells by increasing reactive oxygen species.

Adjuvant chemotherapy for solid tumors based on platinum-derived compounds such as cisplatin is the treatment of choice in most cases. Cisplatin triggers signaling pathways that lead to cell death, but it also induces changes in tumor cells that modify the therapeutic response, thereby leading to cisplatin resistance. We have recently reported that microRNA-7 is silenced by DNA methylation and is involved in the resistance to platinum in cancer cells through the action of the musculoaponeurotic fibrosarcoma oncogene family, protein G (MAFG). In the present study, we first confirm the miR-7 epigenetic regulation of MAFG in 44 normal- and/or tumor-paired samples in non-small-cell lung cancer (NSCLC). We also provide translational evidence of the role of MAFG and the clinical outcome in NSCLC by the interrogation of two extensive in silico databases of 2019 patients. Moreover, we propose that MAFG-mediated resistance could be conferred due to lower reactive oxygen species production after cisplatin exposure. We developed specifically selected aptamers against MAFG, with high sensitivity to detect the protein at a nuclear level probed by aptacytochemistry and histochemistry analyses. The inhibition of MAFG activity through the action of the specific aptamer apMAFG6F increased the levels of reactive oxygen species production and the sensitivity to cisplatin. We report first the specific nuclear identification of MAFG as a novel detection method for diagnosis in NSCLC, and then we report that MAFG modulates the redox response and confers cell protection against free radicals generated after platinum administration, thus also being a promising therapeutic target.

A Transcriptome-Wide Association Study Among 97,898 Women to Identify Candidate Susceptibility Genes for Epithelial Ovarian Cancer Risk.

Large-scale genome-wide association studies (GWAS) have identified approximately 35 loci associated with epithelial ovarian cancer (EOC) risk. The majority of GWAS-identified disease susceptibility variants are located in noncoding regions, and causal genes underlying these associations remain largely unknown. Here, we performed a transcriptome-wide association study to search for novel genetic loci and plausible causal genes at known GWAS loci. We used RNA sequencing data (68 normal ovarian tissue samples from 68 individuals and 6,124 cross-tissue samples from 369 individuals) and high-density genotyping data from European descendants of the Genotype-Tissue Expression (GTEx V6) project to build ovarian and cross-tissue models of genetically regulated expression using elastic net methods. We evaluated 17,121 genes for their cis-predicted gene expression in relation to EOC risk using summary statistics data from GWAS of 97,898 women, including 29,396 EOC cases. With a Bonferroni-corrected significance level of P < 2.2 × 10-6, we identified 35 genes, including FZD4 at 11q14.2 (Z = 5.08, P = 3.83 × 10-7, the cross-tissue model; 1 Mb away from any GWAS-identified EOC risk variant), a potential novel locus for EOC risk. All other 34 significantly associated genes were located within 1 Mb of known GWAS-identified loci, including 23 genes at 6 loci not previously linked to EOC risk. Upon conditioning on nearby known EOC GWAS-identified variants, the associations for 31 genes disappeared and three genes remained (P < 1.47 × 10-3). These data identify one novel locus (FZD4) and 34 genes at 13 known EOC risk loci associated with EOC risk, providing new insights into EOC carcinogenesis.Significance: Transcriptomic analysis of a large cohort confirms earlier GWAS loci and reveals FZD4 as a novel locus associated with EOC risk. Cancer Res; 78(18); 5419-30. ©2018 AACR.

Epidemiology of ovarian cancer: a review.

Ovarian cancer (OC) is the seventh most commonly diagnosed cancer among women in the world and the tenth most common in China. Epithelial OC is the most predominant pathologic subtype, with five major histotypes that differ in origination, pathogenesis, molecular alterations, risk factors, and prognosis. Genetic susceptibility is manifested by rare inherited mutations with high to moderate penetrance. Genome-wide association studies have additionally identified 29 common susceptibility alleles for OC, including 14 subtype-specific alleles. Several reproductive and hormonal factors may lower risk, including parity, oral contraceptive use, and lactation, while others such as older age at menopause and hormone replacement therapy confer increased risks. These associations differ by histotype, especially for mucinous OC, likely reflecting differences in etiology. Endometrioid and clear cell OC share a similar, unique pattern of associations with increased risks among women with endometriosis and decreased risks associated with tubal ligation. OC risks associated with other gynecological conditions and procedures, such as hysterectomy, pelvic inflammatory disease, and polycystic ovarian syndrome, are less clear. Other possible risk factors include environmental and lifestyle factors such as asbestos and talc powder exposures, and cigarette smoking. The epidemiology provides clues on etiology, primary prevention, early detection, and possibly even therapeutic strategies.

Integration of Population-Level Genotype Data with Functional Annotation Reveals Over-Representation of Long Noncoding RNAs at Ovarian Cancer Susceptibility Loci.

BACKGROUND: Genome-wide association studies (GWAS) have identified multiple loci associated with epithelial ovarian cancer (EOC) susceptibility, but further progress requires integration of epidemiology and biology to illuminate true risk loci below genome-wide significance levels (P < 5 × 10-8). Most risk SNPs lie within non-protein-encoding regions, and we hypothesize that long noncoding RNA (lncRNA) genes are enriched at EOC risk regions and represent biologically relevant functional targets. METHODS: Using imputed GWAS data from about 18,000 invasive EOC cases and 34,000 controls of European ancestry, the GENCODE (v19) lncRNA database was used to annotate SNPs from 13,442 lncRNAs for permutation-based enrichment analysis. Tumor expression quantitative trait locus (eQTL) analysis was performed for sub-genome-wide regions (1 × 10-5 > P > 5 × 10-8) overlapping lncRNAs. RESULTS: Of 5,294 EOC-associated SNPs (P < 1.0 × 10-5), 1,464 (28%) mapped within 53 unique lncRNAs and an additional 3,484 (66%) SNPs were correlated (r2 > 0.2) with SNPs within 115 lncRNAs. EOC-associated SNPs comprised 130 independent regions, of which 72 (55%) overlapped with lncRNAs, representing a significant enrichment (P = 5.0 × 10-4) that was more pronounced among a subset of 5,401 lncRNAs with active epigenetic regulation in normal ovarian tissue. EOC-associated lncRNAs and their putative promoters and transcription factors were enriched for biologically relevant pathways and eQTL analysis identified five novel putative risk regions with allele-specific effects on lncRNA gene expression. CONCLUSIONS: lncRNAs are significantly enriched at EOC risk regions, suggesting a mechanistic role for lncRNAs in driving predisposition to EOC. IMPACT: lncRNAs represent key candidates for integrative epidemiologic and functional studies. Further research on their biologic role in ovarian cancer is indicated. Cancer Epidemiol Biomarkers Prev; 26(1); 116-25. ©2016 AACR.

Exome genotyping arrays to identify rare and low frequency variants associated with epithelial ovarian cancer risk.

Rare and low frequency variants are not well covered in most germline genotyping arrays and are understudied in relation to epithelial ovarian cancer (EOC) risk. To address this gap, we used genotyping arrays targeting rarer protein-coding variation in 8,165 EOC cases and 11,619 controls from the international Ovarian Cancer Association Consortium (OCAC). Pooled association analyses were conducted at the variant and gene level for 98,543 variants directly genotyped through two exome genotyping projects. Only common variants that represent or are in strong linkage disequilibrium (LD) with previously-identified signals at established loci reached traditional thresholds for exome-wide significance (P < 5.0 × 10 - 7). One of the most significant signals (Pall histologies = 1.01 × 10 - 13;Pserous = 3.54 × 10 - 14) occurred at 3q25.31 for rs62273959, a missense variant mapping to the LEKR1 gene that is in LD (r2 = 0.90) with a previously identified 'best hit' (rs7651446) mapping to an intron of TIPARP. Suggestive associations (5.0 × 10 - 5 > P≥5.0 ×10 - 7) were detected for rare and low-frequency variants at 16 novel loci. Four rare missense variants were identified (ACTBL2 rs73757391 (5q11.2), BTD rs200337373 (3p25.1), KRT13 rs150321809 (17q21.2) and MC2R rs104894658 (18p11.21)), but only MC2R rs104894668 had a large effect size (OR = 9.66). Genes most strongly associated with EOC risk included ACTBL2 (PAML = 3.23 × 10 - 5; PSKAT-o = 9.23 × 10 - 4) and KRT13 (PAML = 1.67 × 10 - 4; PSKAT-o = 1.07 × 10 - 5), reaffirming variant-level analysis. In summary, this large study identified several rare and low-frequency variants and genes that may contribute to EOC susceptibility, albeit with possible small effects. Future studies that integrate epidemiology, sequencing, and functional assays are needed to further unravel the unexplained heritability and biology of this disease.

Cross-Cancer Genome-Wide Analysis of Lung, Ovary, Breast, Prostate, and Colorectal Cancer Reveals Novel Pleiotropic Associations.

Identifying genetic variants with pleiotropic associations can uncover common pathways influencing multiple cancers. We took a two-stage approach to conduct genome-wide association studies for lung, ovary, breast, prostate, and colorectal cancer from the GAME-ON/GECCO Network (61,851 cases, 61,820 controls) to identify pleiotropic loci. Findings were replicated in independent association studies (55,789 cases, 330,490 controls). We identified a novel pleiotropic association at 1q22 involving breast and lung squamous cell carcinoma, with eQTL analysis showing an association with ADAM15/THBS3 gene expression in lung. We also identified a known breast cancer locus CASP8/ALS2CR12 associated with prostate cancer, a known cancer locus at CDKN2B-AS1 with different variants associated with lung adenocarcinoma and prostate cancer, and confirmed the associations of a breast BRCA2 locus with lung and serous ovarian cancer. This is the largest study to date examining pleiotropy across multiple cancer-associated loci, identifying common mechanisms of cancer development and progression. Cancer Res; 76(17); 5103-14. ©2016 AACR.

STAT3 polymorphisms may predict an unfavorable response to first-line platinum-based therapy for women with advanced serous epithelial ovarian cancer.

Cancer stem cells (CSC) contribute to epithelial ovarian cancer (EOC) progression and therapeutic response. We hypothesized that germline single nucleotide polymorphisms (SNPs) in CSC-related genes may predict an initial therapeutic response for women newly diagnosed with EOC. A nested case-control design was used to study 361 women with advanced-stage serous EOC treated with surgery followed by first-line platinum-based combination therapy at Moffitt Cancer Center or as part of The Cancer Genome Atlas Study. "Cases" included 102 incomplete responders (IRs) and "controls" included 259 complete clinical responders (CRs) to therapy. Using Illumina genotyping arrays and imputation, DNA samples were evaluated for 5,509 SNPs in 24 ovarian CSC-related genes. We also evaluated the overall significance of each CSC gene using the admixture maximum likelihood (AML) test, and correlated genotype with EOC tumor tissue expression. The strongest SNP-level associations with an IR to therapy were identified for correlated (r(2) > 0.80) SNPs within signal transducer and activator of transcription 3 (STAT3) [odds ratio (OR), 2.24; 95% confidence interval (CI), 1.32-3.78; p = 0.0027], after adjustment for age, population stratification, grade and residual disease. At the gene level, STAT3 was significantly associated with an IR to therapy (pAML = 0.006). rs1053004, a STAT3 SNP in a putative miRNA-binding site, was associated with STAT3 expression (p = 0.057). This is the first study to identify germline STAT3 variants as independent predictors of an unfavorable therapeutic response for EOC patients. Findings suggest that STAT3 genotype may identify high-risk women likely to respond more favorably to novel therapeutic combinations that include STAT3 inhibitors.

A functional variant in HOXA11-AS, a novel long non-coding RNA, inhibits the oncogenic phenotype of epithelial ovarian cancer.

The homeobox A (HOXA) region of protein-coding genes impacts female reproductive system embryogenesis and ovarian carcinogenesis. The 5-prime end of HOXA includes three long non-coding RNAs (lncRNAs) (HOXA10-AS, HOXA11-AS, and HOTTIP) that are underexplored in epithelial ovarian cancer (EOC). We evaluated whether common genetic variants in these lncRNAs are associated with EOC risk and/or have functional roles in EOC development. Using genome-wide association study data from 1,201 serous EOC cases and 2,009 controls, an exonic variant within HOXA11-AS, rs17427875 (A>T), was marginally associated with reduced serous EOC risk (OR = 0.88 (95% CI: 0.78-1.01, p = 0.06). Functional studies of ectopic expression of HOXA11-AS minor allele T in EOC cells showed decreased survival, proliferation, migration, and invasion compared to common allele A expression. Additionally, stable expression of HOXA11-AS minor allele T reduced primary tumor growth in mouse xenograft models to a greater extent than common allele A. Furthermore, HOXA11-AS expression levels were significantly lower in human EOC tumors than normal ovarian tissues (p < 0.05), suggesting that HOXA11-AS has a tumor suppressor function in EOC which may be enhanced by the T allele. These findings demonstrate for the first time a role for HOXA11-AS in EOC with effects that could be modified by germline variants.

Evidence of differential effects of vitamin d receptor variants on epithelial ovarian cancer risk by predicted vitamin d status.

INTRODUCTION: Experimental studies suggest vitamin D inhibits ovarian carcinogenesis. Yet, epidemiologic studies of ovarian cancer risk and lifestyle correlates of vitamin D status, plasma 25-hydroxyvitamin D [25(OH)D], or vitamin D receptor (VDR) variants have been inconsistent. OBJECTIVE: To evaluate VDR genetic associations by high vs. low predicted 25(OH)D, scores derived from known determinants of plasma 25(OH)D. To assess ovarian cancer associations with variants identified in genome-wide association studies (GWAS) of plasma 25(OH)D. METHODS: We genotyped up to seven VDR and eight 25(OH)D GWAS variants in the Nurses' Health Studies (562 cases, 1,553 controls) and New England Case-Control study (1,821 cases, 1,870 controls). We estimated haplotype scores using expectation-maximization-based algorithms. We used unconditional logistic regression to calculate odds ratios (ORs) and 95% confidence intervals (CI). We combined study results using DerSimonian and Laird meta-analysis. RESULTS: Ovarian cancer risk increased per A allele of rs7975232 (VDR; OR = 1.12, 95% CI = 1.01-1.25) among all women. When stratified by predicted 25(OH)D, ovarian cancer was associated with rs731236 (VDR; per C allele OR = 1.31) and rs7975232 (OR = 1.38) among women with high predicted 25(OH)D, but not among women with low levels (P ≤ 0.009). We also observed heterogeneity by predicted 25(OH)D for the ovarian cancer association with VDR 3' end haplotypes (P = 0.009). Of 25(OH)D-associated GWAS loci, rs7041 was associated with reduced ovarian cancer risk (per T allele OR = 0.92, 95% CI = 0.85-0.99), which did not differ by predicted 25(OH)D status. CONCLUSION: Our study suggests an influence of VDR 3' end variants on ovarian cancer risk may be observed in women with high predicted 25(OH)D, which remained even after taking multiple comparisons into consideration. Future studies are needed to confirm our results and explore further the relation between vitamin D exposure, genetic variants, and ovarian cancer risk.