Lissowska, Jolanta

Abstract Genome-wide association studies have identified 40 ovarian cancer risk loci. However, the mechanisms underlying these associations remain elusive. In this study, we conducted a two-pronged approach to identify candidate causal SNPs and assess underlying biological mechanisms at chromosome 9p22.2, the first and most statistically significant associated locus for ovarian cancer susceptibility. Three transcriptional regulatory elements with allele-specific effects and a scaffold/matrix attachment region were characterized and, through physical DNA interactions, BNC2 was established as the most likely target gene. We determined the consensus binding sequence for BNC2 in vitro, verified its enrichment in BNC2 ChIP-seq regions, and validated a set of its downstream target genes. Fine-mapping by dense regional genotyping in over 15,000 ovarian cancer cases and 30,000 controls identified SNPs in the scaffold/matrix attachment region as among the most likely causal variants. This study reveals a comprehensive regulatory landscape at 9p22.2 and proposes a likely mechanism of susceptibility to ovarian cancer. Significance: Mapping the 9p22.2 ovarian cancer risk locus identifies BNC2 as an ovarian cancer risk gene. See related commentary by Choi and Brown, p. 439

Thymidylate synthase (TYMS) is a crucial enzyme for DNA synthesis. TYMS expression is regulated by its antisense mRNA, ENOSF1. Disrupted regulation may promote uncontrolled DNA synthesis and tumor growth. We sought to replicate our previously reported association between rs495139 in the TYMS-ENOSF1 3′ gene region and increased risk of mucinous ovarian carcinoma (MOC) in an independent sample. Genotypes from 24,351 controls to 15,000 women with invasive OC, including 665 MOC, were available. We estimated per-allele odds ratios (OR) and 95% confidence intervals (CI) using unconditional logistic regression, and meta-analysis when combining these data with our previous report. The association between rs495139 and MOC was not significant in the independent sample (OR = 1.09; 95% CI = 0.97–1.22; p = 0.15; N = 665 cases). Meta-analysis suggested a weak association (OR = 1.13; 95% CI = 1.03–1.24; p = 0.01; N = 1019 cases). No significant association with risk of other OC histologic types was observed (p = 0.05 for tumor heterogeneity). In expression quantitative trait locus (eQTL) analysis, the rs495139 allele was positively associated with ENOSF1 mRNA expression in normal tissues of the gastrointestinal system, particularly esophageal mucosa (r = 0.51, p = 1.7 × 10−28), and nonsignificantly in five MOC tumors. The association results, along with inconclusive tumor eQTL findings, suggest that a true effect of rs495139 might be small.

Abstract Background: Pelvic inflammatory disease (PID) has been associated with ovarian cancer and Chlamydia trachomatis (chlamydia) is the leading cause of PID in the developed world. Primary infection with chlamydia is often asymptomatic and may persist for several months/years, thus ascertainment of PID in epidemiologic studies is challenging due to pervasive misclassification and under-reporting. To improve our understanding of the role of chlamydia and other infectious agents in developing ovarian cancer, we evaluated serologic markers of infection with incident ovarian cancer using a staged approach in two independent populations. Methods: Studies included: 1) a case-control study conducted in Poland of 278 ovarian cancer cases diagnosed from 2000 to 2003 and age- and study site-matched controls (n=556) and 2) a nested case-control study from the Prostate, Lung, Colorectal, and Ovarian (PLCO) Cancer Screening Trial including 160 women who eventually developed ovarian cancer and 159 matched controls. We calculated odds ratios (OR) and 95% confidence intervals (CI) for the association between serologic marker titers and ovarian cancer risk using logistic regression adjusted for matching factors and potential confounders. We evaluated laboratory cutpoints indicative of prior chlamydia infection and tested more stringent marker thresholds, identified in Poland and applied in PLCO, that could be relevant for a PID-like infection. Results: In the Polish study, positivity for the chlamydia plasmid-encoded Pgp3 protein was associated with increased ovarian cancer risk [OR (95% CI): 1.63 (1.20-2.22)]; when a positive result was redefined at a higher titer, ovarian cancer risk increased [cutpoint 2: 2.00 (1.38-2.89); cutpoint 3 (max OR): 2.19 (1.29-3.73)]. All other chlamydia markers measured were associated with increased risk at the more stringent positivity threshold in Poland. In the prospective PLCO study, Pgp3 was associated with elevated risk at the laboratory cutpoint [1.43 (0.78-2.63)]; using higher thresholds, Pgp3 was associated with increased ovarian cancer risk [cutpoint 2: 2.25 (1.07-4.71); cutpoint 3: 2.53 (0.63-10.08)]. In both studies, markers of other infections (i.e., Mycoplasma genitalium, herpes simplex virus-2, human papillomavirus, herpes simplex virus-1, polyomavirus, hepatitis B, hepatitis C, Epstein Barr virus, cytomegalovirus) measured were not associated with risk. Conclusions: In two independent populations, the gold standard serologic marker of prior/current chlamydia infection (Pgp3) was associated with a doubling in ovarian cancer risk, whereas markers of other infectious agents were unrelated. Although these findings need to be replicated, they suggest potential ovarian cancer risk reduction through targeted treatment of chlamydia infections. Citation Format: Britton Trabert, Tim Waterboer, Sally B. Coburn, Louise A. Brinton, Mark E. Sherman, Jolanta Lissowska, Beata Peplonska, Patricia Hartge, Michael Pawlita, Nicolas Wentzensen. Serologic markers of infectious agents and ovarian cancer: Markers of prior Chlamydia trachomatis infection associated with increased ovarian cancer risk in two independent populations [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4942.

Limited epidemiological evidence suggests that the etiology of hormone receptor positive (HR+) breast cancer may differ by levels of histologic grade and proliferation. We pooled risk factor and pathology data on 5,905 HR+ breast cancer cases and 26,281 controls from 11 epidemiological studies. Proliferation was determined by centralized automated measures of KI67 in tissue microarrays. Odds ratios (OR), 95% confidence intervals (CI) and p‐values for case–case and case–control comparisons for risk factors in relation to levels of grade and quartiles (Q1–Q4) of KI67 were estimated using polytomous logistic regression models. Case–case comparisons showed associations between nulliparity and high KI67 [OR (95% CI) for Q4 vs. Q1 = 1.54 (1.22, 1.95)]; obesity and high grade [grade 3 vs. 1 = 1.68 (1.31, 2.16)] and current use of combined hormone therapy (HT) and low grade [grade 3 vs. 1 = 0.27 (0.16, 0.44)] tumors. In case–control comparisons, nulliparity was associated with elevated risk of tumors with high but not low levels of proliferation [1.43 (1.14, 1.81) for KI67 Q4 vs. 0.83 (0.60, 1.14) for KI67 Q1]; obesity among women ≥50 years with high but not low grade tumors [1.55 (1.17, 2.06) for grade 3 vs. 0.88 (0.66, 1.16) for grade 1] and HT with low but not high grade tumors [3.07 (2.22, 4.23) for grade 1 vs. 0.85 (0.55, 1.30) for grade 3]. Menarcheal age and family history were similarly associated with HR+ tumors of different grade or KI67 levels. These findings provide insights into the etiologic heterogeneity of HR+ tumors.

Background The rarity of mutations in PALB2, CHEK2 and ATM make it difficult to estimate precisely associated cancer risks. Population-based family studies have provided evidence that at least some of these mutations are associated with breast cancer risk as high as those associated with rare BRCA2 mutations. We aimed to estimate the relative risks associated with specific rare variants in PALB2, CHEK2 and ATM via a multicentre case-control study. Methods We genotyped 10 rare mutations using the custom iCOGS array: PALB2 c.1592delT, c.2816T>G and c.3113G>A, CHEK2 c.349A>G, c.538C>T, c.715G>A, c.1036C>T, c.1312G>T, and c.1343T>G and ATM c.7271T>G. We assessed associations with breast cancer risk (42 671 cases and 42 164 controls), as well as prostate (22 301 cases and 22 320 controls) and ovarian (14 542 cases and 23 491 controls) cancer risk, for each variant. Results For European women, strong evidence of association with breast cancer risk was observed for PALB2 c.1592delT OR 3.44 (95% CI 1.39 to 8.52, p=7.1×10−5), PALB2 c.3113G>A OR 4.21 (95% CI 1.84 to 9.60, p=6.9×10−8) and ATM c.7271T>G OR 11.0 (95% CI 1.42 to 85.7, p=0.0012). We also found evidence of association with breast cancer risk for three variants in CHEK2, c.349A>G OR 2.26 (95% CI 1.29 to 3.95), c.1036C>T OR 5.06 (95% CI 1.09 to 23.5) and c.538C>T OR 1.33 (95% CI 1.05 to 1.67) (p≤0.017). Evidence for prostate cancer risk was observed for CHEK2 c.1343T>G OR 3.03 (95% CI 1.53 to 6.03, p=0.0006) for African men and CHEK2 c.1312G>T OR 2.21 (95% CI 1.06 to 4.63, p=0.030) for European men. No evidence of association with ovarian cancer was found for any of these variants. Conclusions This report adds to accumulating evidence that at least some variants in these genes are associated with an increased risk of breast cancer that is clinically important.

AbstractAutomated methods are needed to facilitate high‐throughput and reproducible scoring of Ki67 and other markers in breast cancer tissue microarrays (TMAs) in large‐scale studies. To address this need, we developed an automated protocol for Ki67 scoring and evaluated its performance in studies from the Breast Cancer Association Consortium. We utilized 166 TMAs containing 16,953 tumour cores representing 9,059 breast cancer cases, from 13 studies, with information on other clinical and pathological characteristics. TMAs were stained for Ki67 using standard immunohistochemical procedures, and scanned and digitized using the Ariol system. An automated algorithm was developed for the scoring of Ki67, and scores were compared to computer assisted visual (CAV) scores in a subset of 15 TMAs in a training set. We also assessed the correlation between automated Ki67 scores and other clinical and pathological characteristics. Overall, we observed good discriminatory accuracy (AUC = 85%) and good agreement (kappa = 0.64) between the automated and CAV scoring methods in the training set. The performance of the automated method varied by TMA (kappa range= 0.37–0.87) and study (kappa range = 0.39–0.69). The automated method performed better in satisfactory cores (kappa = 0.68) than suboptimal (kappa = 0.51) cores (p‐value for comparison = 0.005); and among cores with higher total nuclei counted by the machine (4,000–4,500 cells: kappa = 0.78) than those with lower counts (50–500 cells: kappa = 0.41; p‐value = 0.010). Among the 9,059 cases in this study, the correlations between automated Ki67 and clinical and pathological characteristics were found to be in the expected directions. Our findings indicate that automated scoring of Ki67 can be an efficient method to obtain good quality data across large numbers of TMAs from multicentre studies. However, robust algorithm development and rigorous pre‐ and post‐analytical quality control procedures are necessary in order to ensure satisfactory performance.