Naoko Sasamoto

Patterns of Associations with Epidemiologic Factors by High-Grade Serous Ovarian Cancer Gene Expression Subtypes

Abstract Background: Ovarian high-grade serous carcinomas (HGSC) comprise four distinct molecular subtypes based on mRNA expression patterns, with differential survival. Understanding risk factor associations is important to elucidate the etiology of HGSC. We investigated associations between different epidemiologic risk factors and HGSC molecular subtypes. Methods: We pooled data from 11 case–control studies with epidemiologic and tumor gene expression data from custom NanoString CodeSets developed through a collaboration within the Ovarian Tumor Tissue Analysis consortium. The PrOTYPE-validated NanoString-based 55-gene classifier was used to assign HGSC gene expression subtypes. We examined associations between epidemiologic factors and HGSC subtypes in 2,070 cases and 16,633 controls using multivariable-adjusted polytomous regression models. Results: Among the 2,070 HGSC cases, 556 (27%) were classified as C1.MES, 340 (16%) as C5.PRO, 538 (26%) as C2.IMM, and 636 (31%) as C4.DIF. The key factors, including oral contraceptive use, parity, breastfeeding, and family history of ovarian cancer, were similarly associated with all subtypes. Heterogeneity was observed for several factors. Former smoking [OR = 1.25; 95% confidence interval (CI) = 1.03, 1.51] and genital powder use (OR = 1.42; 95% CI = 1.08, 1.86) were uniquely associated with C2.IMM. History of endometriosis was associated with C5.PRO (OR = 1.46; 95% CI = 0.98, 2.16) and C4.DIF (OR = 1.27; 95% CI = 0.94, 1.71) only. Family history of breast cancer (OR = 1.44; 95% CI = 1.16, 1.78) and current smoking (OR = 1.40; 95% CI = 1.11, 1.76) were associated with C4.DIF only. Conclusions: This study observed heterogeneous associations of epidemiologic and modifiable factors with HGSC molecular subtypes. Impact: The different patterns of associations may provide key information about the etiology of the four subtypes.

Prospective Analysis of Circulating Biomarkers and Ovarian Cancer Risk in the UK Biobank

Abstract Background: Risk factors have a limited ability to predict individuals at high risk of developing ovarian cancer among average-risk women, highlighting the need for discovery of novel biomarkers. In the UK Biobank, we investigated serum biomarkers commonly measured in clinical laboratory tests and ovarian cancer risk. Methods: We conducted a prospective analysis of 20 serum biomarkers and ovarian cancer risk in 232,037 female UK Biobank participants (including 1,122 incident ovarian cancer cases diagnosed from 2006 to 2020). Multivariable adjusted Cox proportional hazards models were used to examine associations between biomarkers and ovarian cancer risk overall and by histotype. FDR was used to account for multiple testing. Results: Overall, higher levels of insulin-like growth factor (IGF)-1 [RRquartile 4 vs. 1 = 0.73; 95% confidence interval (CI), 0.60–0.87; P-trend = 0.002/FDR = 0.04], HbA1c (RRquartile 4 vs. 1 = 0.74; 95% CI, 0.62–0.89; P-trend = 0.002/FDR = 0.04), and alanine aminotransferase (RRquartile 4 vs. 1 = 0.76; 95% CI, 0.63–0.91; P-trend = 0.002/FDR = 0.04) were significantly associated with lower ovarian cancer risk. When stratified by histotype, higher IGF1 levels were associated with lower risk of serous (RRquartile 4 vs. 1 = 0.73; 95% CI, 0.58–0.91; P-trend = 0.01/FDR = 0.20) and clear cell tumors (RRquartile 4 vs. 1 = 0.18; 95% CI, 0.07–0.49; P-trend = 0.001/FDR = 0.02), and higher HbA1c levels were associated with lower risk of serous tumors (RRquartile 4 vs. 1 = 0.73; 95% CI, 0.59–0.90; P-trend = 0.004/FDR = 0.08). Conclusions: We observed that higher levels of circulating IGF1, HbA1c, and alanine aminotransferase were associated with lower ovarian cancer risk. Impact: These results suggest metabolism of glucose/amino acid and insulin/IGF1 signaling pathway may be contributing to ovarian carcinogenesis. Further research is needed to replicate our findings and elucidate how systemic changes in metabolism impact ovarian carcinogenesis.

Associations between common contraceptive use and circulating inflammatory biomarkers

Abstract Ovarian cancer incidence has declined in recent decades, due in part to oral contraceptive (OC) use and tubal ligation. However, intrauterine device (IUD) use has increasingly replaced OC use. As ovarian cancer is an inflammation-related disease, we examined the association of OC use, IUD use, and tubal ligation with plasma levels of C-reactive protein (CRP), interleukin 6, and soluble tumor necrosis factor α receptor 2 in the Nurses’ Health Study (NHS) and NHSII. After adjusting for reproductive, hormonal, and lifestyle factors and mutual adjustment for other methods of contraception, there were no differences in inflammatory markers between ever and never use of each method. However, CRP levels decreased from an average of 30.4% (95% CI, –53.6 to 4.4) with every 5 years since initial IUD use (P-trend = .03), while CRP increased an average of 9.9% (95% CI, 5.7, 14.3) with every 5 years of use of OC (P-trend < .0001) as well as differences by body mass index and menopausal status. Our results suggest IUD use and tubal ligation are not associated with higher circulating inflammatory markers long term, although long duration of OC use may increase generalized inflammation, which may in part explain why its protective effect wanes over time. This article is part of a Special Collection on Gynecological Cancer.

A Prospective Study Consortium for the Discovery and Validation of Early Detection Markers for Ovarian Cancer – Baseline Findings for CA125

Abstract Purpose: Epithelial ovarian cancer (EOC) is a lethal malignancy. Cancer antigen 125 (CA125), the “best” available marker for detecting EOC, has insufficient sensitivity and specificity for earlier-stage disease and is not a meaningful screening tool, motivating the search for further biomarkers. Cancer biomarker discovery is enhanced by “omics” technologies. Discovery studies for EOC biomarkers should be conducted in prediagnosis blood samples from prospective cohorts to maximize the likelihood of identifying markers that can detect disease before usual diagnosis and in earlier disease stage while reducing methodologic biases. Experimental Design: Individual cohorts with prediagnosis blood samples have insufficient sample size for such studies. Thus, we established “Prospective Early Detection Consortium for Ovarian Cancer” (“PREDICT”)—a collaboration of nine prospective studies—to assemble a sufficient number of EOC cases with blood samples collected ≤18 months before diagnosis plus controls. The 457 cases and 1,687 controls have circulating CA125 measured using a clinical assay. Results: The discrimination capacity for single CA125 measurements in samples collected <6 months prior to diagnosis was high (AUC; PREDICT overall = 0.92; range across cohorts of nonpregnant individuals = 0.89–0.98) and declined with extended time between blood collection and diagnosis. Between-cohort variability in CA125 levels and predictive performance was observed. Conclusions: Ongoing investigations in PREDICT are evaluating the early detection potential of tumor-associated autoantibodies and miRNAs using CA125 as a benchmark. PREDICT is a well-characterized resource for identifying and validating detection markers for EOC that may then be used in multimodal screening as a complement to CA125 and combined with imaging.

Associations between prediagnostic aspirin use and ovarian tumor gene expression

AbstractBackgroundAspirin use has been associated with reduced ovarian cancer risk, yet the underlying biological mechanisms are not fully understood. To gain mechanistic insights, we assessed the association between prediagnosis low and regular‐dose aspirin use and gene expression profiles in ovarian tumors.MethodsRNA sequencing was performed on high‐grade serous, poorly differentiated, and high‐grade endometrioid ovarian cancer tumors from the Nurses' Health Study (NHS), NHSII, and New England Case–Control Study (n = 92 cases for low, 153 cases for regular‐dose aspirin). Linear regression identified differentially expressed genes associated with aspirin use, adjusted for birth decade and cohort. False discovery rates (FDR) were used to account for multiple testing and gene set enrichment analysis was used to identify biological pathways.ResultsNo individual genes were significantly differentially expressed in ovarian tumors in low or regular‐dose aspirin users accounting for multiple comparisons. However, current versus never use of low‐dose aspirin was associated with upregulation of immune pathways (e.g., allograft rejection, FDR = 5.8 × 10−10; interferon‐gamma response, FDR = 2.0 × 10−4) and downregulation of estrogen response pathways (e.g., estrogen response late, FDR = 4.9 × 10−8). Ovarian tumors from current regular aspirin users versus never users were also associated with upregulation in interferon pathways (FDR <1.5 × 10−4) and downregulation of multiple extracellular matrix (ECM) architecture pathways (e.g., ECM organization, 4.7 × 10−8).ConclusionOur results suggest low and regular‐dose aspirin may impair ovarian tumorigenesis in part via enhancing adaptive immune response and decreasing metastatic potential supporting the likely differential effects on ovarian carcinogenesis and progression by dose of aspirin.

CCNE1 and survival of patients with tubo‐ovarian high‐grade serous carcinoma: An Ovarian Tumor Tissue Analysis consortium study

AbstractBackgroundCyclin E1 (CCNE1) is a potential predictive marker and therapeutic target in tubo‐ovarian high‐grade serous carcinoma (HGSC). Smaller studies have revealed unfavorable associations for CCNE1 amplification and CCNE1 overexpression with survival, but to date no large‐scale, histotype‐specific validation has been performed. The hypothesis was that high‐level amplification of CCNE1 and CCNE1 overexpression, as well as a combination of the two, are linked to shorter overall survival in HGSC.MethodsWithin the Ovarian Tumor Tissue Analysis consortium, amplification status and protein level in 3029 HGSC cases and mRNA expression in 2419 samples were investigated.ResultsHigh‐level amplification (>8 copies by chromogenic in situ hybridization) was found in 8.6% of HGSC and overexpression (>60% with at least 5% demonstrating strong intensity by immunohistochemistry) was found in 22.4%. CCNE1 high‐level amplification and overexpression both were linked to shorter overall survival in multivariate survival analysis adjusted for age and stage, with hazard stratification by study (hazard ratio [HR], 1.26; 95% CI, 1.08‐1.47, p = .034, and HR, 1.18; 95% CI, 1.05‐1.32, p = .015, respectively). This was also true for cases with combined high‐level amplification/overexpression (HR, 1.26; 95% CI, 1.09‐1.47, p = .033). CCNE1 mRNA expression was not associated with overall survival (HR, 1.00 per 1‐SD increase; 95% CI, 0.94‐1.06; p = .58). CCNE1 high‐level amplification is mutually exclusive with the presence of germline BRCA1/2 pathogenic variants and shows an inverse association to RB1 loss.ConclusionThis study provides large‐scale validation that CCNE1 high‐level amplification is associated with shorter survival, supporting its utility as a prognostic biomarker in HGSC.

Measurement of Ovarian Tumor Immune Profiles by Multiplex Immunohistochemistry: Implications for Epidemiologic Studies

Abstract Background: Despite the immunogenic nature of many ovarian tumors, treatment with immune checkpoint therapies has not led to substantial improvements in ovarian cancer survival. To advance population-level research on the ovarian tumor immune microenvironment, it is critical to understand methodologic issues related to measurement of immune cells on tissue microarrays (TMA) using multiplex immunofluorescence (mIF) assays. Methods: In two prospective cohorts, we collected formalin-fixed, paraffin-embedded ovarian tumors from 486 cases and created seven TMAs. We measured T cells, including several sub-populations, and immune checkpoint markers on the TMAs using two mIF panels. We used Spearman correlations, Fisher exact tests, and multivariable-adjusted beta-binomial models to evaluate factors related to immune cell measurements in TMA tumor cores. Results: Between-core correlations of intratumoral immune markers ranged from 0.52 to 0.72, with more common markers (e.g., CD3+, CD3+CD8+) having higher correlations. Correlations of immune cell markers between the whole core, tumor area, and stromal area were high (range 0.69–0.97). In multivariable-adjusted models, odds of T-cell positivity were lower in clear cell and mucinous versus type II tumors (ORs, 0.13–0.48) and, for several sub-populations, were lower in older tissue (sample age > 30 versus ≤ 10 years; OR, 0.11–0.32). Conclusions: Overall, high correlations between cores for immune markers measured via mIF support the use of TMAs in studying ovarian tumor immune infiltration, although very old samples may have reduced antigenicity. Impact: Future epidemiologic studies should evaluate differences in the tumor immune response by histotype and identify modifiable factors that may alter the tumor immune microenvironment.

Common Analgesic Use for Menstrual Pain and Ovarian Cancer Risk

Abstract Menstrual pain has been associated with increased ovarian cancer risk, presumably through increased inflammation, which is known to play a critical role in ovarian carcinogenesis. Analgesic medications are frequently used to treat menstrual pain, some of which lower ovarian cancer risk. In this study, we examined the association between analgesic use for menstrual pain during the premenopausal period and ovarian cancer risk among women with history of menstrual pain. We used data from the New England Case-Control Study, including 1,187 epithelial ovarian cancer cases and 1,225 population-based controls enrolled between 1998 and 2008 with detailed information on analgesic use for their menstrual pain. We used unconditional logistic regression to calculate the odds ratios (ORs) and 95% confidence intervals (CIs) for the association between analgesic use (i.e., aspirin, ibuprofen, acetaminophen) for menstrual pain and ovarian cancer risk. We further conducted a stratified analysis by intensity of menstrual pain (mild/moderate, severe). Among women with menstrual pain during their 20s and 30s, ever use of analgesics for menstrual pain was not significantly associated with ovarian cancer risk. However, among women with severe menstrual pain, ever use of aspirin or acetaminophen for menstrual pain was inversely associated with risk (OR, 0.41; 95% CI, 0.18–0.94 and OR, 0.43; 95% CI, 0.21–0.88 compared with never users, respectively). No significant association was observed between analgesic use and ovarian cancer risk among women with mild/moderate menstrual pain (Pinteraction ≤ 0.03). Our results suggest that use of aspirin or acetaminophen for severe menstrual pain may be associated with lower risk of ovarian cancer. Prevention Relevance: This study investigates whether analgesic use specifically for menstrual pain during the premenopausal period influences ovarian cancer risk. Our results suggest use of aspirin or acetaminophen for severe menstrual pain may be associated with lower risk of ovarian cancer among women with severe menstrual pain.

Prospective Analyses of Lifestyle Factors Related to Energy Balance and Ovarian Cancer Risk by Infiltration of Tumor-Associated Macrophages

Abstract Background: Lifestyle factors related to energy balance have been associated with ovarian cancer risk and influence the tumor immune microenvironment, including tumor-associated macrophages (TAM). However, no studies have assessed whether these factors differentially impact ovarian cancer risk by TAM densities. Methods: We conducted a prospective analysis in the Nurses' Health Studies to examine the associations of physical activity, sitting time, and a food-based empirical dietary inflammatory pattern (EDIP) score with invasive epithelial ovarian cancer risk by TAM density assessed by immunohistochemistry. We considered density of CD68 (marker of total TAMs) and CD163 (marker of pro-carcinogenic M2-type TAMs), and their ratios. We used multivariable Cox proportional hazards regression to calculate hazard ratios (HR) and 95% confidence intervals (CI) of exposures with risk of ovarian tumors with high versus low TAMs, including analyses stratified by body mass index. Results: Analyses included 312 incident ovarian cancer cases with TAM measurements. Physical activity, sitting time, and EDIP score were not differentially associated with ovarian cancer risk by TAM densities (Pheterogeneity > 0.05). Among overweight and obese women, higher EDIP score was associated with increased risk of CD163 low-density tumors (HR comparing extreme tertiles, 1.57; 95% CI, 0.88–2.80; Ptrend = 0.01), but not CD163 high-density tumors (comparable HR, 1.16; 95% CI, 0.73–1.86; Ptrend = 0.24), though this difference was not statistically significant (Pheterogeneity = 0.22). Conclusions: We did not observe differential associations between lifestyle factors and ovarian cancer risk by TAM densities. Impact: Future investigations examining the interplay between other ovarian cancer risk factors and the tumor immune microenvironment may help provide insight into ovarian cancer etiology.

Pre-diagnosis and post-diagnosis dietary patterns and survival in women with ovarian cancer

Evidence is limited on inflammation-related dietary patterns and mortality in ovarian cancer survivors. We examined the associations between pre- and post-diagnosis dietary patterns, including change in diet from before to after diagnosis, and mortality among 1003 ovarian cancer survivors in two prospective cohort studies. Dietary pattern scores for empirical dietary inflammatory pattern (EDIP) and Alternative Healthy Eating Index (AHEI) were calculated based on food frequency questionnaires. We used Cox proportional hazard models to calculate hazard ratios (HRs) and 95% confidence intervals (CIs) for ovarian cancer-specific and all-cause mortality. Pre-diagnosis EDIP score and AHEI were not associated with mortality. Among non-high grade serous cases, a higher post-diagnosis EDIP score was associated with increased risk of all-cause mortality (HR Consuming a more inflammatory dietary pattern post-diagnosis was associated with increased mortality in ovarian cancer survivors, suggesting limiting the inflammatory potential of diet post-diagnosis could lead to enhanced survivorship.

Associations between Parity, History of Breastfeeding, and T-cell Profile of Ovarian Tumors

Abstract Background: Parity and breastfeeding are associated with systemic changes in maternal inflammation and reduced risk of ovarian cancer, but little is known about their impact on the ovarian tumor immune microenvironment. Methods: We evaluated the associations of self-reported parity and history of breastfeeding with tumor-infiltrating T cells among 1,706 ovarian carcinoma cases with tumor tissue collected across four studies. The abundance of tumor-infiltrating T cells was measured by multiplex immunofluorescence in tumor tissue microarrays. ORs and 95% confidence intervals (CI) for the positivity of tumor immune cells were calculated using beta-binomial models and stratified by histotype. Results: Compared with ovarian tumors in nulliparous women, there was no association between parity and ovarian tumor T-cell abundance among all histotypes combined but suggestion of increased cytotoxic T cells and T-cell exhaustion among parous women with clear-cell tumors. When restricted to parous women, history of breastfeeding was associated with increased odds for all T-cell types [i.e., total T, cytotoxic T, helper T (Th), regulatory T, and exhausted T cells], with ORs ranging from 1.11 to 1.42. For every 6 months of breastfeeding, we observed increased odds of activated Th-cell infiltration (CD3+CD4+CD69+; OR, 1.13, 95% CI, 0.99–1.29), with a similar association for high-grade serous tumors, but lower odds in clear-cell tumors (OR, 0.43, 95% CI, 0.21–0.87). Conclusions: History of breastfeeding may alter the ovarian tumor immune microenvironment by modulating the abundance of tumor-infiltrating T cells. Impact: Although replication is required, history of breastfeeding may play a role in the activation of the ovarian tumor immune response.