Esther Elishaev

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.

Assessment of reprocessed ThinPrep cytology cases after glacial acetic acid wash procedure using the Hologic Genius Digital Diagnostics System

This study focuses on ThinPrep Pap tests with a low to borderline number of cells and the performance of AI-assisted digital systems in cases that have undergone the acetic acid wash procedure (AAW). Four hundred sixty-two cases initially interpreted as unsatisfactory and finally interpreted as satisfactory after AAW procedure were included in the study. These ThinPrep Pap slides were scanned using the Genius Digital Diagnostic System (GDDS). Overall agreement between GDDS and Original ThinPrep Interpretation (OTPI) was 63.2% for diagnostic match (Negative for Intraepithelial Lesion, ASCUS, Low Grade Squamous Intraepithelial Lesion, Atypical Squamous Cells, High Grade, Atypical Glandular Cells, or unsatisfactory), and 66.0% when ASCUS + diagnoses are grouped. Out of the 462 cases, 364 (78.8%) were called Negative for Intraepithelial Lesion based upon the manual OTPI, as opposed to 310 (67.1%) reviewed using the GDDS. There were 17.5%, 1.3%, 0.9% and 1.5% cases called Atypical Squamous Cells of Undetermined Significance, Low Grade Squamous Intraepithelial Lesion, Atypical Squamous Cells, High Grade Cannot be Excluded, and Atypical Glandular Cells respectively on OTPI, as opposed to 24.7%, 3.0%, 0.6% and 1.5% respectively by the GDDS. Only 3.0% of the cases were deemed unsatisfactory by GDDS. All the cases with high grade results in the subsequent cervical biopsy were diagnosed as at least Atypical Squamous Cells of Undetermined Significance and above by the GDDS. The diagnostic agreement between GDDS and biopsy was 65.2% compared to 58.7% for OTPI, although this is not statistically significantly different, (χ Our results demonstrate that the GDDS can be successfully used to screen ThinPrep Pap Tests that have undergone the AAW procedure.

Assessment of the efficacy and accuracy of cervical cytology screening with the Hologic Genius Digital Diagnostics System

Abstract Background Medical technologies powered by artificial intelligence are quickly transforming into practical solutions by rapidly leveraging massive amounts of data processed via deep learning algorithms. There is a necessity to validate these innovative tools when integrated into clinical practice. Methods This study evaluated the performance of the Hologic Genius Digital Diagnostics System (HGDDS) with a cohort of 890 previously reviewed and diagnosed ThinPrep Papanicolaou (Pap) tests with the intent to deploy this system for routine clinical use. The study included all diagnostic categories of The Bethesda System, with follow‐up tissue sampling performed within 6 months of abnormal Pap test results to serve as the ground truth. Results The HGDDS demonstrated excellent performance in detecting significant Pap test findings, with close to 100% sensitivity (98.2%–100%) for cases classified as atypical squamous cells of undetermined significance and above within a 95% confidence interval and a high negative predictive value (92.4%–100%). Conclusions The HGDDS streamlined workflow, reduced manual workload, and functioned as a stand‐alone system.

Endometriosis-Associated Mesenchymal Stem Cells Support Ovarian Clear Cell Carcinoma through Iron Regulation

Abstract Ovarian clear cell carcinoma (OCCC) is a deadly and treatment-resistant cancer, which arises within the unique microenvironment of endometriosis. In this study, we identified a subset of endometriosis-derived mesenchymal stem cells (enMSC) characterized by loss of CD10 expression that specifically support OCCC growth. RNA sequencing identified alterations in iron export in CD10-negative enMSCs and reciprocal changes in metal transport in cocultured OCCC cells. CD10-negative enMSCs exhibited elevated expression of iron export proteins hephaestin and ferroportin and donate iron to associated OCCCs, functionally increasing the levels of labile intracellular iron. Iron is necessary for OCCC growth, and CD10-negative enMSCs prevented the growth inhibitory effects of iron chelation. In addition, enMSC-mediated increases in OCCC iron resulted in a unique sensitivity to ferroptosis. In vitro and in vivo, treatment with the ferroptosis inducer erastin resulted in significant death of cancer cells grown with CD10-negative enMSCs. Collectively, this work describes a novel mechanism of stromal-mediated tumor support via iron donation. This work also defines an important role of endometriosis-associated MSCs in supporting OCCC growth and identifies a critical therapeutic vulnerability of OCCC to ferroptosis based on stromal phenotype. Significance: Endometriosis-derived mesenchymal stem cells support ovarian clear cell carcinoma via iron donation necessary for cancer growth, which also confers sensitivity to ferroptosis-inducing therapy.

Molecular Subclasses of Clear Cell Ovarian Carcinoma and Their Impact on Disease Behavior and Outcomes

Abstract Purpose: To identify molecular subclasses of clear cell ovarian carcinoma (CCOC) and assess their impact on clinical presentation and outcomes. Experimental Design: We profiled 421 primary CCOCs that passed quality control using a targeted deep sequencing panel of 163 putative CCOC driver genes and whole transcriptome sequencing of 211 of these tumors. Molecularly defined subgroups were identified and tested for association with clinical characteristics and overall survival. Results: We detected a putative somatic driver mutation in at least one candidate gene in 95% (401/421) of CCOC tumors including ARID1A (in 49% of tumors), PIK3CA (49%), TERT (20%), and TP53 (16%). Clustering of cancer driver mutations and RNA expression converged upon two distinct subclasses of CCOC. The first was dominated by ARID1A-mutated tumors with enriched expression of canonical CCOC genes and markers of platinum resistance; the second was largely comprised of tumors with TP53 mutations and enriched for the expression of genes involved in extracellular matrix organization and mesenchymal differentiation. Compared with the ARID1A-mutated group, women with TP53-mutated tumors were more likely to have advanced-stage disease, no antecedent history of endometriosis, and poorer survival, driven by their advanced stage at presentation. In women with ARID1A-mutated tumors, there was a trend toward a lower rate of response to first-line platinum-based therapy. Conclusions: Our study suggests that CCOC consists of two distinct molecular subclasses with distinct clinical presentation and outcomes, with potential relevance to both traditional and experimental therapy responsiveness. See related commentary by Lheureux, p. 4838

Concurrent RB1 Loss and BRCA Deficiency Predicts Enhanced Immunologic Response and Long-term Survival in Tubo-ovarian High-grade Serous Carcinoma

Abstract Purpose: The purpose of this study was to evaluate RB1 expression and survival across ovarian carcinoma histotypes and how co-occurrence of BRCA1 or BRCA2 (BRCA) alterations and RB1 loss influences survival in tubo-ovarian high-grade serous carcinoma (HGSC). Experimental Design: RB1 protein expression was classified by immunohistochemistry in ovarian carcinomas of 7,436 patients from the Ovarian Tumor Tissue Analysis consortium. We examined RB1 expression and germline BRCA status in a subset of 1,134 HGSC, and related genotype to overall survival (OS), tumor-infiltrating CD8+ lymphocytes, and transcriptomic subtypes. Using CRISPR-Cas9, we deleted RB1 in HGSC cells with and without BRCA1 alterations to model co-loss with treatment response. We performed whole-genome and transcriptome data analyses on 126 patients with primary HGSC to characterize tumors with concurrent BRCA deficiency and RB1 loss. Results: RB1 loss was associated with longer OS in HGSC but with poorer prognosis in endometrioid ovarian carcinoma. Patients with HGSC harboring both RB1 loss and pathogenic germline BRCA variants had superior OS compared with patients with either alteration alone, and their median OS was three times longer than those without pathogenic BRCA variants and retained RB1 expression (9.3 vs. 3.1 years). Enhanced sensitivity to cisplatin and paclitaxel was seen in BRCA1-altered cells with RB1 knockout. Combined RB1 loss and BRCA deficiency correlated with transcriptional markers of enhanced IFN response, cell-cycle deregulation, and reduced epithelial–mesenchymal transition. CD8+ lymphocytes were most prevalent in BRCA-deficient HGSC with co-loss of RB1. Conclusions: Co-occurrence of RB1 loss and BRCA deficiency was associated with exceptionally long survival in patients with HGSC, potentially due to better treatment response and immune stimulation.