BRCA1 Mutation Carriers Research Articles

Abstract 7064: BRCA 1/2 haploinsufficiency-associated metabolic dysfunction in benign fallopian tube epithelia

Abstract In 3-17% of women with BRCA1/2 mutations who undergo risk-reducing surgery, pre-neoplastic and neoplastic lesions are found in their fallopian tubes that display differential loss of heterozygosity of BRCA1/2. Precisely how mutations in BRCA1/2 predispose women to develop high grade serous ovarian carcinoma (HGSC) is not completely understood. Although linked to various processes localized in the nucleus, BRCA1/2 may also have essential roles in the cytoplasm, specifically in the regulation of key processes associated with mitochondrial homeostasis and metabolism. Understanding how haploinsufficiency of BRCA1/2 contributes to the potential neoplastic transformation of fallopian tube epithelial cells should reveal new insights into BRCA1/2 function, and ultimately provide novel opportunities for preventative interventions in individuals with this mutation and new treatment opportunities for HGSC associated with BRCA1/2 heterozygosity. We have identified distinct metabolic and mitochondrial deficits in BRCA2 mutation carriers compared to BRCA1 carriers and non-carriers. Fallopian tube epithelial cell lines derived from BRCA2 mutation carriers showed increased levels of mitochondrial respiration and activity followed by BRCA1 mutation carriers and non-carriers. Western blot analysis of mitochondrial proteins regulating glycolysis and oxidative phosphorylation showed differential expression based on mutation status. Transmission electron microscopy of BRCA2 mutation carriers showed increased disrupted mitochondrial networks when compared to wild type, suggesting that mutation status may contribute to increased mitochondrial fission. These observations reveal an entirely new aspect of BRCA1 and BRCA2 biology and further studies will disaggregate the effects of BRCA1 and BRCA2 on mitochondrial function, versus DNA damage repair. Ultimately, this work will provide new insight into the adaptive changes within FT cells which promote neoplastic transformation and inform how high-grade serous carcinoma (HGSC) cells distinctively respond to microenvironmental stimuli. Citation Format: Jovanka Ravix, Iru Paudel, Gauthami Pulivendala, Ramlogan Sowamber, Elizabeth Smith, Matthew Schlumbrecht, David Lombard, Sophia George. BRCA 1/2 haploinsufficiency-associated metabolic dysfunction in benign fallopian tube epithelia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7064.

Abstract 7237: Peptide-mRNA complex mediated restoration of BRCA-1 tumor suppressor function in BRCA-1 mutated cancers as a new therapeutic strategy

Abstract BRCA1 and BRCA-2 proteins are involved in the repair of DNA damage such as double-strand breaks during S and G2 phases, by intervening in homologous recombination (HR) steps. Breast cancers arising from BRCA-1 mutation carriers (10% of BC) and from epigenetically silenced BRCA-1 (30% of sporadic BC) are associated with a lack of expression and/or function of BRCA-1, which induces genomic instability. Herein, we describe a new potent strategy combining mRNAs with a tumor selective peptide nanocarrier, to selectively rescue BRCA-1 function as potential therapeutic approach in cancers. Proprietary BRCA-1 derived mRNA were complexed with proprietary short amphipathic peptides and the resulting nanoparticles were evaluated on several breast cancer cells, harboring different BRCA-1 alterations. Sensitivity to Veliparib (PARPi) following the nanoparticle treatments were evaluated on PARPI resistant and PARPi sensitive cells. The recruitment of RAD51 to sites of double stranded DNA break was quantified by immunofluorescence. In-vivo efficacy of varying sequences of BRCA-1 mRNA-peptide nanoparticles (2.0 mg/kg, weekly, IV; ADGN-901, -902, -903) and veliparib (50mg/kg, daily, oral) were evaluated on breast cancer (HCC1937-BRCA-1 deficient, PARPi resistant) mouse xenografts. We showed that ADGN-901 inhibits the proliferation of BRCA-1 deficient cell lines from 20 to 60% depending on the type of BRCA-1 alterations. ADGN-902, specifically reduces the proliferation of cells expressing N-terminal truncated BRCA-1, by 50 to 60% and ADGN-903 inhibits proliferation >60% of all cells harboring defects in BRCA-1. Interestingly, ADGN-901 and ADGN-903 were able to resensitize the PARPi resistant cells to veliparib inhibition. We demonstrated that ADGN-903 restores BRCA-1 function in homologous recombination. ADGN-903 prevents RAD51 recruitment to the damage site by reducing RAD51 protein level as well as formation of RAD51 foci following irradiation. IV administration of ADGN-901 and ADGN-903 (2.0 mg/kg) had a strong inhibitory effect on HCC-1937 tumor growth while ADGN-902 and veliparib were inactive. ADGN-901 reduced tumor growth by 95% and ADGN-903 abolished tumor growth with strong tumor regressions. ADGN-901 and ADGN-903, were also evaluated on large tumors of about 500-600 mm3 and were highly effective in preventing further progression and achieving shrinkage of large tumors. ADGN-901 and ADGN-903 treatments are well tolerated, without inducing clinical toxicity. BRCA-1 mRNA-peptide nanoparticles are effective in rescuing BRCA-1 function and inducing tumor regressions in BRCA-1 altered breast cancer cells and tumors. Our study provides a proof-of-concept that restoration of wild type BRCA-1 expression could be a potent therapeutic strategy for BRCA-1 altered tumors whether or not they are PARPi resistant. Citation Format: Gilles Divita, Audrey Grunenberger, Léa Cabrera, Nathalie Durany, Neil Desai. Peptide-mRNA complex mediated restoration of BRCA-1 tumor suppressor function in BRCA-1 mutated cancers as a new therapeutic strategy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7237.

Abstract 2137: Racial disparities in triple-negative breast cancer (TNBC) mortality rates in Hampton Roads Virginia and the effects of SIAH expression on prognosis

Abstract Background: While high-resolution imaging and advancements in therapies have significantly improved breast cancer survival rates, 43,170 breast cancer patients will die in the United States in 2023 alone. Triple-negative breast cancer (TNBC) is the most aggressive breast cancer subtype and disproportionately affects BRCA1 mutation carriers and young black women. Black/African American (AA) patients have the highest mortality and the shortest survival of any racial/ethnic group in the US. Persistent cancer racial disparities remain due to a variety of risk factors. The SEER/CDC data show an 8% lower breast cancer incidence rate and a 41% higher mortality rate in Black/AA patients compared to their white counterparts. Methods: Chart review was conducted using Sentara MD Office/EPIC and VOA iKnowMedicine portals to update tumor relapse, metastasis, and survival in 577 TNBC patients. Supported by strong evidence in developmental, evolutionary, and cancer biology, we hypothesize that persistent EGFR/K-RAS/SIAH pathway activation is a major driving force of TNBC malignancy, racial disparity, early relapse, and high mortality. We propose to integrate SIAH expression to augment the existing clinicopathological parameters to improve patient risk stratification, therapy quantification, and relapse/survival prediction at the 1st-line neoadjuvant settings. Results: We report that cancer disparity and high mortality rates are even more pronounced in our racially-diverse communities in Hampton Roads Virginia. We discovered that SIAH is a tumor-specific, therapy-responsive, and prognostic biomarker in TNBC. High SIAH expression in residual tumors reflects tumor-driving EGFR/K-RAS/SIAH pathway activation (ON) that will predict cancer disparity, treatment resistance, early relapse, and poor survival. Low SIAH expression in residual tumors reflect EGFR/K-RAS/SIAH pathway inactivation (OFF) that will predict tumor remission and prolonged survival. Conclusion: We detect a major racial disparity of TNBC patients at Sentara-EVMS-VOA. Our local Black/AA TNBC patients have a 1.6-fold higher mortality rate than their White counterparts. Encouraged by our preliminary data, we aim to develop a SIAH-centered biomarker panel by measuring the EGFR/K-RAS/SIAH pathway activation (ON)/inactivation (OFF), and use SIAH as a new prognostic biomarker to risk stratify patients, detect cancer racial disparities, forecast tumor relapse, and predict patient survival at 1st-line neoadjuvant settings. By targeting SIAH, individualized treatments can be created to allow for increased survival rates in at risk individuals with more aggressive TNBCs. Citation Format: Claire B. Piatak, Taylor N. Drake, Ashleigh Hannah, Jonathan Baker, Caroline L. Dasom, Emily L. Breeding, Janet S. Winston, Billur Samli, Rick Jansen, Michael Danso, Richard A. Hoefer, Amy H. Tang. Racial disparities in triple-negative breast cancer (TNBC) mortality rates in Hampton Roads Virginia and the effects of SIAH expression on prognosis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2137.

Abstract 5596: Exploring BRCA1-Y1853ter and potential therapeutic targets in BRCA1-BRCT mutated cancers

Abstract The breast cancer gene BRCA1 plays a pivotal role as a tumor suppressor gene. Despite the use of poly (ADP-ribose) polymerase inhibitors (PARPi) in treating BRCA1 mutation carriers, the lack of response or the development of acquired drug resistance underscores the pressing need for new therapeutic targets. One intriguing BRCA1 mutation, BRCA1-Y1853ter, characterized by a minimal truncation, has been identified in multiple individuals with Hereditary Breast and Ovarian Cancer (HBOC) syndrome. We have developed a Halo Tag and adenovirus-based system where wild-type BRCA1 (WT BRCA1) and mutant BRCA1 protein complexes can be studied in multiple cell types. Here, we used this system to affinity purify BRCA1 wild-type and mutant complexes, followed by label-free quantitative proteomics to investigate the dynamic interactome variations between WT BRCA1 and BRCA1 mutant. Contrary to the assumed loss of function, our preliminary findings suggest a potential gain of function, as a significant number of candidate proteins exhibit enhanced binding affinity with the mutant BRCA1. We are currently exploring whether these distinct binding characteristics are shared among various BRCA1-BRCT mutants, which may hint at a common tumorigenesis mechanism. Our overarching hypothesis is that identifying a specific binding partner, shared among various BRCA-BRCT mutants, could unveil a novel therapeutic target with broader applicability in treating BRCA1-related malignancies. Citation Format: Jingjing Chen. Exploring BRCA1-Y1853ter and potential therapeutic targets in BRCA1-BRCT mutated cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5596.

Abstract 7664: Early detection of racial disparity and treatment resistance in triple negative breast cancer

Abstract Introduction: Triple-negative breast cancer (TNBC) is an aggressive breast cancer subtype that disproportionately affects BRCA1 mutation carriers and young Black/African American (AA) women. Black/AA patients have the highest mortality and the shortest survival of any racial/ethnic group in the US. With the approval of pembrolizumab, the addition of immune checkpoint blockade (ICB) to neoadjuvant chemotherapy (NACT) has become the standard of care (SOC) for high-risk early-stage TNBC. The pathologic complete response (pCR) is a reliable prognostic marker that correlates with tumor remission and good outcome, whereas pathologic incomplete response (pIR) with high residual cancer burden (RCB) is often associated with early relapse, treatment resistance, and poor survival. Many similarly-treated patients with identical TNM and RCB classifications still experience racial disparity, different relapse rates, and disparate survival. Current methods fall short in predicting tumor recurrence/chemo-resistance/patient survival with high accuracy in the clinic. Methods: We detected a major racial disparity in our large regional cohorts of TNBC patients (> 1,000 patients), of which 48% patients are Black/AA and 48% patients are white patients, at the Sentara Cancer Network and VCU Massey Comprehensive Cancer Center, that have historically provided excellent healthcare and quality cancer care to the social-economically-disadvantaged, medically underserved, and underinsured racial/ethnic minorities in low income communities in Virginia. Despite Sentara/VCU’s best efforts, Black/AA breast cancer patients still suffer 70-100% higher MBC mortality, and the worst survival compared to their white counterparts, based on our clinical studies and the SEER/CDC databases. Result: We showed that persistent EGFR/RAS/SIAH pathway activation is a major driving force of TNBC malignancy. The most downstream and evolutionarily conserved signaling gatekeeper, SIAH, is an RING domain E3 ligase that is a high-resolution prognostic biomarker for patient risk stratification and racial disparity prediction in TNBC. High SIAH expression in residual tumors reflects ineffective NACT/IO therapy and persistent EGFR/K-RAS/SIAH pathway activation (ON) that drives tumor relapse, resistance, and reduced survival. Low SIAH expression in residual tumors reflects effective NACT/IO therapy and EGFR/K-RAS/SIAH pathway inactivation (OFF) that predicts tumor remission, good outcome, and prolonged survival. We found that SIAH expression is 2.0-fold higher in Black/AA patients’ residual tumors than in the white patients’ residual tumors, mirroring a 1.8-2.0-fold higher death rates in the Black/AA patients than the white patients. Conclusion: In this study, we demonstrate the prognostic power of SIAH for patient risk stratification, therapy quantification, and relationship to cancer racial disparity in TNBC. Citation Format: Amy H. Tang, Richard A. Hoefer, Mary L. Guye, Billur Samli, Janet S. Winston, Jennifer Koblinski, Valentina Robila, Michael O. Idowu, Rick J. Jansen, Harry D. Bear. Early detection of racial disparity and treatment resistance in triple negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7664.

Abstract 4291: Bai1 inhibition promotes ferroptosis of breast cancer cells to prevent tumor growth in human xenograft tumor models

Abstract The adhesion G-protein coupled- receptor 1 ADGRB1/Bai1 was identified as a phosphatidylserine receptor that cooperates with ELMO/Dock180/Rac to promote maximal engulfment of apoptotic cells. This function of Bai1 was also shown to promote myoblasts fusion. Cell-cell fusion has been demonstrated to occur in normal physiological functions as well as in diseases. Studies have shown in cancer that cell-cell fusion increases tumor heterogeneity, drug resistance, and metastasis as hybrids acquire new properties that allow them to migrate and proliferate at new sites. Activation of ELMO-Dock signaling pathway has been associated with breast cancer metastasis. Moreover, ELMO1 has been identified as a modifier of breast cancer risk for BRCA mutation carriers. Rac1 was also shown to be overexpressed in proliferative breast disease, preinvasive and invasive breast carcinoma as well as lymph node metastases. Our recent studies showed that breast cancer cell fusion with bone marrow-derived mesenchymal stem cells was modulated by hypoxic conditions and promoted by apoptosis. Our hypothesis is that hypoxia stress induced apoptosis activates the phosphatidylserine receptor Bai1, and it signals through the ELMO/DOCK180/Rac to promote breast cancer cell fusion-driven tumor heterogeneity and metastasis. In this study, we determined the ability of Bai1 depleted breast cancer cells to induce tumor growth in immunocompromised mice. We used Bai1 sgRNA to inhibit Bai1 expression in the breast cancer cell MDA-MB-231. These cells (1x 106) together with bone marrow-derived mesenchymal stem cells (MSCs) (1x 106) were injected into the fat pad of the breast of 15 NOD SCID female mice. Control mice (15) received the same cells with no inhibition of Bai1. The mice were monitored for eight weeks. We found that Bai1 inhibition significantly reduced tumor growth in these mice in the course of time (P<0.05). The average tumor weight was also significantly reduced when Bai1 was inhibited (P<0.001). RNA sequencing of tumors generated from these mice revealed a significant increase in the expression of genes driving ferroptosis and a reduction in critical oncogenes in the experiment group compared to the control group. These findings will be helpful in the development of new classes of drugs effective in breast cancer treatment. Grant Support The NIH/NIMHD grant 5U54MD015929-03, the Society for Investigative Dermatology (SID) Freinkel Diversity Fellowship, and NSF Award Notice for Award ID 2142465 (awarded to FNK). Citation Format: Felicite Kamdem Noubissi, Oluwatowin Odubanjo, Hung-Chung Huang, Brenda Ogle, Paul Tchounwou. Bai1 inhibition promotes ferroptosis of breast cancer cells to prevent tumor growth in human xenograft tumor models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4291.

Abstract 4531: The synergistic potential of PARP inhibitors and irinotecan in small cell lung cancer

Abstract Background: Small cell lung cancer (SCLC) is highly aggressive and associated with a poor prognosis. While effective treatment is limited after a relapse from platinum-based chemotherapy, recent research has identified novel pathway targets including DNA damage repair (DDR) pathway. Poly (ADP-ribose) polymerase inhibitors (PARPi) have been currently evaluated for their efficacy as a monotherapy and in combination with other agents, such as immunotherapy. Methods: We evaluated the efficacy of combining PARPi and irinotecan in ten SCLC cell lines with diverse genetic backgrounds in the DDR genes. After treating the cells with three different PARPi in combination with irinotecan, we assessed cell viability and the combination effects. Furthermore, we confirmed protein changes in the DDR pathway through western blotting and measured the number of apoptotic cells using Annexin V staining. Results: In our experiments, talazoparib had the most potent effect at low concentrations, while olaparib and venadaparib showed more pronounced synergistic effects (average fold change of IC50: olaparib, 1649±4049; talazoparib, 25±34.21; venadaparib, 336±596.01; see Table). The combination of talazoparib or venadaparib with irinotecan led to a reduction of IC50 values to below 10 nM in 7 out of 10 tested cell lines. Three cell lines carrying BRCA mutations displayed a strong sensitivity to PARPi, whereas no significant correlation found with alterations in other DDR genes. The combined use of both drugs increased p-chk1 and p-p53 proteins, along with apoptosis signaling. Additionally, venadaparib alone did not induce cell toxicity but significantly increased apoptosis when combined with irinotecan, even at very low concentrations. Conclusions: The combination of a PARPi with irinotecan significantly inhibited cell growth more than the use of either agent alone. Our findings indicate that clinical trials are warranted to confirm the effectiveness of combining PARPi with irinotecan in patients with SCLC. IC50 values of PARP inhibitors in SCLC cells w/or w/o irinotecan 50 Nm IC50 (nM) Olaparib Talazoparib Venadaparib BRCA or PTEN status Irinotecan 50 nM - + - + - + BRCA1 deletion NCI-H1694 5,933 148.9 52.38 2.19 >10,000 1.02 BRCA2 deletion NCI-H211 392.8 37.20 2.28 0.99 3.82 0.94 BRCA2 deletion NCI-H1048 1,704 282.7 9.74 2.36 62.01 3.02 PTEN deletion NCI-H1341 >10,000 6,880 452.9 83.9 >10,000 >10,000 PTEN deletion DMS 79 2,382 221.9 18.55 3.4 175.16 3.07 no mutation NCI-H841 5,099 6,202 529.5 380.5 >10,000 >10,000 no mutation NCI-H2029 >10,000 >10,000 478.2 56.16 >10,000 >10,000 no mutation NCI-H2227 3,407 827.5 52.03 2.17 >10,000 0.69 no mutation NCI-H146 5,094 3.31 56.11 0.86 1,295 0.93 no mutation NCI-H209 579.2 0.05 5.24 0.05 8.18 0.04 Citation Format: Songji Oh, Miso Kim, Soyeon Kim, Bhumsuk Keam, Jeonghwan Youk, Tae Min Kim, Dong-Wan Kim, Dae Seog Heo. The synergistic potential of PARP inhibitors and irinotecan in small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4531.

Natural Killer Cell Dysfunction in Premenopausal BRCA1 Mutation Carriers: A Potential Mechanism for Ovarian Carcinogenesis.

Tissue-specificity for fimbrial fallopian tube ovarian carcinogenesis remains largely unknown in BRCA1 mutation carriers. We aimed to assess the cell autonomous and cell-nonautonomous implications of a germline BRCA1 mutation in the context of cancer immunosurveillance of CD3- CD56+ natural killer (NK) cells. Premenopausal BRCA1 mutation carriers versus age-matched non-carriers were compared. Daily urinary 5β-pregnanediol levels were used to determine progesterone metabolomics across an ovarian cycle. Using peripherally acquired NK cells the cell-mediated cytotoxicity of tumor targets (OVCAR-3, K-562) was determined using live cellular impedance (xCELLigence®) and multicolor flow cytometry. Hypoxia-inducible factor 1-alpha (HIF-1α) immunohistochemistry of cancer-free fallopian tube specimens allowed a comparison of proximal versus distal portions. Utilizing these findings the role of environmental factors relevant to the fimbrial fallopian tube (progesterone, hypoxia) on NK cell functional activity were studied in an ovarian phase-specific manner. BRCA1 mutation carriers demonstrate a differential progesterone metabolome with a phase-specific reduction of peripheral NK cell functional activity. Progesterone exposure further impairs NK cell-mediated cytotoxicity in a dose-dependent manner, which is reversed with the addition of mifepristone (1.25 µM). The fimbrial fallopian tube demonstrated significantly higher HIF-1α staining, particularly in BRCA1 mutation carriers, reflecting a site-specific 'hypoxic niche'. Exposure to hypoxic conditions (1% O2) can further impair tumor cytotoxicity in high-risk carriers. Phase-specific differential NK cell activity in BRCA1 mutation carriers, either systemically or locally, may favor site-specific pre-invasive carcinogenesis. These cumulative effects across a reproductive lifecycle in high-risk carriers can have a detrimental effect further supporting epidemiological evidence for ovulation inhibition.

Ovarian Cancer Risk-Reduction and Screening in BRCA1/2 Mutation Carriers.

Objective: To determine the utilization of risk-reducing strategies and screening protocols for ovarian cancer in female BRCA1/2 carriers. Methods: This study was a sub-analysis of female participants from a larger multicenter, cross-sectional survey of BRCA1/2 mutation carriers unaffected by cancer. The questionnaire was administered electronically via email at four institutions located in the northeast United States. Data were analyzed with Fisher's exact test. Results: The survey was completed by 104 female BRCA mutation carriers. BRCA subtypes included 54.3% BRCA2, 41.0% BRCA1, and 2.9% both. The age at which patients underwent genetic testing varied 21.2% were 18-24 years, 25.0% were 25-34 years, 29.8% were 35-44 years, and 24.0% were 45 years or older. Nearly, all respondents (97.1%) reported that a provider had discussed risk-reducing surgeries. Of the 79 females who underwent genetic testing before 45 years of age, 53.2% reported that a health care provider recommended taking combined oral contraceptive pills (COCs) to reduce their risk of ovarian cancer, and, of these women, 88.1% chose to use them. COCs were offered at higher rates among women who were younger at the age of genetic testing (18-24: 86%, 25-34: 62%, 35-44: 23%; p < 0.0001). Approximately half (55.8%) of the respondents reported having been offered increased screening for possible early detection of ovarian cancer, of which 81.0% chose to undergo screening. The majority utilized a combination of transvaginal ultrasound and serum CA125 measurements. There were no differences observed in screening utilization based on BRCA mutation type. Conclusion: In our cohort of female BRCA mutation carriers, risk-reducing surgery was offered to almost all women, whereas only half were offered risk-reducing medication and/or increased screening. Further investigation is needed to identify barriers to the utilization of risk-reducing strategies among this high-risk population.

Abstract A048: Utero-tubal lavage proteomic analysis: detection of ovarian cancer in BRCA mutation carriers

Abstract A048: Utero-tubal lavage proteomic analysis: detection of ovarian cancer in BRCA mutation carriers

Abstract A029: Development of a new iPSC-derived organoid platform to model FTE-derived HGSOC onset and progression

Abstract A029: Development of a new iPSC-derived organoid platform to model FTE-derived HGSOC onset and progression

Abstract A058: Mesenchymal stem cells act as a selective pressure for High Grade Serous Ovarian Cancer initiation

Abstract High grade serous ovarian cancer (HGSOC) is the most common and lethal subtype of ovarian cancer accounting for 80-90% of diagnoses. Despite its high incidence and mortality, the mechanisms of cancer initiation are unclear. The majority of HGSOC originates in the fallopian tube epithelium (FTE) with support of underlying mesenchymal stromal/stem cells (MSCs). MSCs are stromal progenitor cells that play important roles in wound healing and, in the context of cancer, tumor progression. We showed that MSCs within the tumor microenvironment, termed cancer associated MSCs (CA-MSCs) harbor a unique, age-associated epigenetic signature that is essential for tumor growth and metastasis. Surprisingly, we identified rare subsets of MSCs from benign tubes that exhibit similar epigenetic changes to cancer educated CA-MSCs suggesting that the pro-oncogenic MSC phenotype exists prior to cancer initiation. These “high risk” MSCs (hrMSCs) increase with age, are enriched in BRCA mutation carrier patients, promote cancer cell proliferation, and most notably, elicit tumor formation in primary, non-cancerous FTE. To elucidate the mechanism of hrMSC-induced malignant transformation of FTE, we characterized the effect that hrMSCs have on primary FTE. HrMSCs promoted increased growth of FTE relative to normal/low risk MSCs (lrMSCs) derived from the same patient. These MSCs lack the epigenetic signature/transcriptome of hrMSCs/CA-MSCs. Intriguingly, we co-cultured FTE with hrMSCs at a 1:1 ratio and discovered robust DNA damage in the form of DNA double strand breaks (53BP1 foci) in FTE at 24 hours. Furthermore, conditioned media from hrMSCs supported the recovery of FTE following damage with other DNA damaging agents. We hypothesize that the DNA damage function of hrMSCs serves as a selective pressure for mutation in FTE (1st hit). Additionally, we hypothesize that hrMSCs simultaneously provide growth signals to FTE allowing for evasion of DNA damage-induced cell death (2nd hit). Lastly, we hypothesize that both hrMSC functions are required for genesis of HGSOC. Citation Format: Geyon L. Garcia, Huda I. Atiya, Lan G. Coffman. Mesenchymal stem cells act as a selective pressure for High Grade Serous Ovarian Cancer initiation [abstract]. In: Proceedings of the AACR Special Conference on Ovarian Cancer; 2023 Oct 5-7; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(5 Suppl_2):Abstract nr A058.

Risk Factors for Ovarian Cancer by BRCA Status: A Collaborative Case-Only Analysis.

Women with an inherited pathogenic variant in BRCA1 or BRCA2 have a greatly increased risk of developing ovarian cancer, but the importance of behavioral factors is less clear. We used a case-only design to compare the magnitude of associations with established reproductive, hormonal, and lifestyle risk factors between BRCA mutation carriers and noncarriers. We pooled data from five studies from the Ovarian Cancer Association Consortium including 637 BRCA carriers and 4,289 noncarriers. Covariate-adjusted generalized linear mixed models were used to estimate interaction risk ratios (IRR) and 95% confidence intervals (CI), with BRCA (carrier vs. noncarrier) as the response variable. IRRs were above 1.0 for known protective factors including ever being pregnant (IRR = 1.29, 95% CI; 1.00-1.67) and ever using the oral contraceptive pill (1.30, 95% CI; 1.07-1.60), suggesting the protective effects of these factors may be reduced in carriers compared with noncarriers. Conversely, the IRRs for risk factors including endometriosis and menopausal hormone therapy were below 1.0, suggesting weaker positive associations among BRCA carriers. In contrast, associations with lifestyle factors including smoking, physical inactivity, body mass index, and aspirin use did not appear to differ by BRCA status. Our results suggest that associations with hormonal and reproductive factors are generally weaker for those with a pathogenic BRCA variant than those without, while associations with modifiable lifestyle factors are similar for carriers and noncarriers. Advice to maintain a healthy weight, be physically active, and refrain from smoking will therefore benefit BRCA carriers as well as noncarriers.

Abstract A050: CD8+ T cell exhaustion and their DNA damage in BRCA1 mutated TNBC

Abstract PARP Poly (ADP-ribose) polymerase inhibitors (PARPi) are treatment options in TNBC patients with inherited BRCA1 mutations. Recent studies have shown that immune checkpoint therapies (ICT) in combination with PARPi are effective in other solid cancer types such as melanoma, pancreatic and BRCA deficient ovarian cancer. In early and advanced BRCA1 TNBC, ICT studies are limited and not fully elucidated especially for new ICT that target LAG-3, CTLA-4, ICOS. There is a need for identifying potential prognostic markers suggesting which patients could benefit most from ICT. Therefore, we conducted a detailed single cell analysis of the tumor microenvironment (TME) mainly focused on immune checkpoint expression, CD8 T cell exhaustion and DNA damage associated with BRCA1 TNBC compared to WT TNBC. Combining single cell spatial tissue multiplexing (PhenoCycler) (BRCA1 TNBC n=77, WT TNBC n=55), scRNA seq (BRCA2 n=1, WT n=5), FLOW (PBMCs: BRCA1 n=13, BRCA2 n=17, WT n=62) as well as bulk RNA seq (BRCA1 TNBC n=35, WT TNBC n=11), we performed a detailed analysis of the TME associated with BRCA1 and WT breast cancer. We developed a 32-plex antibody panel that evaluated T-cells and other immune cells with markers of exhaustion, DNA damage and immune checkpoint expression. CD8+ T cell frequency was significantly increased in BRCA1 TNBC compared to WT TNBC (p=0.0015). A detailed CD8 T cell characterization revealed increased dysfunctional/exhausted cells (e.g., TOX+/LAG-3+/PD-1+), T cell DNA damage (CD3e+/CD8+/RAD51+, p=0.0047) and LAG-3 transcript levels in BRCA1 TNBC compared to WT TNBC (p=0.0625). Furthermore, immune checkpoint protein expression in tumor- (PD-L1) and immune cells (PD-1, LAG-3, ICOS) was increased in the TME associated with BRCA1 TNBC compared to WT TNBC. Of note, we also found that PBMCs of healthy tumor-free BRCA mutation carriers contained significantly increased CD8+/PD+1 T cells compared to WT (p=0.012). Our data indicates that BRCA1 haploinsufficiency leads to DNA damaged and dysfunctional CD8+ T cells within the TME compared to WT. Thus, the TME of BRCA1 TNBC patients is more immune suppressive due to increased exhausted CD8+ T cell frequency, their DNA damage as well as immune checkpoint expression of both tumor-/and immune cells compared to WT TNBC. The effect of BRCA1 haploinsufficiency on immune cells is important to consider when trials of ICT options are designed. Citation Format: Dana Pueschl, Anupma Nayak, Derek A. Oldridge, William Chandler, Bria Fulmer, Sokratis Apostolidis, Danielle Bragen, Bradley Wubbenhorst, John Pluta, kConFab Investigator, Antonis Antoniou, Georgia Chenevix-Trench, E. John Wherry, Susan M. Domchek, Katherine L. Nathanson. CD8+ T cell exhaustion and their DNA damage in BRCA1 mutated TNBC [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Breast Cancer Research; 2023 Oct 19-22; San Diego, California. Philadelphia (PA): AACR; Cancer Res 2024;84(3 Suppl_1):Abstract nr A050.

BRCA Mutation Patients: Are There Other Predisposing Factors for Ovarian Cancer Occurrence? A Multicenter Retrospective Study

Objectives: The objective of this multicenter retrospective study aimed to evaluate the association of clinical variables and the incidence of ovarian cancer in patients with BRCA 1–2 mutation carriers who underwent risk-reducing salpingo-oophorectomy (RRSO). Design: Patients with a pathogenic mutation of BRCA 1–2 genes and with no evidence of disease are considered eligible. The exclusion criterion was the refusal to undergo the surgery. The retrospective study included all RRSO performed from May 2015 to April 2022 in the three gynecological Institutions of Southern Italy for were included in this retrospective study. Participants/Materials, Setting, Methods: Age, menarche age, BMI, menopause at time of RRSO, breast cancer first- and second-degree relatives, ovarian cancer first- and second-degree relatives, estroprogestin use, pregnancy normal full-term delivery, history of endometriosis, previous breast cancer and histologic type, previous abdominal/pelvic surgery, BRCA 1 or BRCA 2 status, preoperative serum CA-125 levels (IU/mL), age at time of RRSO and histological analysis were collected. Results: 184 were recruited. One was excluded. To assess cancer risk, the outcome variable was classified into three classes: no event, cancer, and other conditions excluding cancer. 14 women presented ovarian cancer and tubal intraepithelial carcinoma (STIC) on histopathologic final report. Ovarian cancer was found in 8 patients, whereas the presence of STIC was found in 6 of them. Limitations: The low incidence of patients diagnosed with ovarian cancer or STIC compared with the total number of patients undergoing RRSO is a potential bias. Conclusions: Our study did not demonstrate a correlation between clinical features and the occurrence of precancerous or cancerous lesions in BRCA mutation carrier patients.

The Concept behind the Suitability of Menstrual Blood-Derived Stem Cells for the Management of Vaginal Atrophy among BRCA Mutation Carriers after RRSO.

Risk-reducing bilateral salpingo-oophorectomy (RRSO) is recommended for breast cancer gene 1 (BRCA1) and 2 (BRCA2) mutation carriers. A major consequence of RRSO is surgical menopause associated with severe menopausal symptoms, mostly genitourinary complaints. Due to the inherent breast cancer risk, estrogen-based therapies are generally avoided in these patients. So far, the non-hormonal approaches available are not efficient to successfully treat the disabling vaginal atrophy-related symptoms. In regenerative medicine, mesenchymal stem cells (MSC) are the most frequently used cell type due to their remarkable and regenerative characteristics. Therapies based on MSC have revealed positive outcomes regarding symptoms and signs associated with vaginal atrophy by promoting angiogenesis, vaginal restoration, and the proliferation of vaginal mucosa cells. Menstrual blood-derived stem cells (MenSC) are a novel source of MSC, with promising therapeutic potential directly linked to their high proliferative rates; low immunogenicity; non-invasive, easy, and periodic acquisition; and almost no associated ethical issues. In this review, we update the current knowledge and research regarding the potential value of previously preserved MenSC in the therapy of vaginal atrophy among BRCA mutation carriers subjected to RRSO.

A cost-effectiveness analysis of an integrated clinical-radiogenomic screening program for the identification of BRCA 1/2 carriers (e-PROBE study)

Current approach to identify BRCA 1/2 carriers in the general population is ineffective as most of the carriers remain undiagnosed. Radiomics is an emerging tool for large scale quantitative analysis of features from standard diagnostic imaging and has been applied also to identify gene mutational status. The objective of this study was to evaluate the clinical and economic impact of integrating a radiogenomics model with clinical and family history data in identifying BRCA mutation carriers in the general population. This cost-effective analysis compares three different approaches to women selection for BRCA testing: established clinical criteria/family history (model 1); established clinical criteria/family history and the currently available radiogenomic model (49% sensitivity and 87% specificity) based on ultrasound images (model 2); same approach used in model 2 but simulating an improvement of the performances of the radiogenomic model (80% sensitivity and 95% specificity) (model 3). All models were trained with literature data. Direct costs were calculated according to the rates currently used in Italy. The analysis was performed simulating different scenarios on the generation of 18-year-old girls in Italy (274,000 people). The main outcome was to identify the most effective model comparing the number of years of BRCA-cancer healthy life expectancy (HLYs). An incremental cost-effectiveness ratio (ICER) was also derived to determine the cost in order to increase BRCA carriers-healthy life span by 1 year. Compared to model 1, model 2 increases the detection rate of BRCA carriers by 41.8%, reduces the rate of BRCA-related cancers by 23.7%, generating over a 62-year observation period a cost increase by 2.51 €/Year/Person. Moreover, model 3 further increases BRCA carriers detection (+ 68.3%) and decrease in BRCA-related cancers (− 38.4%) is observed compared to model 1. Model 3 increases costs by 0.7 €/Year/Person. After one generation, the estimated ICER in the general population amounts to about 3800€ and 653€ in model 2 and model 3 respectively. Model 2 has a massive effect after only one generation in detecting carriers in the general population with only a small cost increment. The clinical impact is limited mainly due to the current low acceptance rate of risk-reducing surgeries. Further multicentric studies are required before implementing the integrated clinical-radiogenomic model in clinical practice.

Signaling Pathway Alterations Driven by BRCA1 and BRCA2 Germline Mutations are Sufficient to Initiate Breast Tumorigenesis by the PIK3CAH1047R Oncogene.

This study provides a single-cell atlas of breast tissues of BRCA1/2 mutation carriers and demonstrates that aberrant signaling due to BRCA1/2 mutations is sufficient to initiate breast cancer by mutant PIK3CA.

Systemic and Local Strategies for Primary Prevention of Breast Cancer.

One in eight women will develop breast cancer in the US. For women with moderate (15-20%) to average (12.5%) risk of breast cancer, there are few options available for risk reduction. For high-risk (>20%) women, such as BRCA mutation carriers, primary prevention strategies are limited to evidence-based surgical removal of breasts and/or ovaries and anti-estrogen treatment. Despite their effectiveness in risk reduction, not many high-risk individuals opt for surgical or hormonal interventions due to severe side effects and potentially life-changing outcomes as key deterrents. Thus, better communication about the benefits of existing strategies and the development of new strategies with minimal side effects are needed to offer women adequate risk-reducing interventions. We extensively review and discuss innovative investigational strategies for primary prevention. Most of these investigational strategies are at the pre-clinical stage, but some are already being evaluated in clinical trials and others are expected to lead to first-in-human clinical trials within 5 years. Likely, these strategies would be initially tested in high-risk individuals but may be applicable to lower-risk women, if shown to decrease risk at a similar rate to existing strategies, but with minimal side effects.

Hormone replacement therapy in gynecological cancer survivors and BRCA mutation carriers: a MITO group survey.

Early iatrogenic menopause in gynecological cancer survivors and BRCA mutation (BRCAm) carriers undergoing risk-reducing salpingo-oophorectomy (RRSO) is a major health concern. Hormone replacement therapy (HRT) is the most effective remedy, but remains underused in clinical practice. The Multicenter Italian Trials in Ovarian cancer and gynecologic malignancies (MITO) group promoted a national survey to investigate the knowledge and attitudes of healthcare professionals regarding the prescription of HRT. The survey consisted of a self-administered, multiple-choice 45-item questionnaire, available online to all MITO members for 2 months starting from January 2022. A total of 61 participants completed the questionnaire (47 out of 180 MITO centers; compliance: 26.1%). Most respondents were female (73.8%), younger than 50 years (65.6%), and gynecologic oncologists (55.7%), working in public general hospitals (49.2%). An 84.4% of specialists actively discuss HRT with patients and 51.0% of patients ask the specialist for an opinion on HRT. The rate of specialists globally in favor of prescribing HRT was 22.9% for ovarian cancer, 49.1% for cervical cancer, and 8.2% for endometrial cancer patients. Most respondents (70.5%) believe HRT is safe for BRCA-mutated patients after RRSO. Nearly 70% of physicians prescribe systemic HRT, while 23.8% prefer local HRT. Most specialists recommend HRT for as long as there is a benefit and generally for up to 5 years. Real-world data suggest that many healthcare professionals still do not easily prescribe HRT for gynecological cancer survivors and BRCA mutation carriers after RRSO. Further efforts are required to implement the use of HRT in clinical practice and to support both clinicians in recommending HRT and patients in accepting it.