Breast Cancer Susceptibility Gene Research Articles

Abstract PO2-08-09: Prevalence of germline genetic variants in breast cancer susceptibility genes among patients with metaplastic breast cancer

Abstract Background: Pathogenic or likely pathogenic variants (PV) in breast cancer (BC) susceptibility genes are present in approximately 5-10% of individuals with breast cancer and approximately 13-15% of those with triple-negative breast cancer (TNBC). Identifying patients with PV in BRCA1 and BRCA2 can inform selection of appropriate candidates for poly(ADP-ribose) polymerase inhibitors (PARPi), which have improved progression-free and overall survival in the metastatic and adjuvant settings, respectively. Metaplastic breast cancer (MpBC) is a rare, aggressive subtype of breast cancer that is often triple-negative and refractory to chemotherapy. To date, limited data describe germline genetic findings in patients with MpBC. Methods: We conducted a retrospective chart review including patients with pathology results at the Hospital of the University of Pennsylvania from 2021 through May 2023 demonstrating MpBC. Germline genetic test results and tumor histopathologic characteristics including estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2) status were evaluated. Descriptive statistics as well as two-tailed t tests were used for data analysis. Results: Of 89 patients identified with MpBC, 51 had germline genetic testing results available for review. Nine patients (17.6%) had MpBC with PV in BC predisposition genes (PV-MpBC) and 42 (82.4%) had PV-negative MpBC (PVneg-MpBC). Seven patients (13.7%) had a PV in BRCA1, 1 (2.0%) in BRCA2, and 1 (2.0%) in CHEK2. Median age at diagnosis was 45 years for patients with PV-MpBC and 56.5 years for those with PVneg-MpBC (p=0.005). Among patients with PV-MpBC, 6 (66.7%) had ER and PR negative disease by ASCO-CAP guidelines and 3 (33.3%) had ER and/or PR positive disease. Among patients with PVneg-MpBC, 31 (73.8%) had ER and PR negative disease and 11 (26.2%) had ER and/or PR positive disease. All 9 patients with PV-MpBC had HER2-negative disease, one (11.1%) of whom had HER2-low disease. Among patients with PVneg-MpBC, 40 (95.2%) had HER2-negative disease, two (4.8%) of whom had HER2-low disease, and 2 (4.8%) had HER2-positive BC. Conclusions: The frequency of PV in breast cancer susceptibility genes among patients with MpBC is comparable to, and possibly higher than, the reported rates of such PV in patients with TNBC, for whom genetic testing is recommended. If confirmed in a larger dataset, germline testing should be considered for all patients with MpBC regardless of ER, PR, and HER2 status, as findings may help identify candidates for PARPi therapy and genetically-directed clinical trials. Citation Format: Kaitlin Demarest, Anupma Nayak, Payal Shah. Prevalence of germline genetic variants in breast cancer susceptibility genes among patients with metaplastic breast cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO2-08-09.

Abstract PO1-20-07: Valosin-containing protein (VCP)/p97: A novel breast cancer susceptibility gene?

Abstract Introduction: Mutations in the valosin-containing protein (VCP, also known as p97) gene are associated with autosomal dominant inclusion-body myopathy with early-onset Paget disease and frontotemporal dementia. VCP is an ATPase that subserves a variety of cellular functions, prominently, organelle biogenesis and ubiquitin-dependent protein degradation. Recently, VCP has also been shown to physically interact with BRCA1 and is postulated to orchestrate the response to DNA double-strand breaks, a critical molecular prelude to homologous recombination DNA repair. Moreover, VCP is also involved in the removal of trapped poly [ADP-ribose] polymerase 1 (PARP1) from chromatin and may play a key role in the response of tumor cells to PARP inhibitors. Whether VCP acts as a breast cancer predisposition gene is currently unknown. These observations raise speculation as to whether pathologic variation in VCP may predispose to breast cancer. Case Description: A 45-year-old female presented with early-age, locally advanced invasive ductal carcinoma, grade 2, ER < 1%, PR 10%, HER2 IHC 3+, FISH ratio 5.4. The patient completed 6 cycles of neoadjuvant PTH and was surgically downstaged to ypT1bN1a at the time of her left mastectomy. She declined adjuvant radiation. She received 1 dose of trastuzumab emtansine which was discontinued due to intolerance and transitioned to complete 1 year of adjuvant trastuzumab and pertuzumab. Her medical history is significant for inclusion body myopathy with Paget disease of bone and/or frontotemporal dementia, which was diagnosed at age 36 after presenting with proximal muscle weakness. Bone scan was consistent with Paget’s disease and neurocognitive testing did not identify any abnormalities. Diagnosis was confirmed via whole genome germline sequencing which identified a pathogenic variant in VCP [c.463C >G (p.Arg155Gly)]. Germline resequencing of 155 known cancer predisposition genes did not identify any pathogenic variants. Whole exome genomic analysis of the patient’s breast cancer revealed an elevated homologous recombination score ( > 42), consistent with molecularly important role of VCP in homologous recombination breast cancer DNA repair. The patient’s family history is significant for muscular dystrophy in multiple family members on her paternal side, each of whom was deceased prior to age 60. Her paternal half-sister has both inclusion body myopathy and breast cancer, and her paternal cousin was diagnosed with breast cancer at age 46 (unknown genetic testing for each). No history of cancer on her maternal side. Discussion: This case raises the intriguing question of whether the VCP gene represents a heretofore unrecognized breast cancer predisposition gene. Further investigation into the role of VCP in tumorigenesis is warranted, and additional population studies are necessary to confirm VCP’s role in breast cancer predisposition. Given that cancers with homologous recombination deficiencies benefit from PARP inhibitors and VCP is involved in the function of the PARP1 protein, patients with mutations in the VCP gene may benefit from PARP inhibition. Citation Format: Alexis LeVee, Duveen Sturgeon, Joanne Mortimer, Kevin McDonnell. Valosin-containing protein (VCP)/p97: A novel breast cancer susceptibility gene? [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO1-20-07.

Abstract PO4-08-03: Prognostic Effect of Ductal Carcinoma in Situ in Breast Cancer with BRCA1/2 Mutations

Abstract Abstract Background: Although Breast Cancer Susceptibility Gene (BRCA)-associated invasive breast cancer has been extensively studied, there are few reports on ductal carcinoma in situ (DCIS) in patients with BRCA1/2 mutations. This study aims to evaluate the prognostic effect of DCIS in breast cancer patients with pathologic variants of BRCA1/2 genes. Methods: Retrospective analysis was performed on a prospective cohort of 157 patients who were found positive for BRCA1/2 mutations through genetic testing from August 2003 to January 2022 at Seoul National University Bundang Hospital. Survival outcomes were compared between patients who had both invasive ductal carcinoma (IDC) and DCIS components (IDC-DCIS group, n = 121) and patients who had IDC only (IDC group, n = 36). Cox regression analysis was performed for evaluation of predictive factors for recurrence. Results: Among the 157 patients, 65 (41.4%) patients showed BRCA1 mutations, 90 (57.3%) patients had pathological variants of BRCA2, and 2 (1.3%) patients tested positive for both BRCA1/2 mutations. There was no difference in baseline clinical characteristics between the IDC-DCIS and IDC groups. When pathological features were compared, the IDC-DCIS group was more likely to express positivity in both estrogen receptor (IDC 5 (13.9%) vs. IDC-DCIS 74 (61.2%), P < 0.001) and progesterone receptor status (IDC 5 (13.9%) vs. IDC-DCIS 67 (55.4%), P < 0.001). Tumors in the IDC group were associated with a higher histologic grade (P = 0.004). There was no statistically significant difference in 5-year disease-free survival between the two groups (IDC 83.6% vs. IDC-DCIS 89.3%, P = 0.989). In univariate Cox regression analysis, younger age at diagnosis was a significant predictor for recurrence (hazard ratio (HR) 2.236, P = 0.049); salpingo-oophorectomy showed a risk-reducing effect (HR 0.106, P < 0.001). Further multivariate analysis found only the effect of salpingo-oophorectomy to be prognostic (HR 0.112, P < 0.001). Presence of DCIS was not a risk factor for recurrence in patients with BRCA1/2 mutations (HR 1.006, P = 0.989). Conclusion: BRCA1/2-positive breast cancer with DCIS components is more likely to be hormone receptor-positive and of lower grade compared to patients with IDC only. A tailored approach might be necessary in establishing treatment options for breast cancer patients with BRCA1/2 mutations according to the presence of DCIS. Citation Format: Kyung-Hwak Yoon, Eun-Kyu Kim, Hee-Chul Shin. Prognostic Effect of Ductal Carcinoma in Situ in Breast Cancer with BRCA1/2 Mutations [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO4-08-03.

Abstract PO5-09-04: Oncotype DX Breast Recurrence Score® analysis in early-stage breast cancer associated with CHEK2, ATM and PALB2 germline pathogenic variants

Abstract Background: The 21-gene assay (Oncotype DX Recurrence Score, RS) is the only clinically validated assay to determine the prognosis in hormone-positive, HER2-negative breast cancer (BC) patients treated with endocrine therapy alone by predicting locoregional and distant recurrence. Outcomes of Oncotype RS in BRCA-associated BC has been studied previously, but the results of RS analysis in BC patients with the other BC susceptibility genes have not been documented. This study evaluates the clinical and pathological characteristics of BC patients with ATM, CHEK2 and PALB2 germline pathogenic variants (PV). Methods: Patients with invasive hormone-positive, HER2-negative BC who have had Oncotype DX Breast Recurrence Score® testing and who underwent genetic testing in the Breast Medical Oncology and the Breast Clinical Cancer Genetics Clinic at the University of Texas MD Anderson Cancer Center (MDACC) were identified from the prospectively maintained research database. Clinical and tumor characteristics were analyzed using descriptive statistics. Results: Between 1996 and June 2023, 5652 patients with Oncotype RS were included in this analysis. Amongst those 828 BC patients had genetic testing and 81 patients were found to have a pathogenic variant: 17 patients with a CHEK2 mutation, 10 patients with an ATM mutation, and 18 patients with a PALB2 mutation. Among patients with CHEK2 PV, median Oncotype RS was 17 (5–32) overall, 21 (13–32) for patients age < 50, and 14.5 (5–28) for patients age ≥50 (p= 0.2). Among patients with ATM PV, median RS was 16.5 (3–54) overall, 25 (10–54) for patients age < 50, and 14.5 (3–29) for patients age ≥50 (p= 0.19). Among patients with PALB2 PV, median RS was 26.5 (15–40) overall, 20 (15–40) for patients age < 50, and 31 (26–37) for patients age ≥50 (p= 0.012). Further patient and tumor characteristics are shown in Table 1. Conclusion: This is a single-institutional cohort of hormone-positive, HER2-negative BC patients who had Oncotype RS and who underwent genetic testing. Further analysis on clinical and tumor pathological characteristics and long-term outcome is ongoing. Table1. Patient and tumor characteristics Citation Format: Fatma Akkoc Mustafayev, Angelica Gutierrez Barrera, Diane Liu, Banu Arun. Oncotype DX Breast Recurrence Score® analysis in early-stage breast cancer associated with CHEK2, ATM and PALB2 germline pathogenic variants [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO5-09-04.

Using genome and transcriptome data from African-ancestry female participants to identify putative breast cancer susceptibility genes

African-ancestry (AA) participants are underrepresented in genetics research. Here, we conducted a transcriptome-wide association study (TWAS) in AA female participants to identify putative breast cancer susceptibility genes. We built genetic models to predict levels of gene expression, exon junction, and 3′ UTR alternative polyadenylation using genomic and transcriptomic data generated in normal breast tissues from 150 AA participants and then used these models to perform association analyses using genomic data from 18,034 cases and 22,104 controls. At Bonferroni-corrected P < 0.05, we identified six genes associated with breast cancer risk, including four genes not previously reported (CTD-3080P12.3, EN1, LINC01956 and NUP210L). Most of these genes showed a stronger association with risk of estrogen-receptor (ER) negative or triple-negative than ER-positive breast cancer. We also replicated the associations with 29 genes reported in previous TWAS at P < 0.05 (one-sided), providing further support for an association of these genes with breast cancer risk. Our study sheds new light on the genetic basis of breast cancer and highlights the value of conducting research in AA populations.

A multi-tissue, splicing-based joint transcriptome-wide association study identifies susceptibility genes for breast cancer.

Splicing-based transcriptome-wide association studies (splicing-TWASs) of breast cancer have the potential to identify susceptibility genes. However, existing splicing-TWASs test the association of individual excised introns in breast tissue only and thus have limited power to detect susceptibility genes. In this study, we performed a multi-tissue joint splicing-TWAS that integrated splicing-TWAS signals of multiple excised introns in each gene across 11 tissues that are potentially relevant to breast cancer risk. We utilized summary statistics from a meta-analysis that combined genome-wide association study (GWAS) results of 424,650 women of European ancestry. Splicing-level prediction models were trained in GTEx (v.8) data. We identified 240 genes by the multi-tissue joint splicing-TWAS at the Bonferroni-corrected significance level; in the tissue-specific splicing-TWAS that combined TWAS signals of excised introns in genes in breast tissue only, we identified nine additional significant genes. Of these 249 genes, 88 genes in 62 loci have not been reported by previous TWASs, and 17 genes in seven loci are at least 1 Mb away from published GWAS index variants. By comparing the results of our splicing-TWASs with previous gene-expression-based TWASs that used the same summary statistics and expression prediction models trained in the same reference panel, we found that 110 genes in 70 loci that are identified only by the splicing-TWASs. Our results showed that for many genes, expression quantitative trait loci (eQTL) did not show a significant impact on breast cancer risk, whereas splicing quantitative trait loci (sQTL) showed a strong impact through intron excision events.

Olaparib and niraparib as maintenance therapy in patients with newly diagnosed and platinum-sensitive recurrent ovarian cancer: A single-center study in China.

Poly adenosine-diphosphate-ribose polymerase (PARP) inhibitors (PARPi) have been approved to act as first-line maintenance (FL-M) therapy and as platinum-sensitive recurrent maintenance (PSR-M) therapy for ovarian cancer in China for >5 years. Herein, we have analyzed the clinical-application characteristics of olaparib and niraparib in ovarian cancer-maintenance therapy in a real-world setting to strengthen our understanding and promote their rational usage. A retrospective chart review identified patients with newly diagnosed or platinum-sensitive recurrent ovarian cancer, who received olaparib or niraparib as maintenance therapy at Sichuan Cancer Hospital between August 1, 2018, and December 31, 2021. Patient medical records were reviewed. We grouped and analyzed patients based on the type of PARPi they used (the olaparib group and the niraparib group) and the line of PARPi maintenance therapy (the FL-M setting and the PSR-M setting). The primary endpoint was the 24-month progression-free survival (PFS) rate. In total, 131 patients (olaparib: n=67, 51.1%; niraparib: n=64, 48.9%) were enrolled. Breast cancer susceptibility genes (BRCA) mutations (BRCAm) were significantly less common in the niraparib group than in the olaparib group [9.4% (6/64) vs. 62.7% (42/67), P<0.001], especially in the FL-M setting [10.4% (5/48) vs. 91.4% (32/35), P<0.001]. The 24-month PFS rates in the FL-M and PSR-M settings were 60.4% and 45.7%, respectively. In patients with BRCAm, the 24-month PFS rates in the FL-M and PSR-M settings were 62.2% and 72.7%, respectively. Olaparib and niraparib were effective in patients with ovarian cancer without any new safety signals except for skin pigmentation. In patients with BRCAm, the 24-month PFS of the PARPi used in the PSR-M setting was even higher than that used in the FL-M setting.

A study on patients’ selection for BRCA1 and BRCA2 mutations testing by different models in Libyan women with breast cancer

ABSTRACT Introduction The BRCA mutation spectrum of familial breast cancer in Libya remains unknown. Several genetic models developed to predict the probability of BRCA1/2 mutations have not been applied in Libya, where the NCCN criteria are used for highly penetrating breast cancer susceptibility genes. We aimed to predict BRCA1/2 mutation probability in familial breast cancer and genetic testing eligibility using BOADICEA and BRCAPRO models and NCCN criteria. Methods BRCA1/2 mutations were retrospectively predicted in 62 women with familial breast cancer between 2018 and 2021. Logistic regression, ROC analysis, and area under the curve were used to compare NCCN referral criteria with the BRCAPRO and BOADICEA scores. Results Thirty-two out of 62 breast cancer patients (51.6%), with a mean age of 43.5 ± 8 years, were predicted as BRCA mutation carriers by both models. BRCAPRO predicted BRCA1 and BRCA2 mutations in 27.4% and 41.9% of the women, respectively. BOADICEA predicted 8% for BRCA1 and 29% for BRCA2. At least one NCCN criterion was met by 50/62 women (80.6%). Three criteria were statistically significant predictors in BRCAPRO and BOADICEA: breast cancer at age ≤50 years with one or more close blood relatives with breast cancer, breast cancer patient with a close relative of male breast cancer, and triple-negative breast cancer. For the three respective criteria, sensitivity was 0.78, 0.89, and 0.75, specificity was 0.33, 0.39, and 0.22, area under curve was 0.72, 0.75, and 0.76, positive predictive value was 55%, 61%, and 51%, and negative predictive value was 58.5%, 77%, and 45%. Conclusions Our study highlights that certain aspects of the NCCN criteria demonstrate variations in significance when compared to the BRCAPR and BOADICEA models. For the first time, these models were used to predict BRCA mutations in Libyan women, and our finding indicates that these models are promising for improving genetic testing decision-making.

Radiosensitizing effect of a novel CTSS inhibitor by enhancing BRCA1 protein stability in triple-negative breast cancer cells.

Triple-negative breast cancer (TNBC) patients harboring wild-type breast cancer susceptibility gene 1 (BRCA1) account for most TNBC patients but lack adequate targeted therapeutic options. Although radiotherapy (RT) is the primary treatment modality for TNBC patients, radioresistance is one of the major challenges. RT-induced increase in cathepsin S (CTSS) causes radioresistance through suppressing BRCA1-mediated apoptosis of tumor cells, which was induced by CTSS-mediated degradation of BRCA1. Targeting CTSS may provide a novel therapeutic opportunity for TNBC patients. Publicly available data and human tissue microarray slides were analyzed to investigate the relationship between CTSS and BRCA1 in breast cancer patients. A CTSS enzyme assay and in silico docking analysis were conducted to identify a novel CTSS inhibitor. RO5461111 was used first to confirm the concept of targeting CTSS for radiosensitizing effects. The MDA-MB-231 TNBC cell line was used for invitro and invivo assays. Western blotting, promoter assay, cell death assay, clonogenic survival assay, and immunohistochemistry staining were conducted to evaluate novel CTSS inhibitors. CTSS inhibitors were further evaluated for their additional benefit of inhibiting cell migration. A novel CTSS inhibitor, TS-24, increased BRCA1 protein levels and showed radiosensitization in TNBC cells with wild-type BRCA1 and invivo in a TNBC xenograft mouse model. These effects were attributed by BRCA1-mediated apoptosis facilitated by TS-24. Furthermore, TS-24 demonstrated the additional effect of inhibiting cell migration. Our study suggests that employing CTSS inhibitors for the functional restoration of BRCA1 to enhance RT-induced apoptosis may provide a novel therapeutic opportunity for TNBC patients harboring wild-type BRCA1.

BRCA1-mediated dual regulation of ferroptosis exposes a vulnerability to GPX4 and PARP co-inhibition in BRCA1-deficient cancers.

Resistance to poly (ADP-ribose) polymerase inhibitors (PARPi) limits the therapeutic efficacy of PARP inhibition in treating breast cancer susceptibility gene 1 (BRCA1)-deficient cancers. Here we reveal that BRCA1 has a dual role in regulating ferroptosis. BRCA1 promotes the transcription of voltage-dependent anion channel 3 (VDAC3) and glutathione peroxidase 4 (GPX4); consequently, BRCA1 deficiency promotes cellular resistance to erastin-induced ferroptosis but sensitizes cancer cells to ferroptosis induced by GPX4 inhibitors (GPX4i). In addition, nuclear receptor coactivator 4 (NCOA4)-mediated ferritinophagy and defective GPX4 induction unleash potent ferroptosis in BRCA1-deficient cancer cells upon PARPi and GPX4i co-treatment. Finally, we show that xenograft tumors derived from BRCA1-mutant breast cancer patients with PARPi resistance exhibit decreased GPX4 expression and high sensitivity to PARP and GPX4 co-inhibition. Our results show that BRCA1 deficiency induces a ferroptosis vulnerability to PARP and GPX4 co-inhibition and inform a therapeutic strategy for overcoming PARPi resistance in BRCA1-deficient cancers.

A Novel Inhibitor of Poly(ADP-Ribose) Polymerase-1 Inhibits Proliferation of a BRCA-Deficient Breast Cancer Cell Line via the DNA Damage-Activated cGAS-STING Pathway.

Loss-of-function mutations in the Breast Cancer Susceptibility Gene (BRCA1 and BRCA2) are often detected in patients with breast cancer. Poly(ADP-ribose) polymerase-1 (PARP1) plays a key role in the repair of DNA strand breaks, and PARP inhibitors have been shown to induce highly selective killing of BRCA1/2-deficient tumor cells, a mechanism termed synthetic lethality. In our previous study, a novel PARP1 inhibitor─(E)-2-(2,3-dibromo-4,5-dimethoxybenzylidene)-N-(4-fluorophenyl) hydrazine-1-carbothioamide (4F-DDC)─was synthesized, which significantly inhibited PARP1 activity with an IC50 value of 82 ± 9 nM. The current study aimed to explore the mechanism(s) underlying the antitumor activity of 4F-DDC under in vivo and in vitro conditions. 4F-DDC was found to selectively inhibit the proliferation of BRCA mutant cells, with highly potent effects on HCC-1937 (BRCA1-/-) cells. Furthermore, 4F-DDC was found to induce apoptosis and G2/M cell cycle arrest in HCC-1937 cells. Interestingly, immunofluorescence and Western blot results showed that 4F-DDC induced DNA double strand breaks and further activated the cGAS-STING pathway in HCC-1937 cells. In vivo analysis results revealed that 4F-DDC inhibited the growth of HCC-1937-derived tumor xenografts, possibly via the induction of DNA damage and activation of the cGAS-STING pathway. In summary, the current study provides a new perspective on the antitumor mechanism of PARP inhibitors and showcases the therapeutic potential of 4F-DDC in the treatment of breast cancer.

Abstract 2998: Regulation of epidermal growth factor receptor signaling by BRCA1-microRNA 146a network

Abstract Inherited defects in the breast cancer susceptibility gene, BRCA1 are associated with an increased risk for hereditary breast, ovarian, and other cancers. BRCA1-associated basal like breast cancers (BLBCs) are high-grade ductal carcinomas that do not express ER, PR or HER2, but frequently overexpress epidermal growth factor receptor (EGFR/ErbB1/HER1). The EGFR-driven signaling network plays a pivotal role in BLBC pathogenesis. EGFR overexpression is associated with tumor progression and metastasis, resistance to radiation and chemotherapy, and poor prognosis. Growth factor receptors are regulated by multiple miRNAs, each having many targets, coordinately controlling the complex pathways of downstream signaling. We have previously reported a post-transcriptional mechanism by which BRCA1 regulates EGFR expression through the induction of miR-146a and provided a rationale for the development of miR-146a based therapeutic strategies. In addition, we have shown that BRCA1 also exerts regulatory control over dozens of important miRNAs that appear to play a critical role in breast neoplasia. From TCGA analysis, we show that low expression of these miRNAs is associated with distinctively poor overall survival of TNBC patients. Using miR-146a loss or gain of function experiments in vitro, we show that BRCA1 and miR-146a regulate EGFR signaling, stemness, epithelial-to-mesenchymal transition (EMT), extracellular matrix (ECM) remodeling and chemoresistance. Using in vivo mouse models, we demonstrate that miR-146a overexpression delays tumor formation leading to better overall survival. BLBC unlike the other subtypes, lacks targeted therapy approaches making cytotoxic chemotherapy essentially the only option for treatment. Though EGFR inhibitors have been used to treat other cancers, clinical trials for breast cancer have so far been unsuccessful due to poor response rates. Since one miRNA can target multiple genes and regulate multiple signaling pathways, our study provides evidence to suggest that restoring miR-146a could suppress EGFR-RAS-MEK signaling and the other compensatory pathways, providing a targeted therapeutic option for BRCA1 associated BLBC. Citation Format: Easwari Kumaraswamy, Raeann M. Koren Shimak, Sumedha Gunewardena, Karen L. Wendt, Dani Alexander, Stacey L. Hembruff, Roy A. Jensen. Regulation of epidermal growth factor receptor signaling by BRCA1-microRNA 146a network [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 2998.

Abstract 5686: Identifying rare microRNA risk variants for hereditary breast cancer in African American individuals

Abstract Background: In the United States, breast cancer is the most frequently diagnosed cancer in women. However, African American women tend to have a higher incidence of triple negative breast cancer cases, and an earlier age of disease onset, which are hallmark characteristics of heredity disease. Even with current breakthroughs in breast cancer research, limited research participation from women of African descent has led to a gap in our knowledge of heredity factors that are involved in disease in this population. Furthermore, most breast cancer research has focused on protein coding genes being the culprit of tumorigenesis; however, within the last three decades, research efforts have begun to focus on noncoding RNAs, like microRNAs (miRNAs), as potential factors to promote carcinogenesis. miRNAs are short, noncoding RNA molecules that control gene expression at the post-translational level. miRNAs have several mechanisms to affect gene expression of cellular processes like cell proliferation, apoptosis, invasion, and metastasis. However, the identification and functional role miRNA risk variants play in African American breast cancer is still not fully understood. Methods: 30X whole genome sequencing (WGS) was carried out on an Illumina platform on blood- or saliva-derived DNA from 120 African American breast cancer cases from the Alabama Hereditary Cancer Cohort. The WGS data was analyzed using our established bioinformatics pipeline, which has been adapted from the Genome Analysis Toolkit (GATK) Best Practice Pipeline. Focusing on miRNA variants as risk factors, a miRNA BED file was used to extract regions harboring miRNA genes. The resulting file was then filtered for rare miRNA variants (minor allele frequency &amp;lt;1%) in the African American population based on the Genome Aggregation Database (gnomAD) allele frequency. P-values for variants of interest were calculated using Fisher’s Exact tests. Results: Sequencing data have revealed three rare miRNA variants from our African American cohort that can be potentially associated with breast cancer risk. Upon statistical analysis comparing allele frequencies between the African American general population and the Alabama Hereditary Cancer Cohort, miR7850, miR3145, and miR8074 had p-values 5.21E-05, 3.58E-05, and 2.19E-04, respectively. Conclusions: We have identified three rare miRNA variants that were not previously known to be associated with breast cancer risk. Interestingly, both miR7850 and miR8074 target TP53 and miR3145 targets GREM1 and PALB2, which these targets are known breast cancer susceptibility genes. Understanding the consequence of these variants is important to add to our knowledge of miRNAs that play critical roles in carcinogenesis leading to better treatment strategies. Citation Format: Sheniqua R. Glover, Troy LoBue, Nancy D. Merner. Identifying rare microRNA risk variants for hereditary breast cancer in African American individuals [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 5686.

Abstract 769: Incorporating a cross-ancestry polygenic risk score into a clinical model improved breast cancer risk prediction in women with pathogenic variants

Abstract Introduction. We previously developed and validated a cross-ancestry polygenic risk score (caPRS) to predict risk of developing breast cancer (BC) in women who do not carry pathogenic variants (PVs) in BC susceptibility genes. Here we aimed to expand upon that study by integrating the caPRS with the Breast and Ovarian Analysis of Disease Incidence and Carrier Estimation Algorithm (BOADICEA) model to combine the effects of genetic and clinical factors in carriers of PVs in BRCA1, BRCA2, PALB2, CHEK2 and ATM. Methods. We explored the association of caPRS among 12,525 women from the UK Biobank and the Consortium of Investigators of Modifiers of BRCA1/2. The effect size of the caPRS was evaluated separately for individuals carrying PVs in BRCA1, BRCA2, CHEK2, ATM and PALB2. We used a logistic regression model adjusted for age, first-degree BC family history (FHx) and cohort for BRCA1 and BRCA2 to examine the association of the caPRS with BC. The effect sizes were expressed as standardized odds ratios (ORs) with 95% confidence intervals (CIs). The estimated absolute risks to age 80 of developing BC were calculated for unaffected women by combining the caPRS-based risk with gene-specific clinical risk estimates from the BOADICEA model based on U.S. incidences. Results. The caPRS was significantly associated with BC risk across all PV carrier groups. Using BOADICEA alone, the estimated absolute BC risk by age 80 for an average unaffected 20-year-old female in the U.S. with an unknown FHx ranged from 23.8% for CHEK2 carriers to 79.4% for BRCA2 carriers. Integration of caPRS into BOADICEA, yielded the distribution of risk for each gene. (Table) Gene N OR per SD (95% CI) Absolute risk by BOADICEA Median absolute risk by BOADICEA + caPRS (Range) ATM 2,037 1.46 (1.25 - 1.71) 24.9% 27.1% (0.1 - 70.5) BRCA1 5,794 1.24 (1.18 - 1.31) 76.1% 75.6% (55.6 - 87.7) BRCA2 4,018 1.40 (1.31 - 1.49) 79.4% 78.7% (45.2 - 92.1) CHEK2 372 1.88 (1.41 - 2.55) 23.8% 28.2% (0.0 - 78.4) PALB2 304 1.81 (1.37 - 2.43) 53.9% 56.3% (2.9 - 90.9) Conclusions. The caPRS significantly modified BC risk for carriers of PVs and could help tailor guidelines for women with PVs, particularly in moderate penetrance genes. More data is needed to increase the precision and reach of risk assessment in diverse populations. Citation Format: Placede Tshiaba, Monika Sun, Dariusz Ratman, Jeffrey N. Weitzel, Premal Shah, Matthew Rabinowitz, Akash Kumar, Kate Im. Incorporating a cross-ancestry polygenic risk score into a clinical model improved breast cancer risk prediction in women with pathogenic variants [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 769.

Inhibition of intracellular ATP synthesis impairs the recruitment of homologous recombination factors after ionizing radiation.

Ionizing radiation (IR)-induced double-strand breaks (DSBs) are primarily repaired by non-homologous end joining or homologous recombination (HR) in human cells. DSB repair requires adenosine-5'-triphosphate (ATP) for protein kinase activities in the multiple steps of DSB repair, such as DNA ligation, chromatin remodeling, and DNA damage signaling via protein kinase and ATPase activities. To investigate whether low ATP culture conditions affect the recruitment of repair proteins at DSB sites, IR-induced foci were examined in the presence of ATP synthesis inhibitors. We found that p53 binding protein 1 foci formation was modestly reduced under low ATP conditions after IR, although phosphorylated histone H2AX and mediator of DNA damage checkpoint 1 foci formation were not impaired. Next, we examined the foci formation of breast cancer susceptibility gene I (BRCA1), replication protein A (RPA) and radiation 51 (RAD51), which are HR factors, in G2 phase cells following IR. Interestingly, BRCA1 and RPA foci in the G2 phase were significantly reduced under low ATP conditions compared to that under normal culture conditions. Notably, RAD51 foci were drastically impaired under low ATP conditions. These results suggest that HR does not effectively progress under low ATP conditions; in particular, ATP shortages impair downstream steps in HR, such as RAD51 loading. Taken together, these results suggest that the maintenance of cellular ATP levels is critical for DNA damage response and HR progression after IR.

The deubiquitinase BRCC3 increases the stability of ZEB1 and promotes the proliferation and metastasis of triple-negative breast cancer cells.

Triple negative breast cancer (TNBC) has a high recurrence rate, metastasis rate and mortality rate. The aim of this study is to identify new targets for the treatment of TNBC. Clinical samples are used for screening deubiquitinating enzymes (DUBs). MDA-MB-231 cells and a TNBC mouse model are used for in vitro and in vivo experiments, respectively. Western blot analysis is used to detect the protein expressions of DUBs, zinc finger E-box binding homeobox 1 (ZEB1), and epithelial-mesenchymal transition (EMT)-related markers. Colony formation and transwell assays are used to detect the proliferation, migration and invasion of TNBC cells. Wound healing assay is used to detect the mobility of TNBC cells. Immunoprecipitation assay is used to detect the interaction between breast cancer susceptibility gene 1/2-containing complex subunit 3 (BRCC3) and ZEB1. ZEB1 ubiquitination levels, protein stability, and protein degradation are also examined. Pathological changes in the lung tissues are detected via HE staining. Our results show a significant positive correlation between the expressions of BRCC3 and ZEB1 in clinical TNBC tissues. Interference with BRCC3 inhibits TNBC cell proliferation, migration, invasion and EMT. BRCC3 interacts with ZEB1 and interferes with BRCC3 to inhibit ZEB1 expression by increasing ZEB1 ubiquitination. Interference with BRCC3 inhibits TNBC cell tumorigenesis and lung metastasis in vivo. In all, this study demonstrates that BRCC3 can increase the stability of ZEB1, upregulate ZEB1 expression, and promote the proliferation, migration, invasion, EMT, and metastasis of TNBC cells, providing a new direction for cancer therapy.

Abstract 1723 Differences in barriers to immortalization in normal breast epithelial cells from donors with and without mutations in breast cancer susceptibility genes

Abstract 1723 Differences in barriers to immortalization in normal breast epithelial cells from donors with and without mutations in breast cancer susceptibility genes

Development and validation of ultrasound-based radiomics model to predict germline BRCA mutations in patients with breast cancer

BackgroundIdentifying breast cancer (BC) patients with germline breast cancer susceptibility gene (gBRCA) mutation is important. The current criteria for germline testing for BC remain controversial. This study aimed to develop a nomogram incorporating ultrasound radiomic features and clinicopathological factors to predict gBRCA mutations in patients with BC.Materials and methodsIn this retrospective study, 497 women with BC who underwent gBRCA genetic testing from March 2013 to May 2022 were included, including 348 for training (84 with and 264 without a gBRCA mutation) and 149 for validation(36 patients with and 113 without a gBRCA mutation). Factors associated with gBRCA mutations were identified to establish a clinicopathological model. Radiomics features were extracted from the intratumoral and peritumoral regions (3 mm and 5 mm) of each image. The least absolute shrinkage and selection operator regression algorithm was used to select the features and logistic regression analysis was used to construct three imaging models. Finally, a nomogram that combined clinicopathological and radiomics features was developed. The models were evaluated based on the area under the receiver operating characteristic curve (AUC), calibration, and clinical usefulness.ResultsAge at diagnosis, family history of BC, personal history of other BRCA-related cancers, and human epidermal growth factor receptor 2 status were independent predictors of the clinicopathological model. The AUC of the imaging radiomics model combining intratumoral and peritumoral 3 mm areas in the validation set was 0.783 (95% confidence interval [CI]: 0.702—0.862), which showed the best performance among three imaging models. The nomogram yielded better performance than the clinicopathological model in validation sets (AUC: 0.824 [0.755—0.894] versus 0.659 [0.563—0.755], p = 0.007).ConclusionThe nomogram based on ultrasound images and clinicopathological factors performs well in predicting gBRCA mutations in BC patients and may help to improve clinical decisions about genetic testing.

Early-stage triple negative breast cancer: the therapeutic role of immunotherapy and the prognostic value of pathological complete response.

Triple negative breast cancer (TNBC) represents an aggressive disease associated with a high risk of recurrence after curative treatment and a poor prognosis in the metastatic setting. Chemotherapy was for years the only treatment available in the early and metastatic setting, due to the lack of actionable targets. Clinical practice has changed following the results obtained with the addition of immunotherapy to standard chemotherapy, the development of novel drugs [i.e. antibody-drug conjugates (ADCs)], and the use of targeted treatments for patients carrying germline pathogenic breast cancer susceptibility genes (BRCA) 1 or BRCA 2 variants. The treatment of early-stage disease has had a shift in clinical practice since July 2021, after the Food and Drug Administration (FDA) approval of pembrolizumab in association with chemotherapy as neoadjuvant treatment for TNBC and as a single agent in the subsequent adjuvant setting. This intensive treatment based on the combination of a poly-chemotherapy and an immune checkpoint inhibitor (ICI) led to the improvement of short- and long-term outcomes, but it has highlighted some new unmet clinical needs in the treatment of early-stage TNBC: the selection of the most effective adjuvant therapy and the integration of pembrolizumab with other therapeutic strategies [capecitabine, poly(ADP-ribose) polymerase (PARP) inhibitors] based on the achievement of pathologic complete response (pCR); the identification of predictive biomarkers to select patients who could most benefit from the addition of ICI, to minimize toxicities and to maximize outcomes; the possibility of de-escalating chemotherapy in favor of immune-combo or novel agents, such as ADCs; the role of immunotherapy in estrogen receptor (ER)-low patients. The advent of immunotherapy not only addresses current challenges in TNBC treatment but also holds the promise of a radical transformation in its therapeutic paradigm, enhancing significantly clinical outcomes and offering new perspectives for patients grappling with this aggressive form of breast cancer.

Germline RECQL gene mutations in Chinese patients with breast cancer.

Breast cancer is the most common malignant tumor in women, seriously threatening health and survival. TP-dependent DNA helicase Q1 (RECQL) is a breast cancer susceptibility gene with possible familial links. However, RECQL gene mutations among Chinese women with breast cancer have not been evaluated. Therefore, this study assessed RECQL mutations and their relationships with clinicopathological and epidemiological characteristics in Chinese women with breast cancer. Clinical information was also obtained via the hospital information system and a follow-up questionnaire. Peripheral venous blood (2 mL) was extracted from all patients and stored at -80°C for future use; the early venous blood samples were from our hospital's sample bank. RECQL gene sequencing were performed by the Shanghai Aishe Gene Company (China). We found that a RECQL mutation is a susceptibility factor for breast cancer. Moreover, patients with RECQL mutations were more likely to have a family history of breast cancer than those without. Also, patients with RECQL variants of uncertain significance (VUS) were less likely to develop invasive ductal carcinoma than those without. In addition, unexplained RECQL mutations occurred more often in patients with human epidermal growth factor receptor 2+ breast cancer than in those with other subtypes. These results provide a basis for creating screening criteria specific to Chinese women. However, the frequency of RECQL mutations was low, and the number of pathogenic mutations was too small and could not be analyzed. Thus, more extensive, long-term studies that include other functional experiments are needed to verify these results.