BRCA-negative Breast Cancer Research Articles

Identification and characterization of ATM founder mutation in BRCA-negative breast cancer patients of Arab ethnicity

Breast cancer (BC) is the most prevalent malignancy among women worldwide with germline pathogenic variants/likely pathogenic variants (PVs/LPVs) in BRCA1/2 accounting for a large portion of hereditary cases. Recently, heterozygous PVs/LPVs in the ATM serine/threonine kinase or Ataxia-telangiectasia mutated gene (ATM) has been identified as a moderate susceptibility factor for BC in diverse ethnicities. However, the prevalence of ATM PVs/LPVs in BC susceptibility in Arab populations remains largely unexplored. This study investigated the prevalence of ATM PVs/LPVs among BC patients from Saudi Arabia, employing capture-sequencing technology for ATM PVs/LPVs screening in a cohort of 715 unselected BC patients without BRCA1/2 PVs/LPVs. In addition, founder mutation analysis was conducted using the PHASE program. In our entire cohort, four unique PVs/LPVs in the ATM gene were identified in six cases (0.8%). Notably, one recurrent LPV, c.6115G > A:p.Glu2039Lys was identified in three cases, for which haplotype analysis confirmed as a novel putative founder mutation traced back to 13 generations on average. This founder mutation accounted for half of all identified mutant cases and 0.4% of total screened cases. This study further reveals a significant correlation between the presence of ATM mutation and family history of BC (p = 0.0127). These findings underscore an approximate 0.8% prevalence of ATM germline PVs/LPVs in Arab BC patients without BRCA1/2 PVs/LPVs and suggest a founder effect of specific recurrent ATM mutation. These insights can help in the design of a genetic testing strategy tailored to the local population in Saudi Arabia, thereby, enabling more accurate clinical management and risk prediction.

Whole-exome sequencing of BRCA-negative breast cancer patients and case–control analyses identify variants associated with breast cancer susceptibility

BackgroundFor the majority of individuals with early-onset or familial breast cancer referred for genetic testing, the genetic basis of their familial breast cancer remains unexplained. To identify novel germline variants associated with breast cancer predisposition, whole-exome sequencing (WES) was performed.MethodsWES on 290 BRCA1/BRCA2-negative Singaporeans with early-onset breast cancer and/or a family history of breast cancer was done. Case–control analysis against the East-Asian subpopulation (EAS) from the Genome Aggregation Database (gnomAD) identified variants enriched in cases, which were further selected by occurrence in cancer gene databases. Variants were further evaluated in repeated case–control analyses using a second case cohort from the database of Genotypes and Phenotypes (dbGaP) comprising 466 early-onset breast cancer patients from the United States, and a Singapore SG10K_Health control cohort.ResultsForty-nine breast cancer-associated germline pathogenic variants in 37 genes were identified in Singapore cases versus gnomAD (EAS). Compared against SG10K_Health controls, 13 of 49 variants remain significantly enriched (False Discovery Rate (FDR)-adjusted p < 0.05). Comparing these 49 variants in dbGaP cases against gnomAD (EAS) and SG10K_Health controls revealed 23 concordant variants that were significantly enriched (FDR-adjusted p < 0.05). Fourteen variants were consistently enriched in breast cancer cases across all comparisons (FDR-adjusted p < 0.05). Seven variants in GPRIN2, NRG1, MYO5A, CLIP1, CUX1, GNAS and MGA were confirmed by Sanger sequencing.ConclusionsIn conclusion, we have identified pathogenic variants in genes associated with breast cancer predisposition. Importantly, many of these variants were significant in a second case cohort from dbGaP, suggesting that the strategy of using case–control analysis to select variants could potentially be utilized for identifying variants associated with cancer susceptibility.

123 (PB-036) Poster - Analysis of rare disruptive germline mutations in 2135 enriched BRCA-negative breast cancer cases excludes additional high-impact susceptibility genes

Background: To better understand the unexplained ∼60% of the genomic architecture of breast cancer susceptibility, we undertook germline whole exome sequencing (WES) of 2,135 BRCA-negative female breast cancer cases. Our series was highly enriched for early-onset, bilateral, family history of disease and concomitant ovarian cancer, substantially boosting power for discovery.

Prognostic Prediction of BRCA Mutations by 18F-FDG PET/CT SUVmax in Breast Cancer

Objectives:This study aimed to investigate the prognostic prediction of germline BRCA1 and BRCA2 mutations by comparing the maximum standardized uptake value (SUVmax) obtained from 18fluoride-fluorodeoxyglucose positron emission tomography/computed tomography (18F-FDG PET/CT), which is considered a prognostic factor in breast cancer (BC).Methods:Retrospective interdisciplinary laboratory results of 92 patients with BC who had germline BRCA1 or BRCA2 mutation profiles and underwent 18F-FDG PET/CT were compared. Genotyping was made by next-generation sequencing, and PET/CT scans were re-evaluated. The histopathological data, genetic results, and clinical demographics of all patients were recorded. Patients were divided into two groups in accordance with the presence of germline BRCA1 and/or BRCA2 mutations. Between-group statistical comparison was performed.Results:In PET/CT performed for primary staging, patients with BRCA-positive BC had significantly higher SUVmax (p=0.039), larger tumor size (p=0.025), and presence of axillary nodal metastases (p=0.023) than patients with BRCA-negative BC. Although the Ki-67 index was higher in the BRCA-positive group than BRCA-negative group, this difference was not significant (p=0.157). Moreover, in the BRCA-positive and negative groups, SUVmax, Ki-67 index, and tumor size, grade, and stage were significantly correlated with each other.Conclusion:The results of this study showed a strong association between BRCA mutations and SUVmax, which indicates the poor prognosis of BC.

Multiple cancer susceptible genes sequencing in BRCA-negative breast cancer with high hereditary risk.

BackgroundHereditary factors contributed to breast cancer susceptibility. Low BRCA mutation prevalence was demonstrated in previous BRCA mutation screening in Chinese breast cancer patients. Multiple-gene sequencing may assist in discovering detrimental germline mutation inBRCAnegative breast cancers.MethodsA total of 384 Chinese subjects with any two of high-risk factors were recruited and screened by next-generation sequencing (NGS) for 30 cancer susceptible genes. Variants with a truncating, initiation codon or splice donor/acceptor effect, or with pathogenicity demonstrated in published literature were classified into pathogenic/likely-pathogenic mutations.ResultsIn total, we acquired 39 (10.2%) patients with pathogenic/likely-pathogenic germline mutations, including one carrying two distinct mutations. Major mutant non-BRCA genes were MUTYH (n=11, 2.9%), PTCH1 (n=7, 1.8%), RET (n=6, 1.6%) and PALB2 (n=5, 1.3%). Other mutant genes included TP53 (n=3, 0.8%), RAD51D (n=2, 0.5%), CHEK2 (n=1, 0.3%), BRIP1 (n=1, 0.3%), CDH1 (n=1, 0.3%), MRE11 (n=1, 0.3%), RAD50 (n=1, 0.3%) and PALLD (n=1, 0.3%). A splicing germline mutation, MUTYH c.934-2A>G, was a hotspot (9/384, 2.3%) in Chinese breast cancer.ConclusionsAmong BRCA-negative breast cancer patients with high hereditary risk in China, 10.2% carried mutations in cancer associated susceptibility genes. MUTYH and PTCH1 had relatively high mutation rates (2.9% and 1.8%). Multigene testing contributes to understand genetic background of BRCA-negative breast cancer patients with high hereditary risk.

Abstract 4723: Genetic ablation of pregnancy zone protein (PZP) promotes breast cancer progression by activating TGF-β/SMAD signalling

Abstract A significant portion of breast cancer (BC) incidence is attributed to a hereditary predisposition to the disease. Germ-line variants in known BC-associated genes explain less than 30% of women with clinical signs of hereditary cancer syndrome. Using the whole-exome sequencing (WES) of genetically enriched BRCA-negative BC patients, we identified six novel candidate variants that are likely to contribute to BC predisposition. Particularly, the obtained data argued that pregnancy zone protein gene PZP might function as a tumor suppressor gene, likewise protein-truncating nonsense variant p.Arg680* could represent a moderate-risk breast cancer susceptibility allele. PZP is reported to bind with TGFβ1/β2 proteins, therefore, we hypothesize that the loss of PZP function may affect TGFβ signalling in breast cancer. We analysed the functional consequences of PZP nonsense mutation on TGFβ/SMAD signalling using CRISPR/Cas9 based genetic ablation of PZP in MCF7 and T47D breast cancer cells. GFP positive cells were sorted using FACS Aria III (BD biosciences) and genome editing events were identified using T7 endonuclease assay followed by DNA sequencing. PZP knockout cells were further verified by Anti-PZP Antibody (Abcam, UK) targeting loop region of PZP. The modulation of TGFβ signalling was quantified using qRT-PCR of TGFβ ligand/receptor mRNAs. The upregulation of TGFβ/SMAD signalling was evaluated by phospho-SMAD2, -SMAD3 protein levels. The wound healing and colony-forming assays were performed to compare cell migration and proliferation index in PZP wild-type [MCF7PZPwt, T47DPZPwt] versus PZP-deficient BC cells. DNA sequencing and Western blot confirmed the successful knockout using CRISPR/Cas9 approach [MCF7PZPsgRNA1, MCF7PZPsgRNA2, T47DPZPsgRNA1, and T47DPZPsgRNA2]. The PZP knockout cells have shown 2-2.8-fold increase in IC50 compared to wild-type cells following tamoxifen treatment (T47DPZPwt vs T47DPZPsgRNA1: 3.45µM vs 7.90µM; MCF7PZPwt vs MCF7PZPsgRNA1: 8.3µM vs 21µM). We documented the significant up-regulation of TGFβ1, β2 and βR2 transcripts in PZP-knockout clones. Of note, a very high up-regulation of TGFβ2 was observed in MCF7PZPsgRNA1 (197-fold), MCF7PZPsgRNA2 (498-fold) cells, whereas T47DPZPsgRNA1, T47DPZPsgRNA2 cells showed 2.8 and 5.5 fold up-regulation compared to wild-type cells (p=0.0001). Further, we reported an increase in SMAD2 and SMAD3 phosphorylation in PZP knockout cells. The relative wound width of PZP knockout cells significantly reduced compared to wildtype (p=0.0045). The clonogenic potential of PZP knockout cells was 3.5-fold higher compared to the wild-type counterpart. Our findings suggest that loss-of-function of PZP may favour breast cancer progression by up-regulating TGF-β/SMAD signalling. The research is supported by Indo Russia grant # INT/RUS/RSF/P-11 and RSF grant # 19-15-00207. Citation Format: Rohit Kumar, Ekaterina Kuligina, Anna Sokolenko, Quadir Siddiqui, Ashok K. Varma, Syed K. Hasan. Genetic ablation of pregnancy zone protein (PZP) promotes breast cancer progression by activating TGF-β/SMAD signalling [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 4723.

Abstract P3-07-11: BRCA mutations and association with discordance in a large oncotype database

Abstract BACKGROUND: BRCA mutated tumors have been shown to be associated with higher recurrence scores as compared to BRCA negative breast cancer patients. Prior studies have demonstrated 7-19% discordance between oncotype RS and tumor grade (TG) in breast cancer patients. We developed a large oncotype database to determine if BRCA mutation status is associated with discordance. METHODS: We identified 723 patients (32 (4%) BRCA positive patients and 691 (96%) BRCA negative patients) with early-stage, hormone positive BC treated between 2006-2018, with tumor characteristics available for analysis. Discordance was defined as one or two-step difference between RS (low, intermediate, high risk) and TG (well (WD), moderately (MD) and poorly (PD) differentiated). BRCA status was defined as BRCA 1 deleterious mutation, BRCA 2 deleterious mutation, BRCA mutation of unknown type, BRCA variant of undetermined significance (VUS) or other mutation. N(%) or median were used to describe patients’ characteristics between groups and compared by Mann-Whitney U test at the confidence level of 5%. RESULTS: Among these patients, there were 32 patients (4% of total) who had any documented mutation or VUS. 16% had documented deleterious mutation in BRCA 1, 22% BRCA 2, 6% BRCA unknown type, 25% BRCA VUS, and 31% other VUS (most commonly CHEK2 and ATM). Median RS was 23.5 in patients with deleterious BRCA mutations (1, 2 or unknown) vs. 16 in patients in the BRCA negative database which was statistically significant (p&amp;lt;0.01). One step and two-step discordance was present in 46% and 8% respectively of patients with deleterious BRCA mutations vs. 53% and 11% respectively in the BRCA negative database. CONCLUSIONS: Patients with deleterious BRCA mutations demonstrated no difference in rates of discordance as compared to BRCA negative patients. We further demonstrated that patients with BRCA positive tumors display higher recurrence scores than patients with BRCA negative tumors. Citation Format: Julia Blanter, Brittney Zimmerman, Serena Tharakan, Krystal Cascetta, Meng Ru, Amy Tiersten. BRCA mutations and association with discordance in a large oncotype database [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P3-07-11.

The Frequency and Management of TP53 Mutation Carriers in Turkish Patients with BRCA-Negative Breast Cancer Under 50 Years of Age

The Frequency and Management of TP53 Mutation Carriers in Turkish Patients with BRCA-Negative Breast Cancer Under 50 Years of Age

Germline variants in cancer genes in high-risk non-BRCA patients from Puerto Rico

Inherited pathogenic variants in genes that confer moderate to high risk of breast cancer may explain up to 50% of familial breast cancer. This study aimed at identifying inherited pathogenic variants in breast cancer cases from Puerto Rico that were not linked to BRCA1 or BRCA2. Forty-eight breast cancer patients that met the clinical criteria for BRCA testing but had received a negative BRCA1/2 result were recruited. Fifty-three genes previously implicated in hereditary cancer predisposition were captured using the BROCA Agilent cancer risk panel followed by massively parallel sequencing. Missense variants of uncertain clinical significance in CHEK2 were evaluated using an in vitro kinase assays to determine their impact on function. Pathogenic variants were identified in CHEK2, MUTYH, and RAD51B in four breast cancer patients, which represented 8.3% of the cohort. We identified three rare missense variants of uncertain significance in CHEK2 and two variants (p.Pro484Leu and p.Glu239Lys) showed markedly decreased kinase activity in vitro comparable to a known pathogenic variant. Interestingly, the local ancestry at the RAD51B locus in the carrier of p.Arg47* was predicted to be of African origin. In this cohort, 12.5% of the BRCA-negative breast cancer patients were found to carry a known pathogenic variant or a variant affecting protein activity. This study reveals an unmet clinical need of genetic testing that could benefit a significant proportion of at-risk Latinas. It also highlights the complexity of Hispanic populations as pathogenic factors may originate from any of the ancestral populations that make up their genetic backgrounds.

BRCA1/BRCA2Pathogenic Variant Breast Cancer: Treatment and Prevention Strategies

Hereditary breast cancer is known for its strong tendency of inheritance. Most hereditary breast cancers are related to BRCA1/BRCA2 pathogenic variants. The lifelong risk of breast cancer in pathogenic BRCA1 and BRCA2 variant carriers is approximately 65% and 45%, respectively, whereas that of ovarian cancer is estimated to be 39% and 11%, respectively. Therefore, understanding these variants and clinical knowledge on their occurrence in breast cancers and carriers are important. BRCA1 pathogenic variant breast cancer shows more aggressive clinicopathological features than the BRCA2 pathogenic variant breast cancer. Compared with sporadic breast cancer, their prognosis is still debated. Treatments of BRCA1/BRCA2 pathogenic variant breast cancer are similar to those for BRCA-negative breast cancer, mainly including surgery, radiotherapy, and chemotherapy. Recently, various clinical trials have investigated poly (adenosine diphosphate [ADP]-ribose) polymerase (PARP) inhibitor treatment for advanced-stage BRCA1/BRCA2 pathogenic variant breast cancer. Among the various PARP inhibitors, olaparib and talazoparib, which reached phase III clinical trials, showed improvement of median progression-free survival around three months. Preventive and surveillance strategies for BRCA pathogenic variant breast cancer to reduce cancer recurrence and improve treatment outcomes have recently received increasing attention. In this review, we provide an information on the clinical features of BRCA1/BRCA2 pathogenic variant breast cancer and clinical recommendations for BRCA pathogenic variant carriers, with a focus on treatment and prevention strategies. With this knowledge, clinicians could manage the BRCA1/BRCA2 pathogenic variant breast cancer patients more effectively.

Abstract P5-09-18: Multiple cancer susceptible genes sequencing in BRCA-negative breast cancers with high hereditary risk

Abstract Background Breast cancer susceptibility is strongly associated with a patient's hereditary background. Comprehensive BRCA mutation screening in our cancer center shows a lower BRCA mutation prevalence (9.1%) in Chinese breast cancer patients with hereditary risk. Multiple cancer susceptible genes sequencing can assist in discovering detrimental germline mutation in BRCA-negative breast cancers. Methods From 2005-2014, BRCA-negative breast cancer patients with any two of the following risk factors were recruited: (1) pathological diagnosis of triple-negative breast cancer, (2) male breast cancer, (3) primary bilateral breast cancers in one individual, regardless of synchrony or asynchrony, (4) early-age onset breast cancer (less than or equal to 40 years of age at diagnosis), or (5) patients with a family history of breast and/or ovarian cancer (at least one first- and/or second-degree relative with breast cancer or ovarian cancer). A total of 384 Chinese subjects were screened by next-generation sequencing for 32 breast cancer associated susceptibility genes (APC, ATM, BARD1, BMPR1A, BRIP1, CDH1, CDK4, CDKN2A, CHEK2, EPCAM, MEN1, MET, MLH1, MRE11A, MSH2, MSH6, MUTYH, NBN, NF1, PALB2, PALLD, PMS2, PTCH1, PTEN, RAD50, RAD51C, RAD51D, RET, SMAD4, STK11, TP53, VHL). Mutations were classified as pathogenic/likely-pathogenic if they had a truncating, initiation codon or splice donor/acceptor effect, or if pathogenicity was demonstrated in published literature. Results In total, we acquired 39 patients (10.2%) with pathogenic/likely-pathogenic germline mutations, including one carrying two distinct mutations. Major mutant non-BRCA genes were MUTYH (n=11), PTCH1 (n=7), RET (n=6) and PALB2 (n=5). Other mutant genes included TP53 (n=3), RAD51D (n=2), CHEK2 (n=1), BRIP1 (n=1), CDH1 (n=1), MRE11 (n=1), RAD50 (n=1) and PALLD (n=1). In addition, PTCH1 mutation carriers were found to be associated with the clinical features of triple-negative and early-age onset breast cancer. Conclusions Among BRCA-negative breast cancer patients with high hereditary risk in China, 10.2% carried mutations in breast cancer associated susceptibility genes. MUTYH and PTCH1 had relatively high mutation rates (2.9% and 1.8%). Some clinical features might be associated with germline mutations of particular genes in breast cancer. Key words germline mutation, BRCA-negative, hereditary breast cancer, multiple-gene sequencing Citation Format: Lang G-T, Hu X, Shi J-X, Huang W, Shao Z-M. Multiple cancer susceptible genes sequencing in BRCA-negative breast cancers with high hereditary risk [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P5-09-18.

BRCA carriers have similar reproductive potential at baseline to noncarriers: comparisons in cancer and cancer-free cohorts undergoing fertility preservation

To investigate whether BRCA carriers with and without malignancy have decreased ovarian reserve at baseline compared with BRCA noncarriers. Retrospective cohort study. Academic medical center. Seven-hundred and ninety-five oocyte cryopreservation patients, comprising BRCA carriers with and without malignancy (n = 57) and BRCA noncarriers (n = 738). Fertility preservation with oocyte cryopreservation. Antral follicle count (AFC), antimüllerian hormone (AMH) concentration, day-3 follicle-stimulating hormone (FSH) level, number of harvested oocytes, and number of mature/cryopreserved oocytes. In the cancer cohort we compared BRCA-positive breast cancer (n = 38) with BRCA-negative breast cancer (n = 53) and with non-breast-cancer malignancies (n = 85). In the cancer-free cohort we compared BRCA carriers (n = 19) with women undergoing elective egg freezing (n = 600). We also compared the BRCA1 (n = 31) versus the BRCA2 carriers (n = 18). The patients' mean ages were 32.4 ± 3.6years and 35.5 ± 4.3years in the BRCA carrier and noncarrier cohorts, respectively. BRCA status was associated with a higher day-3 FSH level in the cancer cohort, but we found no changes in the other outcomes compared with the BRCA-negative cancer groups. BRCA carriers without cancer exhibited a higher AFC and number of mature oocytes compared with the patients undergoing planned egg freezing. Overall (cancer and cancer-free cohorts), the BRCA carriers had an increased AFC (15.5 ± 4.6 vs. 12.6 ± 5.7) and number of mature/cryopreserved oocytes (14.0 ± 7.9 vs. 10.4 ± 6.9) compared with the BRCA noncarriers but had no differences in other outcomes. BRCA carriers with and without malignancy exhibit comparable ovarian reserve and responses to ovarian stimulation compared with women with BRCA-negative cancers and cancer-free controls.

Abstract 1236: Insight into genetic susceptibility to BRCA-negative male breast cancer by multigene panel testing: Results from a multicenter study in Italy

Abstract Background: Male breast cancer (MBC) is a rare disease, representing less than 1% of all breast cancers (BC). Although rare, MBC remains a substantial cause for morbidity and mortality in men. MBC etiology appears to be largely associated with genetic factors. Inherited mutations in BRCA2, PALB2 and BRCA1 genes, predispose to MBC and account for about 15% of all cases. Thus, a fraction of MBC cases remains to be assigned to specific genetic factors. Here, we aimed at investigating the genetic component of BRCA1/2 mutation negative MBC cases and at identifying germline mutations that could further explain MBC genetic susceptibility. Materials and methods: We performed a genomic screening of a well-characterized series of 502 BRCA1/2 mutation negative Italian MBC cases by a comprehensive multi-gene custom panel of 50 cancer-related genes, using MiniSeq platform (Illumina). We also compared the main clinical-pathologic characteristics of MBCs in mutation carriers and non-carriers using Fisher exact test and t-test, where appropriate. All statistical analyses were performed with the R software (www.r-project.org). Results: Overall 6% of BRCA1/2 mutation negative MBC cases were found to carry a pathogenic variant in the genes analyzed. In particular, PALB2 and ATM were the most frequently altered genes (1.1% and 0.7%, respectively). Mutations in known/proposed BC genes, such as BARD1, BLM, BRIP1, CASP8, CHEK2, FANCM, NBN, NF1, RAD50, RAD51C and RAD51D, as well as in genes considered not closely related to BC predisposition, such as APC, EPCAM and MUTYH, were also identified. Intriguingly, the same NBN mutation was identified in three unrelated MBC cases. Mutation carriers were more likely to have a personal history of cancer in addition to MBC (p=0.0045) and family history of cancer other than breast and ovarian cancer (p=0.0004). Conclusions: This study may help to gain more insight into the genetic susceptibility of MBC. Our results support the central role of PALB2 in MBC susceptibility, point to a relevant role of ATM and confirm a low impact of CHEK2 mutations on MBC predisposition in the Italian population. Considering that pathogenic mutations were found only in a fraction of the genes analyzed, our data indicate that the identification of the more appropriate genes for the genomic screening of MBC cases is essential in order to develop a comprehensive and specific MBC susceptibility panel with implications for clinical management and counselling of patients and their families. Moreover, our results suggest that multigene testing approach may benefit appropriately selected patients, especially those with a personal or family history of cancer other than breast/ovarian cancer. Study supported by AIRC (IG 16933) to L.O. Citation Format: Piera Rizzolo, Veronica Zelli, Valentina Silvestri, Virginia Valentini, Alessandro Spinelli, Maria Grazia Tibiletti, Antonio Russo, Liliana Varesco, Giuseppe Giannini, Daniele Calistri, Laura Cortesi, Alessandra Viel, Marco Montagna, Paolo Peterlongo, Paolo Radice, Domenico Palli, Laura Ottini. Insight into genetic susceptibility to BRCA-negative male breast cancer by multigene panel testing: Results from a multicenter study in Italy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1236.

PO-076 Molecular analysis of BRCA-negative breast and/or ovarian cancer families by multigene panel testing

IntroductionAbout 5%–10% of the hereditary breast and/or ovarian cancer (BC/BOC) is associated with an autosomal dominant genetic susceptibility due to highly penetrant mutations of the BRCA1/2 genes. In particular, BRCA1/2 gene mutations are found in 25%–30% of the BC families subjected to genetic testing. These numbers suggest the possible involvement of other genes in BC/BOC genetic predisposition and a fraction of these cases remains to be assigned to specific genetic factors. Here we report on the application of the NGS multigene panel to a group of BRCA1/2 mutation negative BC/BOC cases, in order to identify germline mutations that could further explain BC/BOC genetic susceptibility.Material and methodsWe selected a group of 27 BRCA1/2 negative BC and BOC families on the basis of a clear dominant inheritance pattern and/or a moderate/high BRCAPro score. We performed a genomic screening by a comprehensive multi-gene custom panel of 29 cancer-related genes, using Ion Torrent platform (Thermo Fisher Scientific).Results and discussionsIn three cases (11%) we found mutations described as pathogenic (https://www.ncbi.nlm.nih.gov/clinvar/) in ATM, MUTYH and PALB2 genes. In the series analysed, the most frequently altered genes were APC and ATM (15%) but were also identified mutations in MSH6 and TP53 (11%), MUTYH and RAD51B (7%), MRE11, EPCAM, BRIP1, CHEK2, PALB2, BARD1, STK11 and RAD50 (4%). In particular, we found six genomic variants of uncertain significance (VUS) in MSH6, ATM, BRIP1, RAD50 and APC genes; nine genomic variants of conflicting interpretations of pathogenicity in MUTYH, MRE11, TP53, APC, MSH6, CHEK2, EPCAM and ATM genes and eight genomic variants not reported in ClinVar in APC, RAD51B, STK11, TP53, ATM and BARD1 genes predicted deleterious by in silico analysis. Their biological significance and involvement in the development of the pathology is still unknown today. Only six patients were negative for the presence of mutations in the 29 genes analysed.ConclusionPreliminary results of this study suggest that NGS could offer a great contribution to identify the genetic component of susceptibility to BC/BOC and could potentially be used with implications for clinical management and counselling of patients and their families. Moreover, our results suggest that multigene testing approach may benefit appropriately selected patients, especially those with increased risk of BC/BOC development.

Germline mutations of TP53 gene among Chinese families with high risk for breast cancer

To evaluate the role of germline mutations of TP53 gene among a Chinese population with high risk for breast cancer. A total of 81 BRCA-negative breast cancer probands from cancer families were analyzed using targeted capture and next-generation sequencing. Candidate mutations were verified with Sanger sequencing. Co-segregation analyses were carried out to explore the likely pathogenicity of the mutation. Of the 81 BRCA-negative patients, 3 exonic mutations in the TP53 gene were identified in 3 breast cancer patients. Of these, 2 mutations were previously reported and 1 was novel. One family with TP53 mutation has met the criteria for Li-Fraumeni syndrome (LFS) and accounted for 9.1% of all families who fulfilled the diagnostic criteria for LFS. Two of the carriers were diagnosed with breast cancer under the age of 30, and have accounted for 11.8% (2/17) of all very young (≤30 years) breast cancer patients in our study. The TP53 germline mutation is more common in Chinese population with a high risk for breast cancer than previously thought. TP53 gene mutation screening should be considered particularly for patients with a family history of LFS and very young age of onset.

Abstract 2760: Impact of race and age on choice of surgery in newly diagnosed breast cancer patients who tested negative for BRCA 1/2 mutation

Abstract Introduction: The benefit of bilateral mastectomies in breast cancer patients who test negative for a deleterious BRCA mutation has not been proven. In spite of this, a significant number of women with unilateral breast cancer who test negative for BRCA mutation choose bilateral mastectomies. We aimed to evaluate the role of age and race in such decision making. Methodology: Records of 323 patients referred to Cancer Genetics between January 01,2012 and June 30,2014 for pre-operative genetic evaluation were analyzed. This cohort consisted of patients with newly diagnosed breast cancer who met genetic testing criteria. Patients were excluded if their surgical records were not available, they declined genetic testing, they underwent surgery prior to their test results being available or they were positive for BRCA mutation. A total of 148 patients met our inclusion criteria and were evaluated for final analysis. Statistical analysis was performed using SPSS21. Results: Of the 148 women with breast cancer who tested negative for a deleterious BRCA mutation, 116(78%) were Caucasian, 9(6%) were African-American, 5(3%) were Asian, 4(3%) were Arab, 3(2%) were Indian, 2(2%) were American-Indian and 1(1%) was Hispanic. The race or ethnicity could not be identified in 8(5%) patients. A total of 47(32%) patients ultimately underwent bilateral mastectomies in this cohort. Of these, 40(27%) underwent bilateral mastectomies as their initial surgery while 7(5%) underwent bilateral mastectomies eventually after undergoing partial mastectomy or simple mastectomy initially. There was no statistically significant difference between the mean ages of the group that underwent bilateral mastectomy (48.9 yrs) and the group that underwent partial or simple mastectomy (49.1 yrs). All of the 8 patients with an unknown ethnicity underwent bilateral mastectomy. The rest of the mastectomies (39) were performed in Caucasian patients. None of the non-Caucasian patients underwent bilateral mastectomy. This difference was statistically significant (p&amp;lt;0.001). Conclusion: Our study demonstrates that a significant proportion(32%) of BRCA negative women with breast cancer undergo bilateral mastectomies. Furthermore, Caucasian patients with BRCA negative breast cancer are more likely to choose bilateral mastectomies than non-Caucasians. However, this result has to be interpreted with caution as our sample size of non-Caucasians was small(24). Considering our catchment population, this small sample brings concerns that a significant proportion of non-Caucasian patients may not be undergoing genetic testing despite meeting criteria. We did not find a significant impact of age on choice of surgery in BRCA negative women with breast cancer. Further studies with larger sample size are needed to better understand the role of race and age in access to genetic testing and surgical decision making. Citation Format: Siddhartha Yadav, Heidi Dreyfuss, Jennifer Fulbright, Ashley Reeves, Sarah Campian, Dana Zakalik. Impact of race and age on choice of surgery in newly diagnosed breast cancer patients who tested negative for BRCA 1/2 mutation. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2760. doi:10.1158/1538-7445.AM2015-2760

Cancer risks in women from BRCA-negative hereditary breast and ovarian cancer families

Cancer risks in women from BRCA-negative hereditary breast and ovarian cancer families

Abstract LB-191: Cytological atypia predicts short-term breast cancer risk in women from high-risk families that lack a BRCA mutation.

Abstract Background: Women with deleterious mutations in BRCA1/2 have a 9- to 36-fold increased risk of breast cancer compared with general population rates. However, there are a significant number of women whose families have multi-generation premenopausal breast cancers and lack an identified BRCA-mutation. Currently we lack strategies to predict the development of breast cancer in women from BRCA-negative families. Random Periareolar Fine Needle Aspiration (RPFNA) is a research technique developed to test for “field effects” in high-risk women. Carol Fabian previously demonstrated that atypia in RPFNA predicted breast cancer risk, however, many women in this cohort were known BRCA mutation carriers. We investigated whether cytological atypia in RPFNA predicts the subsequent development of breast cancer in women from BRCA-negative breast cancer families. Results: From 01/01/03 to 07/01/12, we performed RPFNA in 254 high-risk BRCA-negative women. Women underwent yearly mammogram/Magnetic Resonance Imaging (MRI) and bi-annual clinical breast exam. During 6.5 women-years of follow-up, 24 new breast cancers were diagnosed. The median age of the 254 women in this study was 48, and the median BMI was 24. Eighty-six percent (218) of the women were Caucasian, 13% (32) were African-American, and 2 women were Native Americans. Sixty-eight women presented with cytologic-atypia in at least one breast; 16 of these women had cytologic-atypia in both breasts. Fourteen (21%) of the 68 women with baseline cytologic-atypia later developed cancer in a breast that had had baseline cytologic-atypia; none of the 68 women developed cancer in a breast without baseline cytologic-atypia. One hundred eighty-six women did not have cytologic-atypia at baseline; 10 (5%) of these women developed cancer. Ten women had a prophylactic mastectomy and were censored at the date of mastectomy. The median length of follow-up among the uncensored patients was 74 months. In the baseline atypic and non-atypic groups, the 48-month cumulative incidence of cancer development was 0.22 (95% confidence interval [CI], 0.13-0.34) and 0.04 (95% CI, 0.02-0.08), respectively. Atypia was significantly associated with cancer development both with and without controlling for covariates (p &amp;lt; 0.001). The covariate-adjusted hazard ratio (CV-AHR) for cytologic-atypia was 6.3 (95% CI, 1.7-24.2). The hazard ratios for the covariates were as follows: age (1.01; 95% CI, 0.89-1.16), BMI (1.02; 95% CI, 0.91-1.15), menopausal status (0.16; 95% CI, 0.01-2.60), Gail Risk (1.18; 95% CI, 1.03-1.35) and mammographic density for scattered (1.7; 95% CI, 0.27-10.9) and heterogenetic (1.10; 95% CI, 0.20-6.01) compared to the reference type of extreme. Conclusions: This is the first demonstration that in women from high-risk BRCA mutation-negative families, cytologic-atypia in RPFNA is associated with the subsequent development of breast cancer. Citation Format: Victoria L. Seewaldt, Bercedis Peterson, Gloria Broadwater. Cytological atypia predicts short-term breast cancer risk in women from high-risk families that lack a BRCA mutation. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr LB-191. doi:10.1158/1538-7445.AM2013-LB-191

733 Common Breast Cancer Susceptibility Alleles in BRCA-positive and BRCA-negative Male Breast Cancer

733 Common Breast Cancer Susceptibility Alleles in BRCA-positive and BRCA-negative Male Breast Cancer

Identification of new susceptibility genes in familial breast cancer by exome sequencing.

1538 Background: Familial breast cancer was described in the early 1970s with its syndromy confirmed by the discovery of BRCA germline mutations in the 1990s. BRCA mutations account for less than 10% of affected families. Efforts made in the past two decades have resulted in limited results in identifying additional susceptibility genes for familial breast cancer. Methods: Using exome sequencing, we analyzed eight members in a BRCA1-, BRCA2-, p53- and PTEN-negative breast cancer family; five with breast cancer and three unaffected. Mutation candidates were idenified by bioinformatics analysis, experimentally validated by Sanger sequencing and their damaging effect was predicted by the SIFT program. Validated mutations were also tested in 42 additional breast cancer samples from BRCA-negative breast cancer families. Results: We identified 55 non-synonymous germline mutations affecting 49 genes in multiple members of this family, 20 of 22 selected mutations predicted to cause damaging effects were validated. As an exemplar, two mutations in the MYST4 gene were detected at the C terminal (p.D1516Y and p.R1577C), validated, and predicted to cause damaging consequences. MYST4 is a histone acetyltransferase (Champagne, N. et al. JBC. 274, 28528-28536,1999). It contains a C2HC-type finger and a PHD-type zinc finger. Its N-terminal is involved in transcriptional repression while its C-terminal is involved in transcriptional activation and interacts with important transcriptional regulators RUNX1 and RUNX2. MYST4 is involved in DNA replication, transcriptional regulation, and epigenetic modification of chromatin structure. A translocation t(10;16)(q22;p13) between CREBBP and MYST4 was identified in acute myeloid leukemia. The 20 validated germline mutations were tested by Sanger sequencing in 42 additional breast cancer cases from 26 BRCA-negative families. None of the 20 mutations were detected. Conclusions: Results show that remaining unknown susceptibility genes are likely family-specific and can be detected in each affected family using genome sequencing approaches. These results may be the first reported in an exome sequencing study of BRCA-negative breast cancer families.