Breast Cancer BRCA2 Research Articles

Assessment of DNA Repair Gene Expressions in Vitrified Mouse Preantral Follicles.

Objective Vitrification of the ovarian tissue is one of the techniques recommended for preserving the fertility of women who are dealing with infertility. Despite its benefits, our information about the molecular aspects of ovarian follicles vitrification is somehow ambiguous. Therefore, the aim of this study was to evaluate the expression pattern of DNA repair genes in vitrified preantral follicles. Materials and Methods In this experimental study, the isolated preantral follicles (n=906) from 14-16 days old mice (n=12) were divided into three groups: fresh, toxic and vitrified which were cultured in vitro for 12 days. Preantral follicles were vitrified using cryotop followed by exposure to equilibration solution for five minutes and vitrification solution (VS) for 30 seconds. In the toxic group, preantral follicles were only placed in equilibration and vitrification media and they were then placed in the warming solutions without exposure to liquid nitrogen. On the second and sixth days of the culture period, real-time quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was carried out to evaluate expression of the selected genes involved in DNA repair, including Msh6 (MutS homolog 6), Mre11 (Meiotic recombination 11), Brca1 (Breast cancer type 1), Rad51 (RAD51 recombinase), Pcna (Proliferating cell nuclear antigen) and Atm (ATM serine/threonine kinase). In addition, developmental parameters including growth, survival rate, antrum cavity formation and ovulation were analyzed. Results The relative mRNA expression of Msh6, Mre11, Brca1, Rad51, Pcna and Atm on the second and sixth days of the culture period in vitrified group was significantly higher than those of the control and toxic groups, but there was no significant difference between the toxic and control groups. In addition, developmental parameters of follicles were similar in both toxic and control groups, while both were significantly higher than that of vitrified group. ConclusionVitrification changes the expression pattern of DNA repair genes of the mouse preantral follicles.

Next-Generation Sequencing Identifies BRCA1 and/or BRCA2 Mutations in Women at High Hereditary Risk for Breast Cancer with Shorter Telomere Length.

Telomeres, and telomere length in particular, have broad significance for genome biology and thus are prime research targets for complex diseases such as cancers. In this context, BRCA1 and BRCA2 gene mutations have been implicated in relationship to telomere length, and breast cancer susceptibility. Yet, the linkages among human genetic variation and telomere length in persons with high hereditary cancer risk are inadequately mapped. We report here original findings in 113 unrelated women at high hereditary risk for breast cancer, who were characterized for the BRCA1 and BRCA2 mutations using next-generation sequencing. Thirty-one BRCA2 and 21 BRCA1 mutations were identified in 47 subjects (41.6%). The women with a mutation in BRCA1 and/or BRCA2 genes had, on average, 12% shorter telomere compared to women with no BRCA1 or BRCA2 mutation (p = 0.0139). Moreover, the association between telomere length and BRCA mutation status held up upon stratified analysis in those with or without a breast cancer diagnosis. We also indentified two rare mutations, c.536_537insT and c.10078A>G, and a novel mutation c.8680C>G in BRCA2 that was studied further by homology modeling of the DNA binding tower domain of BRCA2 and the structure of the protein. These data collectively lend evidence to the idea that BRCA1 and BRCA2 mutations play a role in telomere length in women at high hereditary risk for breast cancer. Further clinical and diagnostics discovery research on BRCA1 and BRCA2 variation, telomere length, and breast cancer mechanistic linkages are called for in larger study samples.

Response to Olaparib in a Patient with Germline BRCA2 Mutation and Breast Cancer Leptomeningeal Carcinomatosis

Leptomeningeal carcinomatosis (LC) is a devastating complication of metastatic cancer that disproportionately affects patients with advanced breast cancer. Moreover, those with BRCA1/2-mutated disease more often experience leptomeningeal metastasis. Treatment options for LC are limited and often include significant toxicities. PARP inhibitors offer an important potential treatment for patients with BRCA1/2-mutated breast and ovarian cancers, but clinical studies excluded patients with central nervous system (CNS) metastases, including LC. Efficacy data in this area are therefore limited, although a phase I study of olaparib in glioblastoma did show CNS penetration. Here we report a case of a patient with BRCA2-mutated breast cancer and solitary recurrence in the leptomeninges with ongoing complete response to treatment with the PARP inhibitor olaparib. PARP inhibitors may be an important treatment option for patients with BRCA-mutated disease and LC, and warrant further study.

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.

733P - Biliary tract cancers in Portuguese families with BRCA gene mutation: A retrospective study

BackgroundBiliary tract cancers (BTCs) are rare. They are clinically and pathologically heterogeneous and associated with a poor prognosis. BRCA 1/2 are tumor suppressor genes responsible for a high number of hereditary breast and ovarian cancers. BRCA2 mutations are also associated with higher risk of other cancers, including prostate and pancreatic cancers. BTC-associated with BRCA mutation is still unclear. In this study, we analyzed the frequency and characteristics of BTCs in confirmed BRCA Portuguese families. MethodsRetrospective analysis of all BRCA1/2 families registered in our multidisciplinary program since January 2000 was performed. All cases of BTCs, on the mutated side of the family were included. ResultsFrom 513 BRCA1/2 families, 187 were BRCA1 and 325 BRCA2. BRCA2 c.156_157insAlu - the Portuguese founder mutation - was observed in 101 families. We identified BTCs in seven BRCA2 families, five of which were c.1567_157insAlu. Overall, 7 cases of BTCs were identified (5 with pathological confirmation): 5 BRCA2 carriers and 2 first-degree relatives of BRCA2 carriers. Median age at diagnosis was 63 years, with male and female prevalence rates of 57% and 43%, respectively. Two cases of BTC were diagnosed in two female survivors of breast cancer. One female survivor of BT developed peritoneal carcinoma after prophylactic salpingo-oophorectomy. All BRCA2 families revealed a heterogeneous phenotype: 5 out of 7 families have aggregation to pancreatic and/or stomach cancers. Four patients died, three from BTC disease progression and one from peritoneal carcinoma. The other 3 patients are alive without relapse. ConclusionsBRCA-associated BTCs are uncommon. Surveillance and prophylactic surgery reduces the risk of death from breast and ovarian cancer in BRCA2 carries allowing the diagnosis of other type of cancer during the follow-up, such as BTCs. Therefore, it is crucial the development of surveillance protocols for gastrointestinal tumours. Diagnosis of BTC-associated germline BRCA mutations might have clinical implications, expanding roles in treatment planning and allowing application of targeted therapies. Legal entity responsible for the studyThe authors. FundingHas not received any funding. DisclosureAll authors have declared no conflicts of interest.

Abstract 1747: Both somatic and germline BRCA mutations show high degree of bi-allelic loss in breast and ovarian cancers

Abstract PARP inhibitors (PARPi) are promising novel therapies for tumors harboring defects in homologous recombination (HR) DNA repair. Loss of function (LoF) mutations in the tumor suppressors BRCA1 and BRCA2, two critical components in HR pathway, are the most common cause of HR deficiency in tumors. Complete loss of BRCA1/2 function requires bi-allelic loss, and it is hypothesized to be needed for PARPi sensitivity. However, the degrees of bi-allelic loss in breast and ovarian tumors are still lacking consensus, especially when both LoF events are somatic. Recent publications shown a high percentage of germline BRCA2 lacking LOH, including TCGA’s breast and ovarian cohorts. To better understand the loss of wild type BRCA genes, we reanalyzed raw data from TCGA using a novel pipeline to identify both germline and somatic BRCA mutations in breast and ovarian. We then applied mathematical models to determine LOH or bi-allelic loss of those mutations. Our analysis shows a higher degree of bi-allelic loss of BRCA mutations than previously reported, especially germline BRCA2 in breast cancer, which shows nearly 80%. Furthermore, somatic BRCA mutations, which account for about 1/3 of all BRCA mutations, showed a similar degree of bi-allelic loss as germline BRCA mutations. In addition, somatic and germline BRCA were shown to have comparable HRD scores, a measure of tumor genomic instability. Germline BRCA patients are typically diagnosed at younger ages in breast and ovarian cancer, whereas somatic show no difference to non-BRCA patients. In ovarian cancer, patients with somatic or germline BRCA had similar progression-free and overall survival with comparable treatment journeys, suggesting a phenotypic equivalence. Interestingly, BRCA2 tumors showed lower HRD scores than BRCA1 tumors in both breast and ovarian. In breast cancer BRCA1 mutation is significantly enriched in TNBC, but the degree of bi-allelic loss did not show any significant difference between ER+ and TNBC subtypes. To validate our findings, we examined a much larger cohort of breast (n > 5,000) and ovarian (n > 2,500) cancers sequenced by Foundation Medicine, with germline/somatic and LOH status determined computationally using validated methodology. We found similar prevalence of somatic BRCA mutations and the degree of bi-allelic loss and HRD scores in both germline and somatic BRCA mutations. In addition, potential reactivating reversion mutations were discovered in both somatic and germline mutated BRCA in both breast and ovarian cancers, suggesting they are driver mutations. In summary, large scale analysis of BRCA mutations in breast and ovarian cancers suggest that about 1/3 of BRCA mutations are somatic, and both germline and somatic BRCA mutations show high degree of bi-allelic loss and similar genomic instability and may benefit from PARPi therapies. Citation Format: Zhongwu Lai, Darren Hodgson, James Sun, Adrienne Johnson, Carl Barrett, Jonathan Dry. Both somatic and germline BRCA mutations show high degree of bi-allelic loss in breast and ovarian cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1747.

Abstract CT229: Olaparib in patients (pts) with previously treated, homologous recombination repair mutation (HRRm) or homologous recombination deficiency (HRD)-positive advanced cancer: Phase II LYNK-002 study

Abstract Background: Human cancers are associated with defects in DNA damage repair (DDR). Evidence suggests a key role for poly(ADP-ribose) polymerases (PARPs) in repairing certain DNA damage lesions. Olaparib (a PARP inhibitor) is approved for pts with germline BRCA1 and BRCA2-mutated breast and ovarian cancers, and pts with or without BRCA mutation in recurrent ovarian cancer. Mutations in HRR genes (HRRm; beyond BRCA1 or BRCA2) and HRD may sensitize tumors to respond to PARP inhibitors. Olaparib has also shown favorable ORRs in HRRm/HRD-positive (HRD+) cancers, including prostate and ovarian. Thus, this study (NCT03742895) aims to evaluate the use of HRRm/HRD+ as biomarkers of tumor-agnostic response to olaparib monotherapy. Methods: Eligibility for this open-label, Phase II study requires pts ≥18 years of age with a previously treated, histologically/cytologically confirmed, HRRm/HRD+ advanced solid tumor; who have failed, are intolerant to, or ineligible for all available standard of care therapies; with no disease progression during prior platinum-based treatment. At baseline, pts should also have measurable disease per RECIST v1.1 or Prostate Cancer Working Group (PCWG)-modified RECIST v1.1 (for pts with prostate cancer) without CNS metastases, ECOG PS of 0 or 1, and no persistent toxicities (CTCAE grade >2) from prior cancer therapy. Pts may have received any number of prior regimens. Newly obtained or archival tumor samples will be centrally evaluated using the Lynparza HRR Assay (under development at Foundation Medicine, Inc). Pts will be grouped into 2 cohorts: cohort 1 will include pts with any solid tumor type with a BRCA1 or BRCA2 mutation (excluding breast and ovarian cancers); cohort 2 will include pts whose tumor (any type) is BRCA1 or BRCA2 wild-type and HRRm and/or HRD+ as determined by loss of heterozygosity score. Pts will receive olaparib 300 mg orally BID until documented disease progression, unacceptable toxicity, or study withdrawal. Tumor imaging will be performed at baseline, with response assessments Q8W in the first year and Q12W thereafter, per RECIST v1.1 by BICR or PCWG-modified RECIST v1.1, both modified to follow ≤5 target lesions per organ (10 total). AEs will be graded using NCI CTCAE v4.0. The primary endpoint is ORR; the point estimate and 95% CI calculated using the Clopper-Pearson method will be provided. Summary statistics for key secondary efficacy endpoints of duration of response, OS, and PFS will be provided using the Kaplan-Meier method. Safety endpoints will be summarized. Approximately 370 pts will be enrolled (~84 in cohort 1 [BRCA1 and BRCA2 mutated]; ~174 HRRm and ~112 HRD+ in cohort 2 [BRCA1 and BRCA2 wild-type]). Enrollment is ongoing in 5 countries beginning December 12, 2018. Citation Format: David Hyman, Andrew Hendifar, Hyun Cheol Chung, Michele Maio, Alexandra Leary, Iben Spanggaard, Joon Rhee, Matt Marton, Menghui Chen, Suba Krishnan, Ronnie Shapira. Olaparib in patients (pts) with previously treated, homologous recombination repair mutation (HRRm) or homologous recombination deficiency (HRD)-positive advanced cancer: Phase II LYNK-002 study [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr CT229.

Differential cell surface protein expression in response to PARP inhibition in BRCA1 mutant isogenic ovarian and breast cancer cells using a novel proteomic technique

Differential cell surface protein expression in response to PARP inhibition in BRCA1 mutant isogenic ovarian and breast cancer cells using a novel proteomic technique

Inadequate DNA Damage Repair Promotes Mammary Transdifferentiation, Leading to BRCA1 Breast Cancer

Loss of BRCA1 p220 function often results in basal-like breast cancer (BLBC), but the underlying disease mechanism is largely opaque. In mammary epithelial cells (MECs), BRCA1 interacts with multiple proteins, including NUMB and HES1, to form complexes that participate in interstrand crosslink (ICL) DNA repair and MEC differentiation control. Unrepaired ICL damage results in aberrant transdifferentiation to amesenchymal state of cultured, human basal-likeMECs and to a basal/mesenchymal state in primary mouse luminal MECs. Loss of BRCA1, NUMB,or HES1 or chemically induced ICL damage in primarymurine luminal MECs results in persistent DNA damage that triggers luminal to basal/mesenchymal transdifferentiation. Invivo single-cell analysis revealed a time-dependent evolution from normal luminal MECs to luminal progenitor-like tumor cells with basal/mesenchymal transdifferentiation during murine BRCA1 BLBC development. Growing DNA damage accompanied this malignant transformation.

Graphene-Coated Optical Fiber SPR Biosensor for BRCA1 and BRCA2 Breast Cancer Biomarker Detection: a Numerical Design-Based Analysis

This paper provides a simple hybrid design and numerical analysis of the graphene-coated fiber-optic surface plasmon resonance (SPR) biosensor for breast cancer gene-1 early onset (BRCA1) and breast cancer gene-2 early onset (BRCA2) genetic breast cancer detection. Two specific mutations named 916delTT and 6174delT in the BRCA1 and BRCA2 are selected for numerical detection of breast cancer. This sensor is based on the technique of the attenuated total reflection (ATR) method to detect deoxyribonucleic acid (DNA) hybridization along with individual point mutations in BRCA1 and BRCA2 genes. We have numerically shown that momentous changes present in the SPR angle (minimum: 135% more) and surface resonance frequency (SRF) (minimum: 136% more) for probe DNA with various concentrations of target DNA corresponding to a mutation of the BRCA1 and BRCA2 genes. The variation of the SPR angle and SRF for mismatched DNA strands is quite negligible, whereas that for complementary DNA strands is considerable, which is essential for proper detection of genetic biomarkers (916delTT and 6174delT) for early breast cancer. At last, the effect of electric field distribution in inserting graphene layer is analyzed incorporating the finite difference time domain (FDTD) technique by using Lumerical FDTD solution commercial software. To the best of our knowledge, this is the first demonstration of such a highly efficient biosensor for detecting BRCA1 and BRCA2 breast cancer. Therefore, the proposed biosensor opens a new window toward the detection of breast cancers.

Associations with response to poly (ADP-ribose) polymerase (PARP) inhibitors in metastatic breast cancer: Results of a meta-regression analysis.

e12567 Background: PARP inhibitors (PARPi), when given as single agents, have modest antitumor activity in patients with advanced breast cancer and germline mutation in BRCA1 or BRCA2. It is unclear whether some subgroups derive greater benefit from treatment. Methods: Two electronic databases (MEDLINE, CENTRAL) and one registry (Clinicaltrials.gov) were searched from inception to November 2018 to identify trials of PARPi in patients with metastatic breast cancer. The response rates to PARPi were extracted and pooled. Analyses were performed for patients with a germline BRCA mutation. Meta-regression explored the influence of patient and tumor characteristics and previous chemotherapy on the objective response rate (ORR) as reported in individual studies. Analysis comprised of a linear regression weighted by individual study sample size using the weighted least squares (mixed effect) method. Quantitative significance was defined using methods described by Burnand et al. Results: Of 1855 citations, 11 studies comprising 813 patients were included in the analysis. 765 (94%) patients had a germline BRCA mutations; 48% of breast cancers were triple-negative. 76% of patients had received at least 1 previous line of chemotherapy in the metastatic setting and 30% were exposed to platinum-based chemotherapy. Pooled ORR was 47% in patients with a germline BRCA mutation. Meta-regression showed that previous chemotherapy in the metastatic setting ( = -0.94, p = 0.006), especially platinum-based chemotherapy ( = -0.89, p = 0.02) were associated with highly significant negative association with ORR. A highly quantitatively significant negative association was observed for age ( = -0.80, p = 0.10), but this did not meet statistical significance. Performance status ( = 0.44, p = 0.38) and hormone receptor status (hormone receptors positive: = 0.46, p = 0.30) were not associated with response. Conclusions: PARPi therapy is associated with lesser response in patient with prior chemotherapy exposure, especially platinum-based treatment. Younger patients may benefit more from PARPi. There was no association between ORR and hormone receptor status.

Brain metastasis in patients with hereditary BRCA-mutated invasive breast cancer.

1074 Background: In the past six months, two poly(ADP-ribose) polymerase (PARP) inhibitors have been approved for the treatment of patients with metastatic breast cancer and germline pathogenic variants in BRCA1 or BRCA2 ( gBRCA). In addition, recent data from the IMpassion 130 trial lends support for the role of immunotherapy in a subset of patients with triple negative breast cancer (TNBC). Approximately 60% of patients with gBRCA1 have TNBC. There is evidence that both PARP inhibitors and checkpoint inhibitors cross the blood-brain barrier and both classes of agents have entered clinical trials for patients with brain metastases in other tumor types. We studied the clinical course of breast cancer patients with gBRCA and brain metastasis at our institution to inform clinical trial design for this group of patients with poor outcomes. Methods: Patients with stage I to III invasive breast cancer, gBRCA, and eventual development of brain metastasis were identified from clinical databases. Data analyzed included breast cancer subtype and stage at diagnosis, treatment, time to distant recurrence and to discovery of brain metastasis, and overall survival from time of development of brain metastasis. Results: Patients in our cohort (n = 24, to date) were diagnosed at a young age (median age 39) and primarily had TNBC (21/24, 87.5%) with infiltrating ductal carcinoma histology. Nineteen patients had gBRCA1 and 4 patients had gBRCA2. All but 1 patient received anthracycline-based chemotherapy in the neoadjuvant/adjuvant setting. Median time to distant metastasis was 2 years (range: 0.8 – 15) and the brain was the first site of recurrence in 5 of 24 (21%) patients. Median time from diagnosis to development of brain metastasis was 2.6 years (range: 1.2 – 19) and most patients (18/25, 72%) had multiple brain metastases discovered on the initial brain MRI. Median overall survival (OS) was 3.7 years (range: 1.8 – 24) and median OS from the time of brain metastasis was 7 months (range: 1 month – 13 years). Conclusions: Breast cancer patients with germline pathogenic variants in BRCA1/2 who develop brain metastasis have a dismal prognosis. These patients may benefit from an agent with intracranial activity at the time of first distant recurrence.

BRCA1-No Matter How You Splice It.

BRCA1 (breast cancer 1, early onset), a well-known breast cancer susceptibility gene, is a highly alternatively spliced gene. BRCA1 alternative splicing may serve as an alternative regulatory mechanism for the inactivation of the BRCA1 gene in both hereditary and sporadic breast cancers, and other BRCA1-associated cancers. The alternative transcripts of BRCA1 can mimic known functions, possess unique functions compared with the full-length BRCA1 transcript, and in some cases, appear to function in opposition to full-length BRCA1 In this review, we will summarize the functional "naturally occurring" alternative splicing transcripts of BRCA1 and then discuss the latest next-generation sequencing-based detection methods and techniques to detect alternative BRCA1 splicing patterns and their potential use in cancer diagnosis, prognosis, and therapy.

The Potential Role of SnRK1 Protein Kinases in the Regulation of Cell Division in Arabidopsis thaliana

It is known that the sucrose nonfermenting-1 (SNF1)-related protein kinase (SnRK1) subfamily participates in the regulation of carbohydrate metabolism and energy balance. These enzymes are multifunctional and can participate in many other important cellular processes. The role of the SnRK1 protein kinases (KIN10 and KIN11) in the regulation of cell division in Arabidopsis thaliana has been studied in this work. The KIN10 and KIN11 gene knockout lines of A. thaliana were used for this purpose ( http://arabidopsis.info /). A low mitotic index was recorded in cells of these mutant lines and a decreased expression level was shown in the cell proliferation markers—the CYCB1;1 gene (B1 type cyclin) and the plant homolog BRCA1 (Breast Cancer Suppressor Protein). The significantly lower mitotic index and expression level of CYCB1;1 and BRCA1 were observed in the mutants grown in the conditions of energy deficiency. The higher expression of CYCB1;1/BRCA1 and KIN10/KIN11 genes was also recorded in the suspension culture of A. thaliana compared with intact sprouts. These data may confirm a possible role of the KIN10/KIN11 protein kinases in the regulation of cell proliferative activity.

Association of BRCA1- and BRCA2-deficiency with mutation burden, expression of PD-L1/PD-1, immune infiltrates, and T cell-inflamed signature in breast cancer.

Immune checkpoint inhibitors have demonstrated effective anti-tumour response in cancer types with high mutation burden (e.g. melanoma) and in subset of cancers with features of genomic instability (e.g. mismatch-repair deficiency). One possible explanation for this effect is the increased expression of immune checkpoint molecules and pre-existing adaptive immune response in these cancers. Given that BRCA1 and BRCA2 are integral in maintaining genomic integrity, we hypothesise that the inactivation of these genes may give rise to breast cancers with such immunogenic phenotype. Therefore, using two large series of publicly available breast cancer datasets, namely that from The Cancer Genome Atlas and Wellcome Trust Institute, we sought to investigate the association between BRCA1- and BRCA2-deficiency with features of genomic instability, expression of PD-L1 and PD-1, landscape of inferred tumour-infiltrating immune cells, and T-cell inflamed signature in breast cancers. Here, we report that BRCA1 and BRCA2-deficient breast cancers were associated with features of genomic instability including increased mutation burden. Interestingly, BRCA1-, but not BRCA2-, deficient breast cancers were associated with increased expression of PD-L1 and PD-1, higher abundance of tumour-infiltrating immune cells, and enrichment of T cell-inflamed signature. The differences in immunophenotype between BRCA1- and BRCA2-deficient breast cancers can be attributed, in part, to PTEN gene mutation. Therefore, features of genomic instability such as that mediated by BRCA1- and BRCA2- deficiency in breast cancer were necessary, but not always sufficient, for yielding T cell-inflamed tumour microenvironment, and by extension, predicting clinical benefit from immunotherapy.

Therapeutic targeting of BRCA1 and TP53 mutant breast cancer through mutant p53 reactivation

Triple negative breast cancer (TNBC) is an aggressive subset for which effective therapeutic approaches are needed. A significant proportion of TNBC patients harbor either germline or somatic mutations in BRCA1, or epigenetic silencing of BRCA1, which renders them deficient in DNA repair. Virtually all BRCA1 deficient breast cancers harbor mutations in TP53 suggesting that inactivation of p53 is a requirement for tumor progression in the setting of BRCA1 deficiency. Due to this dependency, we hypothesized that restoring wild type p53 function in BRCA1 deficient breast cancer would be therapeutic. The majority of TP53 mutations are missense, which generate a defective protein that potentially can be targeted with small molecules. Zinc metallochaperones (ZMCs) are a new class of anti-cancer drugs that specifically reactivate zinc-deficient mutant p53 by restoring zinc binding. Using ZMC1 in human breast cancer cell lines expressing the zinc deficient p53R175H, we demonstrate that loss of BRCA1 sensitizes cells to mutant p53 reactivation. Using murine breast cancer models with Brca1 deficiency, we demonstrate that ZMC1 significantly improves survival of mice bearing tumors harboring the zinc-deficient Trp53R172H allele but not the Trp53−/− allele. We synthesized a new formulation of ZMC1 (Zn-1), in which the drug is made in complex with zinc to improve zinc delivery, and demonstrate that Zn-1 has increased efficacy. Furthermore, we show that ZMC1 plus olaparib is a highly effective combination for p53R172H tumor growth inhibition. In conclusion, we have validated preclinically a new therapeutic approach for BRCA1 deficient breast cancer through reactivation of mutant p53.

Identification of candidate cancer predisposing variants by performing whole-exome sequencing on index patients from BRCA1 and BRCA2-negative breast cancer families

BackgroundIn the majority of familial breast cancer (BC) families, the etiology of the disease remains unresolved. To identify missing BC heritability resulting from relatively rare variants (minor allele frequency ≤ 1%), we have performed whole exome sequencing followed by variant analysis in a virtual panel of 492 cancer-associated genes on BC patients from BRCA1 and BRCA2 negative families with elevated BC risk.MethodsBC patients from 54 BRCA1 and BRCA2-negative families with elevated BC risk and 120 matched controls were considered for germline DNA whole exome sequencing. Rare variants identified in the exome and in a virtual panel of cancer-associated genes [492 genes associated with different types of (hereditary) cancer] were compared between BC patients and controls. Nonsense, frame-shift indels and splice-site variants (strong protein-damaging variants, called PDAVs later on) observed in BC patients within the genes of the panel, which we estimated to possess the highest probability to predispose to BC, were further validated using an alternative sequencing procedure.ResultsExome- and cancer-associated gene panel-wide variant analysis show that there is no significant difference in the average number of rare variants found in BC patients compared to controls. However, the genes in the cancer-associated gene panel with nonsense variants were more than two-fold over-represented in women with BC and commonly involved in the DNA double-strand break repair process. Approximately 44% (24 of 54) of BC patients harbored 31 PDAVs, of which 11 were novel. These variants were found in genes associated with known or suspected BC predisposition (PALB2, BARD1, CHEK2, RAD51C and FANCA) or in predisposing genes linked to other cancer types but not well-studied in the context of familial BC (EXO1, RECQL4, CCNH, MUS81, TDP1, DCLRE1A, DCLRE1C, PDE11A and RINT1) and genes associated with different hereditary syndromes but not yet clearly associated with familial cancer syndromes (ABCC11, BBS10, CD96, CYP1A1, DHCR7, DNAH11, ESCO2, FLT4, HPS6, MYH8, NME8 and TTC8). Exome-wide, only a few genes appeared to be enriched for PDAVs in the familial BC patients compared to controls.ConclusionsWe have identified a series of novel candidate BC predisposition variants/genes. These variants/genes should be further investigated in larger cohorts/case-control studies. Other studies including co-segregation analyses in affected families, locus-specific loss of heterozygosity and functional studies should shed further light on their relevance for BC risk.

Germline Genetic Testing: What the Breast Surgeon Needs to Know.

The American Society of Breast Surgeons (ASBrS) sought to provide educational guidelines for breast surgeons on how to incorporate genetic information and genomics into their practice. A comprehensive nonsystematic review was performed of selected peer-reviewed literature. The Genetics Working Group of the ASBrS convened to develop guideline recommendations. Clinical and educational guidelines were prepared to outline the essential knowledge for breast surgeons to perform germline genetic testing and to incorporate the findings into their practice, which have been approved by the ASBrS Board of Directors. Thousands of women in the USA would potentially benefit from genetic testing for BRCA1, BRCA2, and other breast cancer genes that markedly increase their risk of developing breast cancer. As genetic testing is now becoming more widely available, women should be made aware of these tests and consider testing. Breast surgeons are well positioned to help facilitate this process. The areas where surgeons need to be knowledgeable include: (1) identification of patients for initial breast cancer-related genetic testing, (2) identification of patients who tested negative in the past but now need updated testing, (3) initial cancer genetic testing, (4) retesting of patients who need their genetic testing updated, (5) cancer genetic test interpretation, posttest counseling and management, (6) management of variants of uncertain significance, (7) cascade genetic testing, (8) interpretation of genetic tests other than clinical cancer panels and the counseling and management required, and (9) interpretation of somatic genetic tests and the counseling and management required.

Identifying Disease Network Dysregulation Through Expression Mean, Variance, and Distribution Changes

Network-based approaches to identifying gene expression signatures of disease have been shown to be more reproducible, accurate, and biologically informative than single-gene markers. Differential expression statistics are most commonly used to rank and select nodes within biological networks. Disease heterogeneity and disease-specific mechanisms may create changes in gene expression distributions that are not captured by differential expression analysis. We implemented a disease network identification algorithm using four unique statistical metrics, based on changes in mean expression, expression variance, and overall expression distribution changes. Using three publicly available gene expression datasets, we evaluated the size, strength, and disease classification performance of the identified subnetworks in cancer and Posttraumatic Stress Disorder (PTSD) cohorts. Identified subnetworks for all three datasets had significant enrichment of disease signal over individual genes, based on ${p}$ -value. Further, novel and unique subnetworks were identified by each statistical metric, including changes in expression variance related to BRCA1 in breast cancer, and changes in blood coagulation in PTSD. Subnetwork-based biomarkers also slightly improved prediction performance over individual gene markers. This approach could provide a novel network-based framework for understanding and predicting gene expression signals of disease.

Plasma RANKL levels are not associated with breast cancer risk in BRCA1 and BRCA2 mutation carriers.

BackgroundAberrant progesterone/receptor activator of nuclear factor κβ (RANK) signaling has been implicated in BRCA1 breast cancer development. Furthermore, lower circulating RANKL has been reported among women with a BRCA mutation compared to non-carriers; however, there have been no reports of plasma RANKL levels and subsequent breast cancer risk. We prospectively evaluated the relationship between plasma RANKL and breast cancer risk among women with a BRCA1 or BRCA2 mutation.MethodsAn enzyme-linked immunosorbent assay was used to quantify plasma RANKL levels in 184 BRCA mutation carriers. Women were stratified into high vs. low RANKL based on the median levels of the cohort (5.24 pg/ml). Kaplan-Meier survival analysis was used to estimate the cumulative incidence of breast cancer by baseline plasma RANKL and cox proportional hazards models were used to estimate the adjusted hazard ratios (HRs) and 95% confidence intervals (CI) for the association between plasma RANKL and risk.ResultsOver a mean follow-up of 6.3 years (0.02-19.24), 15 incident breast cancers were identified. The eight-year cumulative incidence was 10% in the low RANKL group and 12% in the high RANKL group (P-log-rank = 0.85). There was no significant association between plasma RANKL levels and breast cancer risk (multivariate HR high vs. low = 1.06; 95%CI 0.34-3.28; P-trend = 0.86).ConclusionsThese findings suggest that circulating RANKL levels are not associated with breast cancer among BRCA mutation carriers. Pending validation in a larger sample, these findings suggest that RANKL is likely not a biomarker of breast cancer risk among BRCA mutation carriers.