Using next‐generation sequencing to redefine BRCAness in triple‐negative breast cancer

BRCAness is considered a predictive biomarker to platinum and poly(ADP‐ribose) polymerase (PARP) inhibitors. However, recent trials showed that its predictive value was limited in triple‐negative breast cancer (TNBC) treated with platinum. Moreover, tumors with mutations of DNA damage response (DDR) genes, such as homologous recombination (HR) genes, could be sensitive to platinum and PARP inhibitors. Thus, we aim to explore the relationship between mutation status of DDR genes and BRCAness in TNBC. We sequenced 56 DDR genes in 120 TNBC and identified BRCAness by array comparative genomic hybridization. The sequencing results showed that 13, 14, and 14 patients had BRCA, non‐BRCA HR, and non‐HR DDR gene mutations, respectively. Array comparative genomic hybridization revealed that BRCA‐mutated and HR gene‐mutated TNBC shared similar BRCAness features, both having higher numbers and longer length of large‐scale structural aberration (LSA, >10 Mb) and similar altered chromosomal regions of LSA. These suggested non‐BRCA HR gene‐mutated TNBC shared similar characteristics with BRCA‐mutated TNBC, indicating non‐BRCA HR gene‐mutated TNBC sensitive to platinum and PARP inhibitors. Among tumors with mutation of non‐HR DDR genes, 3 PTEN and 1 MSH6 mutation also contained significant LSAs (BRCAness); however, they had different regions of genomic alteration to BRCA and HR gene‐mutated tumors, might explain prior findings that PTEN‐ and MSH6‐mutated cancer cells not sensitive to PARP inhibitors. Therefore, we hypothesize that the heterogeneous genomic background of BRCAness indicates different responsiveness to platinum and PARP inhibitors. Direct sequencing DDR genes in TNBC should be applied to predict their sensitivity toward platinum and PARP inhibitors.     

Treatment outcomes and clinicopathologic characteristics of triple‐negative breast cancer patients who received platinum‐containing chemotherapy

The aim of this study was to evaluate the role of platinum‐containing chemotherapy for metastatic triple‐negative breast cancer (TNBC) patients in terms of the response rate (RR) and progression‐free survival. A second aim was to characterize the clinical behavior at the time of relapse of TNBC. We retrospectively analyzed the clinical outcomes of patients with metastatic breast cancer who received taxane–platinum chemotherapy as the first‐ or second‐line treatment, focusing on the TN phenotype. In total, 257 patients with metastatic breast cancer received platinum‐containing chemotherapy at Samsung Medical Center from 1999 to 2006. Of these patients, 106 patients with available data on estrogen (ER), progesterone (PgR) and human epidermal growth factor receptor‐2 (HER2) receptor status received taxane–platinum regimen as the first‐ or second‐line treatment. The overall RR of patients with TNBC was 39%. This rate did not differ significantly from those of patients with other phenotypes. The time to death after chemotherapy (19 vs. 50 months, p = 0.037) and overall survival (OS) (21 vs. 56 months, p = 0.030) differed significantly between patients with TNBC and non‐TNBC. TNBC showed a unique locoregional infiltration pattern at relapse, which might reflect its aggressive clinical behavior. Despite the similar response to platinum‐containing chemotherapy, patients with TNBC had a shorter OS than patients with non‐TNBC. The implication of TN phenotype as poor prognostic factor is uncertain, because it needs to be defined whether poor outcome is related to the rapid growing characteristics of tumor itself or the resistance to drug therapy. Further prospective studies are warranted. © 2008 Wiley‐Liss, Inc.     

Triple-negative breast cancer: BRCAness and concordance of clinical features with BRCA1-mutation carriers

Background:

BRCAness is defined as shared tumour characteristics between sporadic and BRCA-mutated cancers. However, how to exactly measure BRCAness and its frequency in breast cancer is not known. Assays to establish BRCAness would be extremely valuable for the clinical management of these tumours. We assessed BRCAness characteristics frequencies in a large cohort of triple-negative breast cancers (TNBCs).

Methods:

As a measure of BRCAness, we determined a specific BRCA1-like pattern by array Comparative Genomic Hybridisation (aCGH), and BRCA1 promoter methylation in 377 TNBCs, obtained from 3 different patient cohorts. Clinicopathological data were available for all tumours, BRCA1-germline mutation status and chemotherapy response data were available for a subset.

Results:

Of the tumours, 66–69% had a BRCA1-like aCGH profile and 27–37% showed BRCA1 promoter methylation. BRCA1-germline mutations and BRCA1 promoter methylation were mutually exclusive events (P=1 × 10⁻⁵). BRCAness was associated with younger age and grade 3 tumours. Chemotherapy response was significantly higher in BRCA1-mutated tumours, but not in tumours with BRCAness (63% (12 out of 19) vs 35% (18 out of 52) pathological complete remission rate, respectively).

Conclusion:

The majority of the TNBCs show BRCAness, and those tumours share clinicopathological characteristics with BRCA1-mutated tumours. A better characterisation of TNBC and the presence of BRCAness could have consequences for both hereditary breast cancer screening and the treatment of these tumours.     

Subtyping of triple‐negative breast cancer: Implications for therapy

Triple‐negative breast cancer (TNBC) is a heterogeneous disease; gene expression analyses recently identified 6 distinct TNBC subtypes, each of which displays a unique biology. Exploring novel approaches for the treatment of these subtypes is critical, especially because the median survival for women with metastatic TNBC is less than 12 months, and virtually all women with metastatic TNBC ultimately will die of their disease despite systemic therapy. To date, not a single targeted therapy has been approved for the treatment of TNBC, and cytotoxic chemotherapy remains the standard treatment. In this review, the authors discuss recent developments in subtyping TNBC and the current and upcoming therapeutic strategies being explored in an attempt to target TNBC. Cancer 2015;121:8–16 . © 2014 American Cancer Society.     

BRCAness: finding the Achilles heel in ovarian cancer

Ovarian cancer is the leading cause of death among gynecological cancers. It exhibits great heterogeneity in tumor biology and treatment response. Germline mutations of DNA repair genes BRCA1/2 are the fundamental defects in hereditary ovarian cancer that expresses a distinct phenotype of high response rates to platinum agents, improved disease-free intervals and survival rates, and high-grade serous histology. The term "BRCAness" describes the phenotypic traits that some sporadic ovarian tumors share with tumors in BRCA1/2 germline mutation carriers and reflects similar causative molecular abnormalities. BRCA pathway studies and molecular profiling reveal BRCA-related defects in almost half of the cases of ovarian cancer. BRCA-like tumors are particularly sensitive to DNA-damaging agents (e.g., platinum agents) because of inadequate BRCA-mediated DNA repair mechanisms, such as nucleotide-excision repair and homologous recombination (HR). Additional inhibition of other DNA repair pathways leads to synthetic lethality in HR-deficient cells; this has been employed in the treatment of BRCA-like ovarian tumors with poly(ADP-ribose) polymerase inhibitors with promising results. This article presents a comprehensive review of the relevant literature on the role of BRCAness in ovarian cancer with respect to BRCA function, methods of BRCA epigenetic defect detection and molecular profiling, and the implications of BRCA dysfunction in the treatment of ovarian cancer.     

Low prevalence of germline PALB2 mutations in Australian triple‐negative breast cancer

Triple‐negative breast cancer (TNBC) is a tumour classification that is defined by oestrogen receptor, progesterone receptor and human epidermal growth factor receptor 2 receptor negativity. TNBCs share a similar gene expression profile to BRCA‐mutated tumours, have been shown to carry a high proportion of BRCA mutations and have a more adverse prognosis compared to other types of breast tumours. PALB2 has been shown to be a moderate‐penetrance breast cancer susceptibility gene and is involved in the same DNA damage repair pathway as BRCA1 and BRCA2; this raises the possibility that germline PALB2 mutations may be involved in the pathogenesis of TNBCs. In our study, we sequenced the coding regions of PALB2 (including intron/exon boundaries) in genomic DNA from 347 patients diagnosed with TNBC to determine the prevalence of deleterious mutations in this population. Two novel truncating mutations (c.758dup and c.2390del) and one previously detected truncating mutation (c.3113+5G>C) were found. In addition, five variants predicted to be protein‐affecting were also identified. Our study shows that the prevalence of PALB2 germline mutations in individuals with TNBC is ～1%, similar to the prevalence of PALB2 germline mutation of 1% in familial non‐BRCA1/2 breast cancer cohorts.     

Predictive value of BRCA1/2 mRNA expression for response to neoadjuvant chemotherapy in BRCA‐negative breast cancers

It is well known that BRCA1 and BRCA2 play a central role in DNA repair, but the relationship between BRCA1 and BRCA2 mRNA expression and response to neoadjuvant chemotherapy in sporadic breast cancer patients has not been well established. Here, we investigate the association between BRCA1 or BRCA2 mRNA expression levels and pathological response in 674 BRCA1/2 mutation‐negative breast cancer patients who received neoadjuvant chemotherapy. BRCA1 and BRCA2 mRNA expression were assessed using quantitative real‐time polymerase chain reaction in core biopsy breast cancer tissue obtained prior to the initiation of neoadjuvant chemotherapy. A total 129 patients (19.1%) achieved pathological complete response (pCR) after neoadjuvant chemotherapy. Among patients treated with anthracycline‐based chemotherapy (n = 531), BRCA1 mRNA low expression patients had a significantly higher pCR rate than intermediate or high BRCA1 mRNA expression groups (24.6% vs 16.8% or 14.0%, P = .031) and retained borderline significance (OR = 1.54, 95% CI = 0.93‐2.56, P = .094) in multivariate analysis. Among the 129 patients who received a taxane‐based regimen, pCR rate showed no differences in BRCA1 low, intermediate, and high mRNA level subgroups (19.6%, 26.8% and 21.4%, respectively; P = .71). BRCA2 mRNA level was not associated with pCR rate in the anthracyline‐based treated subgroup (P = .60) or the taxane‐based regimen subgroup (P = .82). Taken together, our findings suggested that BRCA1 mRNA expression could be used as a predictive marker in BRCA1/2 mutation‐negative breast cancer patients who received neoadjuvant anthracycline‐based treatment.     

Alcohol,smoking, and risk of Her2‐overexpressing and triple‐negative breast cancer relative to estrogen receptor‐positive breast cancer

Epidemiological evidence is limited on how alcohol consumption and smoking are associated with risk of different subtypes of breast cancer, such as triple‐negative (TN) and human epidermal growth factor receptor 2‐overexpressing (H2E) breast cancers, which may have different etiologies from more common luminal (estrogen receptor [ER+]) breast cancers. In this population‐based case‐case study, we evaluated the association between alcohol, smoking, and risk of H2E and TN breast cancer, compared with ER+ breast cancers, among women aged 20–69 years. Using polytomous regression, associations between alcohol consumption, smoking, and breast cancer risk were evaluated in 909 ER+, 1,290 TN, and 489 H2E breast cancer patients, with ER+ breast cancer patients as the reference group. Current alcohol consumption at diagnosis was associated with a lower risk of H2E breast cancer (odds ratio = 0.74, 95% confidence interval: 0.58–0.92) relative to ER+ cancers. No difference in association was observed by menopausal status. No association between alcohol consumption and TN breast cancer relative to ER+ breast cancer was observed. Women who smoked did not have an altered risk of TN or H2E breast cancer, relative to ER+ cancer. Our results suggest that alcohol is associated with lower risk of H2E breast cancer relative to ER+ breast cancer. This study adds to the body of epidemiologic evidence that breast cancer etiology differs by breast cancer subtype.     

Clinical outcome of triple negative breast cancer in BRCA1 mutation carriers and noncarriers

BACKGROUND:

Women with BRCA1 mutations develop breast cancer with similar pathologic features to sporadic triple negative (TN) breast cancer, a subtype associated with early disease relapse and poor outcome. The clinical outcome of women with and without BRCA1 mutations who had TN breast cancer treated with conventional chemotherapy were compared.

METHODS:

Women with stage I to III TN breast cancer who had BRCA1 testing within 36 months of diagnosis and received alkylating chemotherapy were identified from clinical databases and a Specialized Program of Research Excellence (SPORE) specimen bank. BRCA2 mutation carriers were excluded, resulting in a study cohort of 46 BRCA1 carriers and 71 noncarriers. Sites of metastasis, relapse rates, and survival were compared among carriers and noncarriers. The median follow‐up was 75 months.

RESULTS:

BRCA1 carriers were younger at diagnosis (P < .001) and had smaller tumors (P = .03) than noncarriers. Freedom from distant metastasis at 5 years was 76% for carriers and 70% for noncarriers (hazard ratio [HR] 0.79, P = .5). Sites of distant recurrence did not differ significantly (P = .15), although BRCA1 carriers had a propensity for brain relapse (58% vs 24%, P = .06). Overall survival at 5 years was 82% for carriers and 74% for noncarriers (HR 0.64, P = .25). Adjusting for age and stage, BRCA1 mutation status was not an independent predictor of survival (HR 0.73, P = .48).

CONCLUSIONS:

BRCA1 mutation carriers with TN disease had similar survival rates to noncarriers when treated with alkylating chemotherapy. Women with BRCA1‐related breast cancer may benefit from novel therapies that target DNA repair, and further study is needed to identify sporadic TN breast cancers with a BRCA‐deficient phenotype. Cancer 2011. © 2011 American Cancer Society.     

Role of inhibitor of yes‐associated protein 1 in triple‐negative breast cancer with taxol‐based chemoresistance

Triple‐negative breast cancer (TNBC) is highly clinically aggressive and taxol‐based chemoresistance remains a big TNBC therapeutic problem to be solved. Verteporfin, a small molecular yes‐associated protein 1 (YAP1) inhibitor, is little known as an antitumor drug for TNBC. Our data showed that YAP1 expression was associated with early relapse in tissue samples of patients with TNBC taxol chemoresistance (P < .001). Verteporfin reduced migration and enhanced apoptosis or autophagy of a taxol‐resistant MDA‐MB‐231 cell line in vitro. Knockdown of YAP1 increased epithelial‐mesenchymal transition response in a taxol‐resistant TNBC cell line. In an in vivo experiment, we found that verteporfin was able to shrink tumor weight and volume and decreased Ki67 expression in a taxol‐resistant mouse model. Our results provide evidence that verteporfin could be a chemosensitizer for TNBC patients with taxol‐based treatment.     

BRCA1和MDR1在三阴性乳腺癌中的表达及其临床意义

目的:观察BRCA1和MDR1在三阴性乳腺癌与非三阴性乳腺癌中的表达,研究药物敏感性,分析对临床预后判断的意义.方法:采用免疫组化法(SP二步法)检测手术切除的45例三阴性乳腺癌组织和30例非三阴性乳腺癌组织中BRCA1和MDR1的表达情况,并采用X2检验方法,分析BRCA1、MDR1与三阴性乳腺癌临床病理特征的关系及药物敏感性研究.结果:在临床病理特征方面,三阴性乳腺癌患者中绝经前所占比例为80％,高于非三阴性乳腺癌的43.3％,差异有统计学意义(P＜0.05);组织学Ⅱ级以上者三阴性乳腺癌组为71.1％,非三阴性乳腺癌组为33.3％,两者之间差异有统计学意义(P＜0.05);而在肿瘤大小、淋巴结是否转移、临床分期上两组相比差异无统计学意义(P＞0.05).三阴性乳腺癌患者的BRCA1低表达,应用含铂类方案化疗患者生存期长(P＜0.05).结论:相对于非三阴性乳腺癌,三阴性乳腺癌患者具有在绝经前发病率高、组织分级高等高危特点;BRCA1和MDR1高表达与其临床预后及对药物敏感度呈负相关,提示在临床预防、治疗和预后中可能有一定的指导意义.     

MicroRNA expression signatures for the prediction of BRCA1/2 mutation‐associated hereditary breast cancer in paraffin‐embedded formalin‐fixed breast tumors

Screening for germline mutations in breast cancer‐associated genes BRCA1 and BRCA2 is indicated for patients with breast cancer from high‐risk breast cancer families and influences both treatment options and clinical management. However, only 25% of selected patients test positive for BRCA1/2 mutation, indicating that additional diagnostic biomarkers are necessary. We analyzed 124 formalin‐fixed paraffin‐embedded (FFPE) tumor samples from patients with hereditary (104) and sporadic (20) invasive breast cancer, divided into two series (A and B). Microarray expression profiling of 829 human miRNAs was performed on 76 samples (Series A), and bioinformatics tool Prophet was used to develop and test a microarray classifier. Samples were stratified into a training set (n = 38) for microarray classifier generation and a test set (n = 38) for signature validation. A 35‐miRNA microarray classifier was generated for the prediction of BRCA1/2 mutation status with a reported 95% (95% CI = 0.88–1.0) and 92% (95% CI: 0.84–1.0) accuracy in the training and the test set, respectively. Differential expression of 12 miRNAs between BRCA1/2 mutation carriers versus noncarriers was validated by qPCR in an independent tumor series B (n = 48). Logistic regression model based on the expression of six miRNAs (miR‐142‐3p, miR‐505*, miR‐1248, miR‐181a‐2*, miR‐25* and miR‐340*) discriminated between tumors from BRCA1/2 mutation carriers and noncarriers with 92% (95% CI: 0.84–0.99) accuracy. In conclusion, we identified miRNA expression signatures predictive of BRCA1/2 mutation status in routinely available FFPE breast tumor samples, which may be useful to complement current patient selection criteria for gene testing by identifying individuals with high likelihood of being BRCA1/2 mutation carriers.     

聚腺苷酸二磷酸核糖转移酶-1抑制剂在三阴性乳腺癌中的应用现状

目的 对国内外聚腺苷酸二磷酸核糖转移酶-1[Poly (ADP-ribose) polymerase-1,PARP1]抑制剂在三阴性乳腺癌中的应用现状进行综述分析.方法 应用PubMed及CNKI期刊全文数据库检索系统,以“聚腺苷酸二磷酸核糖转移酶-1、PARP1抑制剂、三阴性乳腺癌”等为关键词,检索2009-01-2014-02相关文献,共检索到英文文献322条,中文文献201条.纳入标准:1)PARP1分子的生物学功能;2)PARP1抑制剂在BRCA突变相关乳腺癌以及三阴性乳腺癌方面的基础与临床研究;剔除标准:1)PARP1抑制剂的药理学特性;2)PARP1与乳腺癌发生相关性研究.最后纳入分析33篇文献.结果 “合成致死”原理是PARP1抑制剂在BRCA突变相关乳腺癌中发挥抗肿瘤活性的理论基础,PARP1抑制剂治疗BRCA突变相关的乳腺癌的Ⅰ、Ⅱ临床试验取得良好的成果,但是在三阴性乳腺癌的治疗方面,Ⅲ期临床试验并未得到预期的试验结果,在应用PARP1抑制剂之前重组修复通路的功能状态以及PARP1的活性应该得到合理评估.结论 三阴性乳腺癌与BRCA突变相关乳腺癌存在表型的相似性,但是三阴性乳腺癌患者是否可以受益于PARP1抑制剂的治疗,还需要进一步深入研究,同时寻找可靠生物标记分子预测PARP1抑制剂治疗的敏感性,是目前PARP1抑制剂应用于临床所面临的挑战.     

HB‐EGF orchestrates the complex signals involved in triple‐negative and trastuzumab‐resistant breast cancer

A number of therapeutic strategies targeting epidermal growth factor receptor (EGFR) have not always led to success in the present state of breast cancer therapy. Notably, there is currently no way to treat trastuzumab‐resistant and triple‐negative breast cancer (TNBC). Here, we found that heparin‐binding epidermal growth factor‐like growth factor (HB‐EGF), a member of the EGFR ligands, was predominantly expressed in breast cancer and that treatment with crossreacting material 197 (CRM197), a specific inhibitor of HB‐EGF, blocked ERK as well as AKT activation via complexes of EGFR and unknown growth factor receptors in TNBC or through complexes of EGFR and human epidermal growth factor receptor‐2 in trastuzumab‐resistant breast cancer, caused significant cell apoptosis and inhibited tumor growth. Accordingly, we can provide a novel concept that a certain EGFR ligand is recognized as a rational target against breast cancer. In addition, it is plausible that CRM197 could be an effective anticancer agent for molecularly targeted therapies.     

MicroRNA‐493 is a prognostic factor in triple‐negative breast cancer

Breast cancer is one of the most common malignant diseases in women. Triple‐negative breast cancer (TNBC) shows higher aggressiveness and recurrence rates than other subtypes, and there are no effective targets or tailored treatments for TNBC patients. Thus, finding effective prognostic markers for TNBC could help clinicians in their ability to care for their patients. We used tissue microarrays (TMAs) to detect microRNA‐493 (miR‐493) expression in breast cancer samples. A miRCURY LNA detection probe specific for miR‐493 was used in in situ hybridization assays. Staining results were reviewed by two independent pathologists and classified as high or low expression of miR‐493. Kaplan–Meier survival plots and multivariate Cox analysis were carried out to clarify the relationship between miR‐493 and survival. In the Kaplan–Meier analysis, patients with high miR‐493 expression had better disease‐free survival than patients with low miR‐493 expression. After adjusting for common clinicopathological factors in breast cancer, the expression level of miR‐493 was still a significant prognostic factor in breast cancer. Further subtype analysis revealed that miR‐493 expression levels were only significantly prognostic in TNBC patients. These results were validated in the Molecular Taxonomy of Breast Cancer International Consortium database for overall survival. We proved the prognostic role of miR‐493 in TNBC by using one of the largest breast cancer TMAs available and validated it in a large public RNA sequencing database.     

BRCA1 andBRCA2 germline mutations in Japanese with hereditary breast cancer families

We examined germline mutations inBRCA1 andBRCA2 in 23 Japanese breast cancer families, using PCR-SSCP analysis. The same nonsense mutation (exon 5, Leu63ter) ofBRCA1 was detected in two different families. Three different mutations resulting in a truncatedBRCA2 protein (exon 9, 20, 24) were detected in three different families, including one male case of breast cancer. One base substitution mutation inBRCA2, A10462G, was detected in the other two families. Although the mean age of onset for breast cancer in families with theBRCA1- mutation was 50 years, the age of onset in families with theBRCA2-mutation was from 28 to 43 years. Among the 23 families examined, two families had members with ovarian cancers, three had members with prostate cancers, and one had a pancreatic cancer. However, none of these families was positive for theBRCA1 orBRCA2 mutation. Histopathologically, we observed a prevalence of histological grade 3 inBRCA2-associated familial breast cancers, because of nuclear atypia, structural atypia and mitotic activity. It is suggested thatBRCA2 may play a more important role thanBRCA1 in Japanese familial breast cancers, and these mutations are related to the aggressive nature and highly proliferative activity of the tumors.     

miRNA expression patterns in normal breast tissue and invasive breast cancers of BRCA1 and BRCA2 germ-line mutation carriers

miRNA deregulation has been found to promote carcinogenesis. Little is known about miRNA deregulation in hereditary breast tumors as no miRNA expression profiling studies have been performed in normal breast tissue of BRCA1 and BRCA2 mutation carriers. miRNA profiles of 17 BRCA1- and 9 BRCA2-associated breast carcinomas were analyzed using microarrays. Normal breast tissues from BRCA1 and BRCA2 mutation carriers (both n = 5) and non-mutation carriers (n = 10) were also included. Candidate miRNAs were validated by qRT-PCR. Breast carcinomas showed extensive miRNA alteration compared to normal breast tissues in BRCA1 and BRCA2 mutation carriers. Moreover, normal breast tissue from BRCA1 mutation carriers already showed miRNA alterations compared to non-mutation carriers. Chromosomal distribution analysis showed several hotspots containing down- or up-regulated miRNAs. Pathway analysis yielded many similarities between the BRCA1 and BRCA2 axes with miRNAs involved in cell cycle regulation, proliferation and apoptosis. Lesser known pathways were also affected, including cellular movement and protein trafficking. This study provides a comprehensive insight into the potential role of miRNA deregulation in BRCA1/2-associated breast carcinogenesis. The observed extensive miRNA deregulation is likely the result of genome-wide effects of chromosomal instability caused by impaired BRCA1 or BRCA2 function. This study''s results also suggest the existence of common pathways driving breast carcinogenesis in both BRCA1 and BRCA2 germ-line mutation carriers.