The Retinoblastoma Gene Undergoes Rearrangements in BRCA1-Deficient Basal-like Breast Cancer

Breast tumors from BRCA1 germ line mutation carriers typically exhibit features of the basal-like molecular subtype. However, the specific genes recurrently mutated as a consequence of BRCA1 dysfunction have not been fully elucidated. In this study, we used gene expression profiling to molecularly subtype 577 breast tumors, including 73 breast tumors from BRCA1/2 mutation carriers. Focusing on the RB1 locus, we analyzed 33 BRCA1-mutated, 36 BRCA2-mutated, and 48 non-BRCA1/2-mutated breast tumors using a custom-designed high-density oligomicroarray covering the RB1 gene. We found a strong association between the basal-like subtype and BRCA1-mutated breast tumors and the luminal B subtype and BRCA2-mutated breast tumors. RB1 was identified as a major target for genomic disruption in tumors arising in BRCA1 mutation carriers and in sporadic tumors with BRCA1 promoter methylation but rarely in other breast cancers. Homozygous deletions, intragenic breaks, or microdeletions were found in 33% of BRCA1-mutant tumors, 36% of BRCA1 promoter-methylated basal-like tumors, 13% of non-BRCA1-deficient basal-like tumors, and 3% of BRCA2-mutated tumors. In conclusion, RB1 was frequently inactivated by gross gene disruption in BRCA1 hereditary breast cancer and BRCA1-methylated sporadic basal-like breast cancer but rarely in BRCA2 hereditary breast cancer and non-BRCA1-deficient sporadic breast cancers. Together, our findings show the existence of genetic heterogeneity within the basal-like breast cancer subtype that is based upon BRCA1 status. Cancer Res; 72(16); 4028-36. ?2012 AACR.     

An integrative hypothesis about the origin and development of sporadic and familial breast cancer subtypes

Do breast cancer tumours have a common cell origin? Do different breast cancer molecular phenotypes arise from distinct cell types? The studies we have performed during the last few years in familial breast tumours (BRCA1, BRCA2 and non-BRCA1/2) widen questions about the development of sporadic breast cancer to hereditary breast cancer. Array-comparative genomic hybridisation (CGH) studies show universal genomic aberrations in both familial and sporadic breast cancer subtypes that may be selected in the breast tumour development. The inactivation of BRCA1 seems to play a critical role in oestrogen receptor (ER)-negative cancer stem cells (CSCs), driving the tumour development mostly towards a basal-like or, in some cases, to a luminal B phenotype, but other carcinogenetic events are proposed to explain the remaining tumour subtypes. The existence of common genomic alterations in basal-like, ERBB2 and luminal B breast tumours may suggest a common cell origin or clonal selection of these tumour subtypes, arising from an ER-negative CSC or from a progenitor cell (PC). Finally, specific genomic aberrations in ER-positive tumours could provide cellular proliferation advantages when the cells are exposed to oestrogen. We propose a combination of the CSC hypothesis (for the carcinogenesis processes) and the clonal selection model (in terms of tumour development). We uphold that the basal-like-, ERBB2- and luminal B-sporadic and familial tumour subtypes have an ER-negative breast stem/PC origin, whereas luminal A tumours arise from an ER-positive PC, supporting a hierarchical breast carcinogenesis model, whereas crucial genomic imbalances are clonally selected during the tumour development.     

Subtypes of familial breast tumours revealed by expression and copy number profiling

Extensive expression profiling studies have shown that sporadic breast cancer is composed of five clinically relevant molecular subtypes. However, although BRCA1-related tumours are known to be predominantly basal-like, there are few published data on other classes of familial breast tumours. We analysed a cohort of 75 BRCA1, BRCA2 and non-BRCA1/2 breast tumours by gene expression profiling and found that 74% BRCA1 tumours were basal-like, 73% of BRCA2 tumours were luminal A or B, and 52% non-BRCA1/2 tumours were luminal A. Thirty-four tumours were also analysed by single nucleotide polymorphism–comparative genomic hybridization (SNP-CGH) arrays. Copy number data could predict whether a tumour was basal-like or luminal with high accuracy, but could not predict its mutation class. Basal-like BRCA1 and basal-like non-BRCA1 tumours were very similar, and contained the highest number of chromosome aberrations. We identified regions of frequent gain containing potential driver genes in the basal (8q and 12p) and luminal A tumours (1q and 17q). Regions of homozygous loss associated with decreased expression of potential tumour suppressor genes were also detected, including in basal tumours (5q and 9p), and basal and luminal tumours (10q). This study highlights the heterogeneity of familial tumours and the clinical consequences for treatment and prognosis.     

Familial breast cancers without mutations in BRCA1 or BRCA2 have low cyclin E and high cyclin D1 in contrast to cancers in BRCA mutation carriers.

PURPOSE: We analyzed the expression of critical cell cycle regulators cyclin E and cyclin D1 in familial breast cancer, focusing on BRCA mutation-negative tumors. Cyclin E expression in tumors of BRCA1 or BRCA2 carriers is higher, and cyclin D1 expression lower, than in sporadic tumors. In familial non-BRCA1/2 tumors, cyclin E and cyclin D1 expression has not been studied. EXPERIMENTAL DESIGN: Cyclin E and cyclin D1 immunohistochemical expression was studied in tissue microarrays consisting of 53 BRCA1, 58 BRCA2, 798 familial non-BRCA1/2, and 439 sporadic breast tumors. RESULTS: In univariate analysis, BRCA1 tumors had significantly more frequently high cyclin E (88%) and low cyclin D1 (84%) expression than sporadic (54% and 49%, respectively) or familial non-BRCA1/2 (38% and 45%, respectively) tumors. BRCA2 tumors had significantly more frequently low cyclin D1 expression (68%) than sporadic or familial non-BRCA1/2 tumors and significantly more frequently high cyclin E expression than familial non-BRCA1/2 tumors. In a logistic regression model, cyclin expression, early age of onset, and estrogen receptor (ER) and human epidermal growth factor receptor-2 (HER2) status were the independent factors most clearly distinguishing tumors of BRCA1 mutation carriers from other familial breast cancers. High cyclin E and low cyclin D1 expression were also independent predictors of BRCA2 mutation when compared with familial non-BRCA1/2 tumors. Most interestingly, lower frequency of high cyclin E expression independently distinguished familial non-BRCA1/2 tumors also from sporadic ones. CONCLUSIONS: Cyclin E and cyclin D1 expression distinguishes non-BRCA1/2 tumors from both sporadic and BRCA1- and BRCA2-associated tumors and may reflect different predisposition and pathogenesis in these groups.     

Immunohistochemical Analysis of ER,PR, Her2 and CK5/6 in Infiltrative Breast Carcinomas in Indian Patients

Aim: Breast cancer is biologically a heterogeneous disease. Patients with the same diagnostic profile havemarkedly different clinical outcomes. Gene expression studies identified distinct breast cancer subtypes thatdiffer in prognosis. Aim is to identify the immunohistochemical subtypes of breast carcinoma and correlate theresults with pathological features associated with adverse prognosis in our study population. Method: We included107 consecutive cases of invasive breast carcinoma and sub classified using immunohistochemical staining forER, PR, Her2, and CK5/6 into the following subtypes: luminal A, luminal B, basal-like, Her2+ and unclassified.Associations between tumor subtypes and tumor characteristics were examined. Results: The proportion ofeach subtype in our patient population was: luminal A 37.4%, luminal B 11.1%, Her2+ 29% and basal-like7.5%. The following variables were significantly associated with IHC breast cancer subtypes: patient age (p<.05),overall histopathology grade (p<0.001), nuclear grade (p<0.005) and mitotic index (p<0.001). Her2+ and basallike subtypes were associated with poor differentiation (p<0.01), higher nuclear grade (p<0.05) and high mitoticindex (p<0.05). Conclusions: Our data show a higher proportion of patients in the study population undergototal mastectomy and harbor poorly differentiated, node positive tumors than reported. There was also a relativelyhigh percentage of the Her2+ subtype (29%).     

Genomic subtypes of breast cancer identified by array-comparative genomic hybridization display distinct molecular and clinical characteristics

Introduction

Breast cancer is a profoundly heterogeneous disease with respect to biologic and clinical behavior. Gene-expression profiling has been used to dissect this complexity and to stratify tumors into intrinsic gene-expression subtypes, associated with distinct biology, patient outcome, and genomic alterations. Additionally, breast tumors occurring in individuals with germline BRCA1 or BRCA2 mutations typically fall into distinct subtypes.

Methods

We applied global DNA copy number and gene-expression profiling in 359 breast tumors. All tumors were classified according to intrinsic gene-expression subtypes and included cases from genetically predisposed women. The Genomic Identification of Significant Targets in Cancer (GISTIC) algorithm was used to identify significant DNA copy-number aberrations and genomic subgroups of breast cancer.

Results

We identified 31 genomic regions that were highly amplified in > 1% of the 359 breast tumors. Several amplicons were found to co-occur, the 8p12 and 11q13.3 regions being the most frequent combination besides amplicons on the same chromosomal arm. Unsupervised hierarchical clustering with 133 significant GISTIC regions revealed six genomic subtypes, termed 17q12, basal-complex, luminal-simple, luminal-complex, amplifier, and mixed subtypes. Four of them had striking similarity to intrinsic gene-expression subtypes and showed associations to conventional tumor biomarkers and clinical outcome. However, luminal A-classified tumors were distributed in two main genomic subtypes, luminal-simple and luminal-complex, the former group having a better prognosis, whereas the latter group included also luminal B and the majority of BRCA2-mutated tumors. The basal-complex subtype displayed extensive genomic homogeneity and harbored the majority of BRCA1-mutated tumors. The 17q12 subtype comprised mostly HER2-amplified and HER2-enriched subtype tumors and had the worst prognosis. The amplifier and mixed subtypes contained tumors from all gene-expression subtypes, the former being enriched for 8p12-amplified cases, whereas the mixed subtype included many tumors with predominantly DNA copy-number losses and poor prognosis.

Conclusions

Global DNA copy-number analysis integrated with gene-expression data can be used to dissect the complexity of breast cancer. This revealed six genomic subtypes with different clinical behavior and a striking concordance to the intrinsic subtypes. These genomic subtypes may prove useful for understanding the mechanisms of tumor development and for prognostic and treatment prediction purposes.     

Association between mammographic density and basal-like and luminal A breast cancer subtypes

Introduction

Mammographic density is a strong risk factor for breast cancer overall, but few studies have examined the association between mammographic density and specific subtypes of breast cancer, especially aggressive basal-like breast cancers. Because basal-like breast cancers are less frequently screen-detected, it is important to understand how mammographic density relates to risk of basal-like breast cancer.

Methods

We estimated associations between mammographic density and breast cancer risk according to breast cancer subtype. Cases and controls were participants in the Carolina Breast Cancer Study (CBCS) who also had mammograms recorded in the Carolina Mammography Registry (CMR). A total of 491 cases had mammograms within five years prior to and one year after diagnosis and 528 controls had screening or diagnostic mammograms close to the dates of selection into CBCS. Mammographic density was reported to the CMR using Breast Imaging Reporting and Data System categories. The expression of estrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor 1 and 2 (HER1 and HER2), and cytokeratin 5/6 (CK5/6) were assessed by immunohistochemistry and dichotomized as positive or negative, with ER+ and/or PR+, and HER2- tumors classified as luminal A and ER-, PR-, HER2-, HER1+ and/or CK5/6+ tumors classified as basal-like breast cancer. Triple negative tumors were defined as negative for ER, PR and HER2. Of the 491 cases 175 were missing information on subtypes; the remaining cases included 181 luminal A, 17 luminal B, 48 basal-like, 29 ER-/PR-/HER2+, and 41 unclassified subtypes. Odds ratios comparing each subtype to all controls and case-case odds ratios comparing mammographic density distributions in basal-like to luminal A breast cancers were estimated using logistic regression.

Results

Mammographic density was associated with increased risk of both luminal A and basal-like breast cancers, although estimates were imprecise. The magnitude of the odds ratio associated with mammographic density was not substantially different between basal-like and luminal A cancers in case–control analyses and case-case analyses (case-case OR = 1.08 (95% confidence interval: 0.30, 3.84)).

Conclusions

These results suggest that risk estimates associated with mammographic density are not distinct for separate breast cancer subtypes (basal-like/triple negative vs. luminal A breast cancers). Studies with a larger number of basal-like breast cancers are needed to confirm our findings.     

Immunohistochemical expression of DNA repair proteins in familial breast cancer differentiate BRCA2-associated tumors.

PURPOSE: Morphologic and immunohistochemical studies of familial breast cancers have identified specific characteristics associated with BRCA1 mutation-associated tumors when compared with BRCA2 and non-BRCA1/2 tumors, but have not identified differences between BRCA2 and non-BRCA1/2 tumors. Because BRCA1 and BRCA2 genes participate in the DNA repair pathway, we have performed an immunohistochemical study with markers related to this pathway to establish the profile of the three groups. MATERIALS AND METHODS: We have studied two tissue microarrays that include 103 familial and 104 sporadic breast tumors, with a panel of DNA repair markers including ATM, CHEK2, RAD51, RAD50, XRCC3, and proliferating cell nuclear antigen. RESULTS: We found more frequent expression of CHEK2 in BRCA1 and BRCA2 tumors than in non-BRCA1/2 and sporadic tumors. We found absence of nuclear expression and presence of cytoplasmic expression of RAD51 in BRCA2 tumors that differentiate them from other familial tumors. We validated these results with a new series of patient cases. The final study with 253 familial patient cases (74 BRCA1, 71 BRCA2, 108 non-BRCA1/2), and 288 sporadic patient cases, has allowed us to confirm our preliminary results. Because BRCA2 tumors present a specific immunohistochemical profile for RAD51 and CHEK2 markers that is different from non-BRCA1/2 tumors, we have built a multivariate model with these markers that distinguish both tumors with an estimated probability of at least 76%. CONCLUSION: Our results suggest that BRCA2 tumors demonstrate more cytoplasmic and less nuclear RAD51 staining, and increased CHEK2 staining. This pattern may distinguish BRCA2 from familial non-BRCA1/2 tumors.     

Clinical–pathologic significance of cancer stem cell marker expression in familial breast cancers

Human breast cancer cells with a CD44⁺/CD24^?/low or ALDH1+ phenotype have been demonstrated to be enriched for cancer stem cells (CSCs) using in vitro and in vivo techniques. The aim of this study was to determine the association between CD44⁺/CD24^?/low and ALDH1 expression with clinical–pathologic tumor characteristics, tumor molecular subtype, and survival in a well characterized collection of familial breast cancer cases. 364 familial breast cancers from the Ontario Familial Breast Cancer Registry (58 BRCA1-associated, 64 BRCA2-associated, and 242 familial non-BRCA1/2 cancers) were studied. Each tumor had a centralized pathology review performed. TMA sections of all tumors were analyzed for the expression of ER, PR, HER2, CK5, CK14, EGFR, CD44, CD24, and ALDH1. The Chi square test or Fisher’s exact test was used to analyze the marker associations with clinical–pathologic tumor variables, molecular subtype and genetic subtype. Analyses of the association of overall survival (OS) with marker status were conducted using Kaplan–Meier plots and log-rank tests. The CD44⁺/CD24^?/low and ALDH1+ phenotypes were identified in 16% and 15% of the familial breast cancer cases, respectively, and associated with high-tumor grade, a high-mitotic count, and component features of the medullary type of breast cancer. CD44⁺/CD24^?/low and ALDH1 expression in this series were further associated with the basal-like molecular subtype and the CD44⁺/CD24^?/low phenotype was independently associated with BRCA1 mutational status. The currently accepted breast CSCs markers are present in a minority of familial breast cancers. Whereas the presence of these markers is correlated with several poor prognostic features and the basal-like subtype of breast cancer, they do not predict OS.     

Estrogen receptor status could modulate the genomic pattern in familial and sporadic breast cancer.

PURPOSE: Familial breast cancer represents 5% to 10% of all breast tumors. Mutations in the two known major breast cancer susceptibility genes, BRCA1 and BRCA2, account for a minority of familial breast cancer, whereas families without mutations in these genes (BRCAX group) account for 70% of familial breast cancer cases. EXPERIMENTAL DESIGN: To better characterize and define the genomic differences between the three classes of familial tumors and sporadic malignancies, we have analyzed 19 BRCA1, 24 BRCA2, and 31 BRCAX samples from familial breast cancer patients and 19 sporadic breast tumors using a 1-Mb resolution bacterial artificial chromosome array-based comparative genomic hybridization. RESULTS: We found that BRCA1/2 tumors showed a higher genomic instability than BRCAX and sporadic cancers. There were common genomic alterations present in all breast cancer groups, such as gains of 1q and 16p or losses of 8ptel-p12 and 16q. We found that the presence/absence of the estrogen receptor (ER) may play a crucial role in driving tumor development through distinct genomic pathways independently of the tumor type (sporadic or familial) and mutation status (BRCA1 or BRCA2). ER(-) tumors presented higher genomic instability and different altered regions than ER+ ones. CONCLUSIONS: According to our results, the BRCA gene mutation status (mainly BRCA1) would contribute to the genomic profile of abnormalities by increasing or modulating the genome instability.     

Association of Poor Prognosis Subtypes of Breast Cancer with Estrogen Receptor Alpha Methylation in Iranian Women

Breast cancer is a prevalent heterogeneous malignant disease. Gene expression profiling by DNA microarraycan classify breast tumors into five different molecular subtypes: luminal A, luminal B, HER-2, basal and normallikewhich have differing prognosis. Recently it has been shown that immunohistochemistry (IHC) markersincluding estrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor 2(Her2), can divide tumors to main subtypes: luminal A (ER+; PR+/-; HER-2-), luminal B (ER+;PR+/-; HER-2+),basal-like (ER-;PR-;HER2-) and Her2+ (ER-; PR-; HER-2+). Some subtypes such as basal-like subtype have beencharacterized by poor prognosis and reduced overall survival. Due to the importance of the ER signaling pathwayin mammary cell proliferation; it appears that epigenetic changes in the ERα gene as a central component of thispathway, may contribute to prognostic prediction. Thus this study aimed to clarify the correlation of differentIHC-based subtypes of breast tumors with ERα methylation in Iranian breast cancer patients. For this purposeone hundred fresh breast tumors obtained by surgical resection underwent DNA extraction for assessedment oftheir ER methylation status by methylation specific PCR (MSP). These tumors were classified into main subtypesaccording to IHC markers and data were collected on pathological features of the patients. ERα methylation wasfound in 25 of 28 (89.3%) basal tumors, 21 of 24 (87.5%) Her2+ tumors, 18 of 34 (52.9%) luminal A tumors and7 of 14 (50%) luminal B tumors. A strong correlation was found between ERα methylation and poor prognosistumor subtypes (basal and Her2+) in patients (P<0.001). Our findings show that ERα methylation is correlatedwith poor prognosis subtypes of breast tumors in Iranian patients and may play an important role in pathogenesisof the more aggressive breast tumors.     

Genomic analysis of the 8p11-12 amplicon in familial breast cancer

Amplification of 8p11-12 has been recurrently reported in sporadic breast cancer. These studies define a complex molecular structure with a set of minimal amplified regions, and different putative oncogenes that show a strong correlation between amplification and over-expression such as ZNF703/FLJ14299, SPFH2/C8orf2, BRF2 and RAB11FIP. However, none of these studies were carried out on familial breast malignancies. We have studied the incidence, molecular features and clinical value of this amplification in familial breast tumors associated with BRCA1, BRCA2 and non-BRCA1/2 gene mutations. We detected 9 out of 80 familial tumors with this amplicon by chromosomal comparative genomic hybridization. Next, we used a high-resolution comparative genomic hybridization array covering the 8p11-12 region to characterize this chromosomal region. This approach allowed us to define 2 cores of common amplification that largely overlap with those reported in sporadic tumors. Our findings confirm the molecular complexity of this chromosomal region and indicate that this genomic event is a common alteration in breast cancer, present not only in sporadic but also in familial tumors. Finally, we found correlation between the 8p11-12 amplification and proliferation (Ki-67) and cyclin E expression, which further proves in familial tumors the poor prognosis association previously reported in sporadic breast cancer.     

A retrospective study of breast cancer subtypes: the risk of relapse and the relations with treatments

Immunohistochemical markers are often used to classify breast cancer into subtypes that are biologically distinct and behave differently. The aim of this study was to estimate relapse for patients with the major subtypes of breast cancer as classified using immunohistochemical assay and to investigate the patterns of benefit from the therapies over the past years. The study population included primary, operable 2,118 breast cancer patients, all non-specific infiltrative ductal carcinoma, with the median age of 53.2 years. All patients underwent local and/or systemic treatments. The clinicopathological characteristics and clinical outcomes were retrospectively reviewed. The expression of estrogen receptor (ER), progesterone receptor, human epidermal growth factor receptor 2 (HER2), epidermal growth factor receptor (EGFR), and cytokeratin 5/6 were analyzed by immunohistochemistry. All patients were classified into the following categories: luminal A, luminal B, HER2 overexpressing, basal-like, and unclassified subtypes. Ki-67 was detected in luminal A subtype. The median follow-up time was 67.9 months. Luminal A tumors had the lowest rate of relapse (12.7%, P < 0.001), while luminal B, HER2 overexpression, and basal-like subtypes were associated with an increased risk of relapse (15.7, 19.1, 20.9%). Molecular subtypes retained independent prognostic significance (P < 0.001). In luminal A subtype, adjunctive radiotherapy could decrease the risk of relapse (P = 0.005), Ki67 positive was a high-risk factor for relapse (P < 0.001), and adjuvant chemotherapies could reduce the relapse for the patients with risk factors (P < 0.001). Adjuvant hormone therapy was an effective treatment for ER-positive tumors (P < 0.001). Molecular subtypes of breast cancer could robustly identify the risk of recurrence and were significant in therapeutic decision making. The model combined subtype and clinical pathology was a significant improvement. Luminal A tumors might represent two distinct subsets which demonstrated distinct prognosis and therapy response.     

Understanding and treating triple-negative breast cancer

Triple-negative breast cancer is a subtype of breast cancer that is clinically negative for expression of estrogen and progesterone receptors (ER/PR) and HER2 protein. It is characterized by its unique molecular profile, aggressive behavior, distinct patterns of metastasis, and lack of targeted therapies. Although not synonymous, the majority of triple-negative breast cancers carry the "basal-like" molecular profile on gene expression arrays. The majority of BRCA1-associated breast cancers are triple-negative and basal-like; the extent to which the BRCA1 pathway contributes to the behavior of sporadic basal-like breast cancers is an area of active research. Epidemiologic studies illustrate a high prevalence of triple-negative breast cancers among younger women and those of African descent. Increasing evidence suggests that the risk factor profile differs between this subtype and the more common luminal subtypes. Although sensitive to chemotherapy, early relapse is common and a predilection for visceral metastasis, including brain metastasis, is seen. Targeted agents, including epidermal growth factor receptor (EGFR), vascular endothelial growth factor (VEGF), and poly (ADP-ribose) polymerase (PARP) inhibitors, are currently in clinical trials and hold promise in the treatment of this aggressive disease.     

Traditional breast cancer risk factors in relation to molecular subtypes of breast cancer

At least four major categories of invasive breast cancer have been reproducibly identified by gene expression profiling: luminal A, luminal B, HER2-type, and basal-like. These subtypes have been shown to differ in their outcome and response to treatment. Whether this heterogeneity reflects the evolution of these subtypes through distinct etiologic pathways has not been clearly defined. We evaluated the association between traditional breast cancer risk factors and risk of previously defined molecular subtypes of breast cancer in the Nurses’ Health Study. This analysis included 2,022 invasive breast cancer cases for whom we were able to obtain archived breast cancer tissue specimens. Tissue microarrays (TMAs) were constructed, and slides were immunostained for estrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2), cytokeratin 5/6 (CK5/6), and epidermal growth factor receptor (EGFR). Using immunostain results in combination with histologic grade, cases were grouped into molecularly defined subtypes. We used Cox proportional hazards models to estimate hazard ratios (HRs) and 95% confidence intervals (CIs). We observed differences in the association between risk factors and subtypes of breast cancer. In general, many reproductive factors were most strongly associated with the luminal A subtype, although these differences were not statistically significant. Weight gain since age 18 showed significant differences in its association with molecular subtypes (P-heterogeneity = 0.05) and was most strongly associated with the luminal B subtype (P-trend 0.001). Although there was not significant heterogeneity for lactation across subtypes, an inverse association was strongest for basal-like tumors (HR = 0.6, 95% CI 0.4–0.8; P-heterogeneity = 0.88). These results support the hypothesis that different subtypes of breast cancer have different etiologies and should not be considered as a single group. Identifying risk factors for less common subtypes such as luminal B, HER2-type and basal-like tumors has important implications for prevention of these more aggressive subtypes.     

A non-BRCA1/2 hereditary breast cancer sub-group defined by aCGH profiling of genetically related patients

Germline mutations in BRCA1 and BRCA2 explain approximately 25% of all familial breast cancers. Despite intense efforts to find additional high-risk breast cancer genes (BRCAx) using linkage analysis, none have been reported thus far. Here we explore the hypothesis that BRCAx breast tumors from genetically related patients share a somatic genetic etiology that might be revealed by array comparative genomic hybridization (aCGH) profiling. As BRCA1 and BRCA2 tumors can be identified on the basis of specific genomic profiles, the same may be true for a subset of BRCAx families. Analyses used aCGH to compare 58 non-BRCA1/2 familial breast tumors (designated BRCAx) to sporadic (non-familiar) controls, BRCA1 and BRCA2 tumors. The selection criteria for BRCAx families included at least three cases of breast cancer diagnosed before the age of 60 in the family, and the absence of ovarian or male breast cancer. Hierarchical cluster analysis was performed to determine sub-groups within the BRCAx tumor class and family heterogeneity. Analysis of aCGH profiles of BRCAx tumors indicated that they constitute a heterogeneous class, but are distinct from both sporadic and BRCA1/2 tumors. The BRCAx class could be divided into sub-groups. One subgroup was characterized by a gain of chromosome 22. Tumors from family members were classified within the same sub-group in agreement with the hypothesis that tumors from the same family would harbor a similar genetic background. This approach provides a method to target a sub-group of BRCAx families for further linkage analysis studies.     

Short telomeres are frequent in hereditary breast tumors and are associated with high tumor grade

Telomere shortening is a common event involved in malignant transformation. Critically short telomeres may trigger chromosomal aberrations and produce genomic instability leading to cancer development. Therefore, telomere shortening is a frequent molecular alteration in early stages of many epithelial tumors and in breast cancer correlates with stage and prognosis. A better understanding of the involvement of short telomeres in tumors may have a significant impact on patient management and the design of more specific treatments. To understand the role of telomere length (TL) in breast cancer etiology we measured the length of individual telomere signals in single cells by using quantitative telomere in situ hybridization in paraffin-embedded tissue from hereditary and sporadic breast cancers. A total of 104 tumor tissue samples from 75 familial breast tumors (BRCA1, n = 14; BRCA2, n = 13; non-BRCA1/2, n = 48) and 29 sporadic tumors were analyzed. Assessment of telomere signal intensity allowed estimation of the mean TL and related variables, such as percentage of critically short telomeres and percentage of cells with short telomeres. These data were correlated with the immunohistochemical expression of molecular breast cancer markers. Hereditary BRCA1, BRCA2, and non-BRCA1/2 tumors were characterized by shorter TL comparing to sporadic tumors. Considering all tumors, tumor grade was a strong risk factor determining the proportion of short telomeres or short telomere cells. Moreover, some histopathological features appeared to be differentially associated to hereditary or sporadic subgroups. Short telomeres correlated with ER-negative tumors in sporadic cases but not in familial cases, whereas a high level of apoptosis was associated with shorter telomeres in hereditary BRCA1 and BRCA2 tumors. In addition, TL helped to define a subset of non-BRCA1/2 tumors with short telomeres associated with increased expression of antiapoptotic proteins. These findings highlight the potential interest of TL measurements as markers of aggressiveness in breast cancer.     

Contemplating chemosensitivity of basal-like breast cancer based on BRCA1 dysfunction

Gene-expression profiling classified breast cancer to intrinsic subtypes, including luminal A and B, HER2 positive, normal-breast-like, and basal-like tumors. Of these, basal-like tumors that express basal cytokeratins and that are negative for estrogen receptor α, progesterone receptor, and HER2 show the most aggressive phenotype with a poor prognosis. Analyses of clinical samples and basic research indicate that basal-like breast cancer is caused by deficiencies in the breast cancer susceptibility protein, BRCA1. Indeed, conditionally deleting BRCA1 from the mammary gland causes mice to develop basal-like cancers at high rates. One of the major functions of BRCA1 is DNA double-strand break (DSB) repair, and its failure to perform causes increased sensitivity of cells to DNA damage-inducing agents, such as PARP inhibitors, DNA cross-linkers, or topoisomerase inhibitors. Therefore, BRCA1 dysfunction could be a principal target for therapeutic application of basal-like breast cancer. Recently, significant progress has been made in understanding the BRCA1 cascade in response to DSBs, where ubiquitin polymer formation plays critical roles. Ubiquitination was indeed found to be an apparent early response of breast cancer to neoadjuvant treatment with epirubicin and cyclophosphamide. Deducing the role of BRCA1 ubiquitin E3 ligase activity in this pathway is a critical challenge to further clarify its functional mechanism. In individualized treatment of breast cancer, evaluation of the DNA repair capacity by the BRCA1 pathway may be an important issue when determining proper treatment of basal-like breast cancer.