Novel germline MET pathogenic variants in French patients with papillary renal cell carcinomas type I.

Hereditary papillary renal cell carcinoma (HPRC) is a rare inherited renal cancer syndrome characterized by bilateral and multifocal papillary type 1 renal tumors (PRCC1). Activating germline pathogenic variants of the MET gene were identified in HPRC families. We reviewed the medical and molecular records of a large French series of 158 patients screened for MET oncogenic variants. MET pathogenic and likely pathogenic variants rate was 12.4% with 40.6% among patients with familial PRCC1 and 5% among patients with sporadic PRCC1. The phenotype in cases with MET pathogenic and likely pathogenic variants was characteristic: PRCC1 tumors were mainly bilateral (84.3%) and multifocal (87.5%). Histologically, six out of seven patients with MET pathogenic variant harbored biphasic squamoid alveolar PRCC. Genetic screening identified one novel pathogenic variant MET c.3389T>C, p.(Leu1130Ser) and three novel likely pathogenic variants: MET c.3257A>T, p.(His1086Leu); MET c.3305T>C, p.(Ile1102Thr) and MET c.3373T>G, p.(Cys1125Gly). Functional assay confirmed their oncogenic effect as they induced an abnormal focus formation. The genotype-phenotype correlation between MET pathogenic variants and PRCC1 presentation should encourage to widen the screening, especially toward nonfamilial PRCC1. This precise phenotype also constitutes a strong argument for the classification of novel missense variants within the tyrosine kinase domain when functional assays are not accessible.

VOPP1 promotes breast tumorigenesis by interacting with the tumor suppressor WWOX.

BACKGROUND: The WW domain-containing oxidoreductase (WWOX) gene, frequently altered in breast cancer, encodes a tumor suppressor whose function is mediated through its interactions with cancer-related proteins, such as the pro-apoptotic protein p73α. RESULTS: To better understand the involvement of WWOX in breast tumorigenesis, we performed a yeast two-hybrid screen and co-immunoprecipitation assays to identify novel partners of this protein. We characterized the vesicular overexpressed in cancer pro-survival protein 1 (VOPP1) as a new regulator of WWOX. In breast cancer cells, VOPP1 sequestrates WWOX in lysosomes, impairs its ability to associate with p73α, and inhibits WWOX-dependent apoptosis. Overexpressed VOPP1 potentiates cellular transformation and enhances the growth of transplanted tumors in vivo. VOPP1 is overexpressed in breast tumors, especially in tumors that retain WWOX. Moreover, increased expression of VOPP1 is associated with reduced survival of patients with WWOX-positive, but not with WWOX-negative, tumors. CONCLUSIONS: These findings emphasize the importance of the sequestration of WWOX by VOPP1 in addition to WWOX loss in breast tumors and define VOPP1 as a novel oncogene promoting breast carcinogenesis by inhibiting the anti-tumoral effect of WWOX.

A SUMOylation-defective MITF germline mutation predisposes to melanoma and renal carcinoma.

So far, no common environmental and/or phenotypic factor has been associated with melanoma and renal cell carcinoma (RCC). The known risk factors for melanoma include sun exposure, pigmentation and nevus phenotypes; risk factors associated with RCC include smoking, obesity and hypertension. A recent study of coexisting melanoma and RCC in the same patients supports a genetic predisposition underlying the association between these two cancers. The microphthalmia-associated transcription factor (MITF) has been proposed to act as a melanoma oncogene; it also stimulates the transcription of hypoxia inducible factor (HIF1A), the pathway of which is targeted by kidney cancer susceptibility genes. We therefore proposed that MITF might have a role in conferring a genetic predisposition to co-occurring melanoma and RCC. Here we identify a germline missense substitution in MITF (Mi-E318K) that occurred at a significantly higher frequency in genetically enriched patients affected with melanoma, RCC or both cancers, when compared with controls. Overall, Mi-E318K carriers had a higher than fivefold increased risk of developing melanoma, RCC or both cancers. Codon 318 is located in a small-ubiquitin-like modifier (SUMO) consensus site (ΨKXE) and Mi-E318K severely impaired SUMOylation of MITF. Mi-E318K enhanced MITF protein binding to the HIF1A promoter and increased its transcriptional activity compared to wild-type MITF. Further, we observed a global increase in Mi-E318K-occupied loci. In an RCC cell line, gene expression profiling identified a Mi-E318K signature related to cell growth, proliferation and inflammation. Lastly, the mutant protein enhanced melanocytic and renal cell clonogenicity, migration and invasion, consistent with a gain-of-function role in tumorigenesis. Our data provide insights into the link between SUMOylation, transcription and cancer.

Prognostic value of a newly identified MALAT1 alternatively spliced transcript in breast cancer.

BACKGROUND: Epigenetic deregulation is considered as a new hallmark of cancer. The long non-coding RNA MALAT1 has been implicated in several cancers; however, its role in breast cancer is still little known. METHODS: We used RT-PCR, in situ hybridisation, and RPPA methods to quantify (i) the full-length (FL) and an alternatively spliced variant (Δsv) of MALAT1, and (ii) a panel of transcripts and proteins involved in MALAT1 pathways, in a large series of breast tumours from patients with known clinical/pathological status and long-term outcome. RESULTS: MALAT1 was overexpressed in 14% (63/446) of the breast tumours. MALAT1-overexpressed tumour epithelial cells showed marked diffuse nuclear signals and numerous huge nuclear speckles. Screening of the dbEST database led to the identification of Δsv-MALAT1, a major alternatively spliced MALAT1 transcript, with a very different expression pattern compared with FL-MALAT1. This alternative Δsv-MALAT1 transcript was mainly underexpressed (18.8%) in our breast tumour series. Multivariate analysis showed that alternative Δsv-MALAT1 transcript is an independent prognostic factor. Δsv-MALAT1 expression was associated with alterations of the pre-mRNAs alternative splicing machinery, and of the Drosha-DGCR8 complex required for non-coding RNA biogenesis. Alternative Δsv-MALAT1 transcript expression was associated to YAP protein status and with an activation of the PI3K-AKT pathway. CONCLUSIONS: Our results reveal a complex expression pattern of various MALAT1 transcript variants in breast tumours, and suggest that this pattern of expressions should be taken into account to evaluate MALAT1 as predictive biomarker and therapeutic target.

MiR-190b, the highest up-regulated miRNA in ERα-positive compared to ERα-negative breast tumors, a new biomarker in breast cancers?

BACKGROUND: MicroRNAs (miRNAs) show differential expression across breast cancer subtypes and have both oncogenic and tumor-suppressive roles. Numerous microarray studies reported different expression patterns of miRNAs in breast cancers and found clinical interest for several miRNAs but often with contradictory results. Aim of this study is to identify miRNAs that are differentially expressed in estrogen receptor positive (ER(+)) and negative (ER(-)) breast primary tumors to better understand the molecular basis for the phenotypic differences between these two sub-types of carcinomas and to find potential clinically relevant miRNAs. METHODS: We used the robust and reproductive tool of quantitative RT-PCR in a large cohort of well-annotated 153 breast cancers with long-term follow-up to identify miRNAs specifically differentially expressed between ER(+) and ER(-) breast cancers. Cytotoxicity tests and transfection experiments were then used to examine the role and the regulation mechanisms of selected miRNAs. RESULTS: We identified a robust collection of 20 miRNAs significantly deregulated in ER(+) compared to ER(-) breast cancers : 12 up-regulated and eight down-regulated miRNAs. MiR-190b retained our attention as it was the miRNA the most strongly over-expressed in ER(+) compared to ER(-) with a fold change upper to 23. It was also significantly up-regulated in ER(+)/Normal breast tissue and down-regulated in ER(-)/Normal breast tissue. Functional experiments showed that miR-190b expression is not directly regulated by estradiol and that miR-190b does not affect breast cancer cell lines proliferation. Expression level of miR-190b impacts metastasis-free and event-free survival independently of ER status. CONCLUSIONS: This study reveals miR-190b as the highest up-regulated miRNA in hormone-dependent breast cancers. Due to its specificity and high expression level, miR-190b could therefore represent a new biomarker in hormone-dependent breast cancers but its exact role carcinogenesis remains to elucidate.

TPD52 represents a survival factor in ERBB2-amplified breast cancer cells.

TPD52 and ERBB2 co-expression has been persistently reported in human breast cancer and animal models of this disease, but the significance of this is unknown. We identified significant positive associations between relative TPD52 and ERBB2 transcript levels in human diagnostic breast cancer samples, and maximal TPD52 expression in the hormone receptor (HR)- and ERBB2-positive sub-group. High-level TPD52 expression was associated with significantly reduced metastasis-free survival, within the overall cohort (log rank test, P = 8.6 × 10(-4), n = 375) where this was an independent predictor of metastasis-free survival (hazard ratio, 2.69, 95% confidence interval 1.59-4.54, P = 2.2 × 10(-4), n = 359), and the HR- and ERBB2-positive sub-group (log rank test, P = 0.035, n = 47). Transient TPD52 knock-down in the ERBB2-amplified breast cancer cell lines SK-BR-3 and BT-474 produced significant apoptosis, both singly and in combination with transient ERBB2 knock-down. Unlike ERBB2 knock-down, transient TPD52 knock-down produced no reduction in pAKT levels in SK-BR-3 or BT-474 cells. We then derived multiple SK-BR-3 cell lines in which TPD52 levels were stably reduced, and measured significant inverse correlations between pERBB2 and TPD52 levels in viable TPD52-depleted and control cell lines, all of which showed similar proliferative capacities. Our results therefore identify TPD52 as a survival factor in ERBB2-amplified breast cancer cells, and suggest complementary cellular functions for TPD52 and ERBB2.

Toll-like receptor 4-dependent contribution of the immune system to anticancer chemotherapy and radiotherapy.

Conventional cancer treatments rely on radiotherapy and chemotherapy. Such treatments supposedly mediate their effects via the direct elimination of tumor cells. Here we show that the success of some protocols for anticancer therapy depends on innate and adaptive antitumor immune responses. We describe in both mice and humans a previously unrecognized pathway for the activation of tumor antigen-specific T-cell immunity that involves secretion of the high-mobility-group box 1 (HMGB1) alarmin protein by dying tumor cells and the action of HMGB1 on Toll-like receptor 4 (TLR4) expressed by dendritic cells (DCs). During chemotherapy or radiotherapy, DCs require signaling through TLR4 and its adaptor MyD88 for efficient processing and cross-presentation of antigen from dying tumor cells. Patients with breast cancer who carry a TLR4 loss-of-function allele relapse more quickly after radiotherapy and chemotherapy than those carrying the normal TLR4 allele. These results delineate a clinically relevant immunoadjuvant pathway triggered by tumor cell death.

Description and analysis of genetic variants in French hereditary breast and ovarian cancer families recorded in the UMD-BRCA1/BRCA2 databases.

BRCA1 and BRCA2 are the two main genes responsible for predisposition to breast and ovarian cancers, as a result of protein-inactivating monoallelic mutations. It remains to be established whether many of the variants identified in these two genes, so-called unclassified/unknown variants (UVs), contribute to the disease phenotype or are simply neutral variants (or polymorphisms). Given the clinical importance of establishing their status, a nationwide effort to annotate these UVs was launched by laboratories belonging to the French GGC consortium (Groupe Génétique et Cancer), leading to the creation of the UMD-BRCA1/BRCA2 databases (http://www.umd.be/BRCA1/ and http://www.umd.be/BRCA2/). These databases have been endorsed by the French National Cancer Institute (INCa) and are designed to collect all variants detected in France, whether causal, neutral or UV. They differ from other BRCA databases in that they contain co-occurrence data for all variants. Using these data, the GGC French consortium has been able to classify certain UVs also contained in other databases. In this article, we report some novel UVs not contained in the BIC database and explore their impact in cancer predisposition based on a structural approach.

Variations in the mRNA expression of poly(ADP-ribose) polymerases, poly(ADP-ribose) glycohydrolase and ADP-ribosylhydrolase 3 in breast tumors and impact on clinical outcome.

In order to assess the variation in expression of poly(ADP-ribose) polymerase (PARP) family members and the hydrolases that degrade the poly(ADP-ribose) polymers they generate and possible associations with classical pathological parameters, including long-term outcome, the mRNA levels of PARP1, PARP2, PARP3, poly(ADP-ribose) glycohydrolase (PARG) and ADP-ribosylhydrolase 3 (ARH3) were examined using quantitative reverse transcription polymerase chain reaction in 443 unilateral invasive breast cancers and linked to hormonal status, tumor proliferation and clinical outcome. PARP1 mRNA levels were the highest among these five genes in both normal and tumor tissues, with a 2.45-fold higher median level in tumors compared to normal tissues. Tumors (34.1%) showed PARP1 overexpression (>3 fold relative to normal breast tissues) compared to underexpression (<0.33 fold) in only 0.5%. This overexpression was seen in all breast tumor subgroups, with the highest fraction (51%) seen in the HR-positive/ERBB2-positive subgroup and was not highly associated with any other classical predictive factors. No correlation was seen between PARP1 mRNA and PARP-1 protein levels in a subset of 31 tumors. PARP3 was underexpressed in 10.4% of tumors, more frequently in the HR-negative tumors (25.4%) than the HR-positive tumors (5.9%). This PARP3 underexpression was mutually exclusive with a PARP1 overexpression. PARP2 levels were unchanged between normal and tumor tissues and few tumors showed overexpression of PARG (3.8%) or ARH3 (3.4%). Within the subgroup of triple negative tumors, PARG mRNA levels below the median were associated with a higher risk of developing metastases (p = 0.039) raising the possibility this might be marker of clinical outcome.

microRNA expression profile in a large series of bladder tumors: identification of a 3-miRNA signature associated with aggressiveness of muscle-invasive bladder cancer.

The aim of this study was to evaluate the expression levels of microRNAs (miRNAs) in bladder tumors in order to identify miRNAs involved in bladder carcinogenesis with potential prognostic implications. Expression levels of miRNAs were assessed by quantitative real-time RT-PCR in 11 human normal bladder and 166 bladder tumor samples (86 non-muscle-invasive bladder cancer (NMIBC) and 80 muscle-invasive bladder cancer (MIBC)). The expression level of 804 miRNAs was initially measured in a well-defined series of seven NMIBC, MIBC and normal bladder samples (screening set). The most strongly deregulated miRNAs in tumor samples compared to normal bladder tissue were then selected for RT-PCR validation in a well-characterized independent series of 152 bladder tumors (validation set), and in six bladder cancer cell lines. Expression levels of these miRNAs were tested for their association with clinical outcome. A robust group of 15 miRNAs was found to be significantly deregulated in bladder cancer. Except for two miRNAs, miR-146b and miR-9, which were specifically upregulated in MIBC, the majority of miRNAs (n = 13) were deregulated in the same way in the two types of bladder tumors, irrespective of pathological stage : three miRNAs were upregulated (miR-200b, miR-182 and miR-138) and the other 10 miRNAs were downregulated (miR-1, miR-133a, miR-133b, miR-145, miR-143, miR-204, miR-921, miR-1281, miR-199a and miR-199b). A 3-miRNA signature (miR-9, miR-182 and miR-200b) was found to be related to MIBC tumor aggressiveness and was associated with both recurrence-free and overall survival in univariate analysis with a trend to significance in the multivariate analysis (p = 0.05). Our results suggested a promising individual prognostic value of these new markers.

miRNA expression profiling of inflammatory breast cancer identifies a 5-miRNA signature predictive of breast tumor aggressiveness.

IBC (inflammatory breast cancer) is a rare but very aggressive form of breast cancer with a particular phenotype. The molecular mechanisms responsible for IBC remain largely unknown. In particular, genetic and epigenetic alterations specific to IBC remain to be identified. MicroRNAs, a class of small noncoding RNAs able to regulate gene expression, are deregulated in breast cancer and may therefore serve as tools for diagnosis and prediction. This study was designed to determine miRNA expression profiling (microRNAome) in IBC. Quantitative RT-PCR was used to determine expression levels of 804 miRNAs in a screening series of 12 IBC compared to 31 non-stage-matched non-IBC and 8 normal breast samples. The differentially expressed miRNAs were then validated in a series of 65 IBC and 95 non-IBC. From a set of 18 miRNAs of interest selected from the screening series, 13 were differentially expressed with statistical significance in the validation series of IBC compared to non-IBC. Among these, a 5-miRNA signature comprising miR-421, miR-486, miR-503, miR-720 and miR-1303 was shown to be predictive for IBC phenotype with an overall accuracy of 89%. Moreover, multivariate analysis showed that this signature was an independent predictor of poor Metastasis-Free Survival in non-IBC patients.

PIK3R1 underexpression is an independent prognostic marker in breast cancer.

BACKGROUND: The present study focused on the prognostic roles of PIK3CA and PIK3R1 genes and additional PI3K pathway-associated genes in breast cancer. METHODS: The mutational and mRNA expression status of PIK3CA, PIK3R1 and AKT1, and expression status of other genes involved in the PI3K pathway (EGFR, PDK1, PTEN, AKT2, AKT3, GOLPH3, WEE1, P70S6K) were assessed in a series of 458 breast cancer samples. RESULTS: PIK3CA mutations were identified in 151 samples (33.0%) in exons 1, 2, 9 and 20. PIK3R1 mutations were found in 10 samples (2.2%) and underexpression in 283 samples (61.8%). AKT1 mutations were found in 15 samples (3.3%) and overexpression in 116 samples (25.3%). PIK3R1 underexpression tended to mutual exclusivity with PIK3CA mutations (p = 0.00097). PIK3CA mutations were associated with better metastasis-free survival and PIK3R1 underexpression was associated with poorer metastasis-free survival (p = 0.014 and p = 0.00028, respectively). By combining PIK3CA mutation and PIK3R1 expression status, four prognostic groups were identified with significantly different metastasis-free survival (p = 0.00046). On Cox multivariate regression analysis, the prognostic significance of PIK3R1 underexpression was confirmed in the total population (p = 0.0013) and in breast cancer subgroups. CONCLUSIONS: PIK3CA mutations and PIK3R1 underexpression show opposite effects on patient outcome and could become useful prognostic and predictive factors in breast cancer.

Prediction of BRCA1 germ-line mutation status in patients with breast cancer using histoprognosis grade, MS110, Lys27H3, vimentin, and KI67.

Family structure, lack of reliable information, cost, and delay are usual concerns when deciding to perform BRCA analyses. Testing breast cancer tissues with four antibodies (MS110, lys27H3, vimentin, and KI67) in addition to grade evaluation enabled us to rapidly select patients for genetic testing identification. We constituted an initial breast cancer tissue microarray, considered as a learning set, comprising 27 BRCA1 and 81 sporadic tumors. A second independent validation set of 28 BRCA1 tumors was matched to 28 sporadic tumors using the same original conditions. We investigated morphological parameters and 21 markers by immunohistochemistry. A logistic regression model was used to select the minimal number of markers providing the best model to predict BRCA1 status. The model was applied to the validation set to estimate specificity and sensibility. In the initial set, univariate analyses identified 11 markers significantly associated with BRCA1 status. Then, the best multivariate model comprised only grade 3, MS110, Lys27H3, vimentin, and KI67. When applied to the validation set, BRCA1 tumors were correctly classified with a sensitivity of 83% and a specificity of 81%. The performance of this model was superior when compared to other profiles. This study offers a new rapid and cost-effective method for the prescreening of patients at high risk of being BRCA1 mutation carriers, to guide genetic testing, and finally to provide appropriate preventive measures, advice, and treatments including targeted therapy to patients and their families.

Multiple sequence variants of BRCA2 exon 7 alter splicing regulation.

BACKGROUND: Exonic variants of unknown biological significance (VUS) identified in patients can affect mRNA splicing, either by changing 5' or 3' splice sites or by modifying splicing regulatory elements. Bioinformatic predictions of these elements are still inaccurate and only few such elements have been functionally mapped in BRCA2. We studied the effect on splicing of eight exon 7 VUS, selected from the French UMD-BRCA2 mutation database. METHODS: We performed splicing minigene assays and analyses of patient RNA. We also developed a pyrosequencing-based quantitative assay, to measure, in patient RNA, the relative contribution of each allele to the production of exon 7-containing transcripts. Moreover, an exonic splicing enhancer (ESE)-dependent minigene assay was used to evaluate the splicing regulatory properties of wild-type and mutant segments. RESULTS: Six out of the eight variants induced splicing defects. In the minigene assay, c.517G>T and c.631G>A altered the natural splice sites, c.572A>G created a new 5' splice site, and c.520C>T, c.587G>A and c.617C>G induced exon 7 skipping (66%, 25% and 46%, respectively). Pyrosequencing of patient RNA confirmed these levels of exon skipping for c.520C>T and c.617C>G. Results from the ESE-dependent minigene assay indicated that c.520C>T and c.587G>A disturb splicing regulatory elements. CONCLUSIONS: BRCA2 exon 7 splicing is regulated by multiple exonic elements and is sensitive to disease-associated sequence variations. Measurements of allelic imbalance in patient-derived RNA and/or quantitative analyses using minigene assays provide valuable estimates of the extent of partial splicing defects. Assessment of pathogenicity of variants with partial splicing effect awaits additional evidence and especially the completion of segregation analyses.

BRCA1 R1699Q variant displaying ambiguous functional abrogation confers intermediate breast and ovarian cancer risk.

BACKGROUND: Clinical classification of rare sequence changes identified in the breast cancer susceptibility genes BRCA1 and BRCA2 is essential for appropriate genetic counselling of individuals carrying these variants. We previously showed that variant BRCA1 c.5096G>A p.Arg1699Gln in the BRCA1 transcriptional transactivation domain demonstrated equivocal results from a series of functional assays, and proposed that this variant may confer low to moderate risk of cancer. METHODS: Measures of genetic risk (report of family history, segregation) were assessed for 68 BRCA1 c.5096G>A p.Arg1699Gln (R1699Q) families recruited through family cancer clinics, comparing results with 34 families carrying the previously classified pathogenic BRCA1 c.5095C>T p.Arg1699Trp (R1699W) mutation at the same residue, and to 243 breast cancer families with no BRCA1 pathogenic mutation (BRCA-X). RESULTS: Comparison of BRCA1 carrier prediction scores of probands using the BOADICEA risk prediction tool revealed that BRCA1 c.5096G>A p.Arg1699Gln variant carriers had family histories that were less 'BRCA1-like' than BRCA1 c.5095C>T p.Arg1699Trp mutation carriers (p<0.00001), but more 'BRCA1-like' than BRCA-X families (p=0.0004). Further, modified segregation analysis of the subset of 30 families with additional genotyping showed that BRCA1 c.5096G >A p.Arg1699Gln had reduced penetrance compared with the average truncating BRCA1 mutation penetrance (p=0.0002), with estimated cumulative risks to age 70 of breast or ovarian cancer of 24%. CONCLUSIONS: Our results provide substantial evidence that the BRCA1 c.5096G>A p.Arg1699Gln (R1699Q) variant, demonstrating ambiguous functional deficiency across multiple assays, is associated with intermediate risk of breast and ovarian cancer, highlighting challenges for risk modelling and clinical management of patients of this and other potential moderate-risk variants.

A methodological approach to unravel organ-specific breast cancer metastasis.

Breast cancer is the most commonly diagnosed and the second highest cause of cancer-related mortality. Although major breakthroughs have emerged during the past decades concerning the characterization of major malignant tumors hallmarks, little is known about the molecular process that sustains the most deadly feature of cancer: metastasis to distant organs. In fact, this colonization of tumor cells to secondary sites is not random but rather orientated, and depends on several signalling events that are not fully elucidated yet. Understanding the precise molecular and cellular mechanisms accountable for the specific invasion of tissues by breast cancer cells is likely to be important for developing new therapeutic strategies to effectively prevent metastasis in patients diagnosed with early cancer lesions. Here, we briefly describe a multidisciplinary approach based on the molecular profiling of breast cancer metastases, the elaboration of prognostic gene signatures, the clinical validation and the experimental confirmation using cell and animal models to better address breast cancer metastasis. This methodology can be considered as a useful workflow to identify and validate the genes that trigger and support organ tropism of breast cancer cells during metastasis.

WWOX binds MERIT40 and modulates its function in homologous recombination, implications in breast cancer.

The tumor suppressor gene WWOX is localized in an unstable chromosomal region and its expression is decreased or absent in several types of cancer. A low expression of WWOX is associated with a poor prognosis in breast cancer (BC). It has recently been shown that WWOX contributes to genome stability through its role in the DNA damage response (DDR). In breast cancer cells, WWOX inhibits homologous recombination (HR), and thus promotes the repair of DNA double-stranded breaks (DSBs) by non-homologous end joining (NHEJ). The fine-tuning modulation of HR activity is crucial. Its under or overstimulation inducing genome alterations that can induce cancer. MERIT40 is a positive regulator of the DDR. This protein is indispensable for the function of the multi-protein complex BRCA1-A, which suppresses excessive HR activity. MERIT40 also recruits Tankyrase, a positive regulator of HR, to the DSBs to stimulate DNA repair. Here, we identified MERIT40 as a new molecular partner of WWOX. We demonstrated that WWOX inhibited excessive HR activity induced by overexpression of MERIT40. We showed that WWOX impaired the MERIT40-Tankyrase interaction preventing the role of the complex on DSBs. Furthermore, we found that MERIT40 is overexpressed in BC and that this overexpression is associated to a poor prognosis. These results strongly suggest that WWOX, through its interaction with MERIT40, prevents the deleterious impact of excessive HR on BC development by inhibiting MERIT40-Tankyrase association. This inhibitory effect of WWOX would oppose MERIT40-dependent BC development.

HORMAD1 overexpression predicts response to anthracycline-cyclophosphamide and survival in triple-negative breast cancers.

Triple negative breast cancers (TNBCs) represent 15-20% of all breast cancers and are associated with higher recurrence and distant metastasis rate. Standard of care for early stage TNBC is anthracyclines combined with cyclophosphamide (AC) followed by taxanes, in the neo-adjuvant or adjuvant setting. This work aimed to identify predictive biomarkers of AC response in patient-derived xenograft (PDX) models of TNBC and to validate them in the clinical setting. By gene and protein expression analysis of 39 PDX with different responses to AC, we found that high expression of HORMAD1 was associated with better response to AC. Both gene and protein expression were associated with promoter hypomethylation. In a cohort of 526 breast cancer patients, HORMAD1 was overexpressed in 71% of TNBC. In a second cohort of 186 TNBC patients treated with AC, HORMAD1 expression was associated with longer metastasis-free survival (MFS). In summary, HORMAD1 overexpression was predictive of an improved response to AC in PDX and is an independent prognostic factor in TNBC patients treated with AC.

A diagnostic genetic test for the physical mapping of germline rearrangements in the susceptibility breast cancer genes BRCA1 and BRCA2.

The BRCA1 and BRCA2 genes are involved in breast and ovarian cancer susceptibility. About 2 to 4% of breast cancer patients with positive family history, negative for point mutations, can be expected to carry large rearrangements in one of these two genes. We developed a novel diagnostic genetic test for the physical mapping of large rearrangements, based on molecular combing (MC), a FISH-based technique for direct visualization of single DNA molecules at high resolution. We designed specific Genomic Morse Codes (GMCs), covering the exons, the noncoding regions, and large genomic portions flanking both genes. We validated our approach by testing 10 index cases with positive family history of breast cancer and 50 negative controls. Large rearrangements, corresponding to deletions and duplications with sizes ranging from 3 to 40 kb, were detected and characterized on both genes, including four novel mutations. The nature of all the identified mutations was confirmed by high-resolution array comparative genomic hybridization (aCGH) and breakpoints characterized by sequencing. The developed GMCs allowed to localize several tandem repeat duplications on both genes. We propose the developed genetic test as a valuable tool to screen large rearrangements in BRCA1 and BRCA2 to be combined in clinical settings with an assay capable of detecting small mutations.