Functional mechanisms underlying pleiotropic risk alleles at the 19p13.1 breast-ovarian cancer susceptibility locus

A locus at 19p13 is associated with breast cancer (BC) and ovarian cancer (OC) risk. Here we analyse 438 SNPs in this region in 46,451 BC and 15,438 OC cases, 15,252 BRCA1 mutation carriers and 73,444 controls and identify 13 candidate causal SNPs associated with serous OC (P=9.2 × 10-20), ER-negative BC (P=1.1 × 10-13), BRCA1-associated BC (P=7.7 × 10-16) and triple negative BC (P-diff=2 × 10-5). Genotype-gene expression associations are identified for candidate target genes ANKLE1 (P=2 × 10-3) and ABHD8 (P<2 × 10-3). Chromosome conformation capture identifies interactions between four candidate SNPs and ABHD8, and luciferase assays indicate six risk alleles increased transactivation of the ADHD8 promoter. Targeted deletion of a region containing risk SNP rs56069439 in a putative enhancer induces ANKLE1 downregulation; and mRNA stability assays indicate functional effects for an ANKLE1 3′-UTR SNP. Altogether, these data suggest that multiple SNPs at 19p13 regulate ABHD8 and perhaps ANKLE1 expression, and indicate common mechanisms underlying breast and ovarian cancer risk

The FANCM:p.Arg658* truncating variant is associated with risk of triple-negative breast cancer

Abstract: Breast cancer is a common disease partially caused by genetic risk factors. Germline pathogenic variants in DNA repair genes BRCA1, BRCA2, PALB2, ATM, and CHEK2 are associated with breast cancer risk. FANCM, which encodes for a DNA translocase, has been proposed as a breast cancer predisposition gene, with greater effects for the ER-negative and triple-negative breast cancer (TNBC) subtypes. We tested the three recurrent protein-truncating variants FANCM:p.Arg658*, p.Gln1701*, and p.Arg1931* for association with breast cancer risk in 67,112 cases, 53,766 controls, and 26,662 carriers of pathogenic variants of BRCA1 or BRCA2. These three variants were also studied functionally by measuring survival and chromosome fragility in FANCM−/− patient-derived immortalized fibroblasts treated with diepoxybutane or olaparib. We observed that FANCM:p.Arg658* was associated with increased risk of ER-negative disease and TNBC (OR = 2.44, P = 0.034 and OR = 3.79; P = 0.009, respectively). In a country-restricted analysis, we confirmed the associations detected for FANCM:p.Arg658* and found that also FANCM:p.Arg1931* was associated with ER-negative breast cancer risk (OR = 1.96; P = 0.006). The functional results indicated that all three variants were deleterious affecting cell survival and chromosome stability with FANCM:p.Arg658* causing more severe phenotypes. In conclusion, we confirmed that the two rare FANCM deleterious variants p.Arg658* and p.Arg1931* are risk factors for ER-negative and TNBC subtypes. Overall our data suggest that the effect of truncating variants on breast cancer risk may depend on their position in the gene. Cell sensitivity to olaparib exposure, identifies a possible therapeutic option to treat FANCM-associated tumors

A case-only study to identify genetic modifiers of breast cancer risk for BRCA1/BRCA2 mutation carriers

Abstract: Breast cancer (BC) risk for BRCA1 and BRCA2 mutation carriers varies by genetic and familial factors. About 50 common variants have been shown to modify BC risk for mutation carriers. All but three, were identified in general population studies. Other mutation carrier-specific susceptibility variants may exist but studies of mutation carriers have so far been underpowered. We conduct a novel case-only genome-wide association study comparing genotype frequencies between 60,212 general population BC cases and 13,007 cases with BRCA1 or BRCA2 mutations. We identify robust novel associations for 2 variants with BC for BRCA1 and 3 for BRCA2 mutation carriers, P < 10−8, at 5 loci, which are not associated with risk in the general population. They include rs60882887 at 11p11.2 where MADD, SP11 and EIF1, genes previously implicated in BC biology, are predicted as potential targets. These findings will contribute towards customising BC polygenic risk scores for BRCA1 and BRCA2 mutation carriers

Functional mechanisms underlying pleiotropic risk alleles at the 19p13.1 breast-ovarian cancer susceptibility locus

A locus at 19p13 is associated with breast cancer (BC) and ovarian cancer (OC) risk. Here we analyse 438 SNPs in this region in 46,451 BC and 15,438 OC cases, 15,252 BRCA1 mutation carriers and 73,444 controls and identify 13 candidate causal SNPs associated with serous OC (P = 9.2 x 10(-20)), ER-negative BC (P = 1.1 x 10(-13)), BRCA1-associated BC (P = 7.7 x 10(-16)) and triple negative BC (P-diff = 2 x 10(-5)). Genotype-gene expression associations are identified for candidate target genes ANKLE1 (P = 2 x 10(-3)) and ABHD8 (PPeer reviewe

A case-only study to identify genetic modifiers of breast cancer risk for BRCA1/BRCA2 mutation carriers

Breast cancer (BC) risk for BRCA1 and BRCA2 mutation carriers varies by genetic and familial factors. About 50 common variants have been shown to modify BC risk for mutation carriers. All but three, were identified in general population studies. Other mutation carrier-specific susceptibility variants may exist but studies of mutation carriers have so far been underpowered. We conduct a novel case-only genome-wide association study comparing genotype frequencies between 60,212 general population BC cases and 13,007 cases with BRCA1 or BRCA2 mutations. We identify robust novel associations for 2 variants with BC for BRCA1 and 3 for BRCA2 mutation carriers, P < 10−8, at 5 loci, which are not associated with risk in the general population. They include rs60882887 at 11p11.2 where MADD, SP11 and EIF1, genes previously implicated in BC biology, are predicted as potential targets. These findings will contribute towards customising BC polygenic risk scores for BRCA1 and BRCA2 mutation carriers

Rôle de la Survivine et de ses transcrits alternatifs dans la progression tumorale et la résistance à la chimiothérapie (étude du cancer du sein)

La survivine est impliquée dans l apoptose et la régulation du cycle cellulaire. Par épissage alternatif, son gène code cinq transcrits dont les fonctions sont controversées. Notre étude a mis en évidence que des mutations de P53 sont impliquées dans l augmentation de l expression des trois transcrits anti-apoptotiques de la survivine dans le cancer du sein. De plus, l expression de survivine pourrait être contrôlée par de nouveaux gènes proches des microsatellites D3S1478 et D6S264. Enfin, la survivine est contrôlée par GATA-1 via un site constitutif et un site alternatif de fixation dans son promoteur. La seconde partie de notre travail a montré que les transcrits alternatifs de survivine pourraient avoir un rôle prédictif et pronostique pour la réponse à certains traitements de chimiothérapie et que l étude de la tumeur après une cure de chimiothérapie ajoute une information importante pour la prédiction et le pronostic de la maladie. La dernière partie de notre étude a mis en évidence les fonctions, dans deux lignées cellulaires mammaires, des cinq isoformes de survivine et notamment la survivine-3B. Cette dernière inhibe différentes voies de l apoptose et présente un intérêt thérapeutique particulier dû à sa spécificité tumorale. Des résultats encourageants ont été obtenus lors de son extinction in vivo. L ensemble de ces données montre l intérêt de l étude de la survivine et de ses transcrits alternatifs dans le cancer du sein, et notamment dans la réponse aux traitements. Ces résultats indiquent que la survivine-3B pourrait être une cible thérapeutique prometteuse.Survivin is involved in apoptosis and cell cycle regulation. By alternative splicing, its gene gives rise to five transcripts with controversial functions. Our study highlighted that P53 mutations are implied in anti-apoptotic survivin transcript over-expression. In addition, survivin expression could be controlled by new genes located closed to D3S1478 and D6S264 microsatellites. Moreover, survivin is controlled by GATA-1 via both constitutive and alternative binding sites in its promoter. The second part of our work showed that survivin alternative transcripts could have predictive and prognostic roles for response to chemotherapy and study of tumours after one course of chemotherapy could be beneficial for disease prediction and prognosis. The last part of our research highlighted that functions, in two breast tumour cell lines, of the five survivin isoforms, especially of survivin-3B. This last inhibits different apoptosis pathways and shows a therapeutic interest due to its tumour specificity. Interesting results were obtained in vivo. These data shows the survivin transcript study interest in breast cancer and treatment response. These results indicate survivin-3B could be a useful therapeutic target.DIJON-BU Sciences Economie (212312102) / SudocSudocFranceF

Impact des marqueurs de l'apoptose sur la progression tumorale et la réponse aux traitements dans les cancers du sein

Pour contourner le problème de chimiorésistance cancers du sein, il faut mettre en évidence de nouveaux marqueurs. Ainsi les gènes impliqués dans l apoptose pourraient servir de marqueurs prédictifs. Notre étude, portant sur la survivine, la caspase-3 et leurs transcrits alternatifs a monté que la survivine-2B pourraient servir de marqueur prédictif et la survivine-3B de marqueur pronostic. Nous montrons également que l expression des transcrits anti apoptotiques la survivine est régulée par p53. Par ailleurs, un site GATA-1 formé la présence d un polymorphisme dans le promoteur de la survivine semble induire sa surexpression. Nos résultats indiquent que la survivine-3B pourrait inhiber la voie de récepteurs de mort et que la caspase-3s a une fonction fortement anti-apoptotique par interaction avec la caspase-3 après un stimulus apoptotique. L ensemble de ces données met l accent sur le rôle majeur des transcrits alternatifs des gènes de l apoptose dans la réponse des cancers du sein à la chimiothérapie. Ils permettent d envisager ces protéines comme de nouvelles cibles thérapeutiques.To avoid chemoresistance of breast cancers, prognosis and predictive markers are required. Apoptosis is highly involved in treatment response, that is why apoptosis genes could become predictive markers. Our study, dealing with survivin, caspase-3, and their alternative splice variants shows that survivin-2B could be a predictive marker and survivin-3B a prognosis marker. We also show that survivin and its anti-apoptotic variant over-expression could be linked to p53 mutations. Moreover, a polymorphism in survivin promoter induces a GATA-1 binding site able to overexpress survivin. Our results showed that survivin-3B could inhibit death receptors pathway and that caspase-3s is strongly anti-apoptotic by interaction of caspase-3 after an apoptotic stimulus. This work shows the major role of alternative splice variants of apoptosis genes in breast cancer response and strongly supports that these proteins as new therapeutic targets.DIJON-BU Médecine Pharmacie (212312103) / SudocPARIS-BIUP (751062107) / SudocSudocFranceF

A Short Caspase-3 Isoform Inhibits Chemotherapy- Induced Apoptosis by Blocking Apoptosome Assembly

Alternative splicing of caspase-3 produces a short isoform caspase-3s that antagonizes caspase-3 apoptotic activity. However, the mechanism of apoptosis inhibition by caspase-3s remains unknown. Here we show that exogenous caspase-3 sensitizes MCF-7 and HBL100 breast cancers cells to chemotherapeutic treatments such as etoposide and methotrexate whereas co-transfection with caspase-3s strongly inhibits etoposide and methotrexate-induced apoptosis underlying thus the anti-apoptotic role of caspase-3s. In caspase-3 transfected cells, lamin-A and a-fodrin were cleaved when caspase-3 was activated by etoposide or methotrexate. When caspase-3s was co-transfected, this cleavage was strongly reduced. Depletion of caspase-3 by RNA interference in HBL100 containing endogenous caspase-3s caused reduction in etoposide and methotrexate-induced apoptosis, whereas the depletion of caspase-3s sensitized cells to chemotherapy. In the presence of caspase-3s, a lack of interaction between caspase-3 and caspase-9 was observed. Immunoprecipitation assays showed that caspase-3s binds the pro-forms of caspase-3. This result suggested that the absence of interaction with caspase-9 when both variants of caspase-3 are present contribute to block the apoptosome assembly and inhibit apoptosis. These data support that caspases-3s negatively interferes with caspase-3 activation and apoptosis in breast cancer, and tha