Biomedical subjects

Dominique Stoppa-Lyonnet

Publications and source records attributed to Dominique Stoppa-Lyonnet.

At least 55 records · Page 3Linked to original sources

PubMed2003

Phenotypic cellular characterization of an ataxia telangiectasia patient carrying a causal homozygous missense mutation.

Most disease-causing mutations in Ataxia telangiectasia (AT) patients correspond to truncating mutations in the ATM gene with very few cases of AT patients carrying two missense sequence alterations being reported. The cellular phenotype of a lymphoblastoid cell line established from an AT patient (AT173) who showed classical clinical AT features, and carried two homozygous missense alterations, the 378T>A variant and 9022C>T located within the ATM kinase domain, has been characterized. ATM mRNA was detectable and the ATM protein level was approximately 50% of that seen in normal cell lines. Functional analysis of this protein revealed a total absence of ATM kinase activity measured either in vitro or in vivo, before and after exposure to ionizing radiation. The AT173 cell line was hypersensitive to ionizing radiation and exhibited a G1 cell cycle arrest defect and an accumulation of cells in G2 phase of the cell cycle after irradiation, a response that is identical to that seen in AT cell lines carrying truncating mutations. These phenotypic features strongly suggest that the 9022C>T (R3008C) missense mutation is the disease-causing mutation and that the presence of ATM protein is not always predictive of a normal cellular phenotype.

Ataxia Telangiectasia↗

PubMed2003

Haplotype analysis of the BRCA2 9254delATCAT recurrent mutation in breast/ovarian cancer families from Spain.

A frame-shift 9254del5 mutation was independently identified in 12 families, eleven of them with Spanish ancestors, in a BRCA2 screening performed in 841 breast and/or ovarian cancer families and in 339 women with breast cancer diagnosed before the age of 40 at different centers in France and Spain. We sought to analyze in detail the haplotype and founder effects of the 9254del5 and to estimate the time of origin of the mutation. Eight polymorphic microsatellite markers and two BRCA2 polymorphisms were used for the haplotype analyses. The markers were located flanking the BRCA2 gene spanning a region of 6.1 cM. Our results suggest that these families shared a common ancestry with BRCA2 9254del5, which is a founder mutation originating in the Northeast Spanish, with an estimated age of 92 (95% CI 56-141) generations.

Adult↗

PubMed2003

Characterisation of a 161 kb deletion extending from the NBR1 to the BRCA1 genes in a French breast-ovarian cancer family.

A large germline deletion removing exons 1 to 22 of the BRCA1 gene has been previously detected using quantitative PCR based methods (QMPSF and real time PCR gene dosage assay) in a woman affected with breast and ovarian cancer. Here, we report its characterisation by using colour bar code on combed DNA of the BRCA1 region. The 5' boundary is located in a Alu Y sequence in NBR1 intron 18 whereas the 3' boundary is located in a Alu Sc sequence in BRCA1 intron 22. This 161 kb deletion encompassing the NBR1, PsiBRCA1, NBR2 and BRCA1 genes is the largest BRCA1 deletion reported so far. No specific phenotype was associated with the hemizygosity of these four genes.

BRCA1 Protein↗

PubMed2003

Impact of gene patents on the cost-effective delivery of care: the case of BRCA1 genetic testing.

BACKGROUND: In 1994/95, two genes, BRCA1/2, associated with a predisposition to breast or ovarian cancer were identified. Genetic testing of deleterious BRCA1/2 mutations consequently can be proposed to individuals with a family history of breast or ovarian cancer to identify who is at risk. The granting of U.S. patents on BRCA1/2 to a privately owned company has led to the monopoly use of a unique technique (Direct Sequencing of the gene, DS) for BRCA1/2 testing in this country. Alternative strategies using prescreening techniques, however, have been experienced worldwide. METHODS: On the basis of data collected at three laboratories of French public hospitals, we carried out a cost-effectiveness study comparing DS to 19 alternative strategies with the number of deleterious BRCA1 mutations detected as the outcome. RESULTS: Results show that the DS strategy presents the highest average cost per mutation detected (9,882.5 Euro) and that there exist strategies using prescreening techniques that can reach similar effectiveness while reducing total costs. Moreover, other strategies can obtain a four- to sevenfold reduction in the average cost per mutation detected as soon as some rates of false negatives (2% to 13%) are deemed to be acceptable. CONCLUSIONS: Results suggest that gene patents with a very broad scope, covering all potential medical applications, may prevent health care systems from identifying and adopting the most efficient genetic testing strategies due to the monopoly granted for the exploitation of the gene. Policy implications for regulatory authorities, in the current context of the extension of BRCA1/2 patents in other countries, are discussed.

Breast Neoplasms↗

PubMed2003

Testing participation in BRCA1/2-positive families: initiator role of index cases.

The objectives of this study were to: (1) describe diffusion of information by affected women in whom a mutation has been identified (index cases) to their families and testing participation among high-risk relatives; (2) assess information recall and understanding by index cases and their satisfaction with the testing process; and (3) determine the factors associated with higher/lower testing decision in the family. Thirty index cases completed a self-administered questionnaire assessing their personal and family characteristics and their satisfaction with their own genetic testing process and a telephone interview to evaluate their knowledge about the risk of a genetic predisposition to breast and ovarian cancer, the type and number of close relatives that they informed, and the difficulties that they encountered. Information about breast/ovarian cancer risk and test availability was generally well transmitted (75%), predominantly (88%) to first-degree relatives. In contrast, testing participation was low (15%) and essentially occurred among sisters and daughters. There was a general lack of knowledge despite a high level of satisfaction regarding the information given by the geneticist. Family support and the knowledge of index cases about the risk of transmission of BRCA1/2 mutations by women were found to be positively and significantly associated with the testing decision among first-degree relatives. Difficulties in informing relatives appeared to be related to poor understanding of the information by index cases, as well as fear, and avoidance among close relatives. A major challenge for genetic counseling is to ensure that consulting patients not only receive complete understanding but also understand this information and anticipate the impact of the test result before deciding to take the test.

Adult↗

PubMed2003

BRCA1 wild-type allele modifies risk of ovarian cancer in carriers of BRCA1 germ-line mutations.

Strong inter- and intrafamilial variation of penetrance of breast and ovarian cancer is observed in BRCA1 mutation carriers. The wild-type copy of the BRCA1 gene is a plausible candidate as a cancer risk modifier given that the residual function corresponding to the intact BRCA1 allele may influence the process of tumor formation in BRCA1 carriers. Indeed, growing evidence is now becoming available on impaired reparation of double-strand DNA breaks in cells heterozygous for BRCA1 mutations, implying an enhanced mutability of BRCA1(+/-) cells. To determine whether certain variant forms of the wild-type BRCA1 allele are implicated in variation of the BRCA1-related cancer risk, their effect was studied in a panel of 591 women with BRCA1 germ-line mutations. We found that BRCA1 carriers with the wild-type BRCA1 copy bearing a common Gly1038 variant were at increased risk of ovarian cancer (hazards ratio, 1.50; 95% confidence interval, 1.03-2.19). The results of our study imply that a quite significant proportion of the interindividual variability in ovarian cancer penetrance in BRCA1 carriers may be explained by a common BRCA1 Gly1038 wild-type allele, given its high frequency (0.27).

Adult↗

PubMed2002

The nonsense-mediated mRNA decay pathway triggers degradation of most BRCA1 mRNAs bearing premature termination codons.

Germline mutations in the BRCA1 gene are scattered over the 22 coding exons and most of them generate premature termination codons (PTCs). A mechanism called nonsense-mediated mRNA decay (NMD) is known to specifically degrade transcripts with PTCs; however, steady-state amounts of mutant BRCA1 mRNAs have very rarely been measured. Although growing evidence implicates downstream exon-exon junctions (EEJs) as critical determinants for discrimination between normal stop codons and PTCs, requirements concerning the minimal and maximal distance between PTCs and downstream EEJs are still debated. We assessed the relative amount of transcripts encoded by BRCA1 alleles harbouring 30 different truncating mutations in lymphoblastoid cell lines established from carriers from breast/ovarian cancer families. We found that NMD is triggered by 80% of PTC(+) alleles and results in a 1.5- to 5-fold reduction in mRNA abundance. All truncating mutations located in the 3.4 kb long central exon are subject to NMD, irrespective of their distance to the downstream EEJ (305 to 3395 nt). PTCs not leading to NMD are either located in the last exon or very close to the translation initiation codon. We hypothesize that reinitiation could explain why transcripts carrying early PTCs escape NMD. This is the first study challenging the NMD rules, which have been established through the study of minigenes, by analysing a large series of mutant endogenous alleles.

Amino Acid Substitution↗

PubMed2002

Significant contribution of large BRCA1 gene rearrangements in 120 French breast and ovarian cancer families.

Genetic linkage data have shown that alterations of the BRCA1 gene are responsible for the majority of hereditary breast-ovarian cancers. However, BRCA1 germline mutations are found much less frequently than expected, especially as standard PCR-based mutation detection approaches focus on point and small gene alterations. In order to estimate the contribution of large gene rearrangements to the BRCA1 mutation spectrum, we have extensively analysed a series of 120 French breast-ovarian cancer cases. Thirty-eight were previously found carrier of a BRCA1 point mutation, 14 of a BRCA2 point mutation and one case has previously been reported as carrier of a large BRCA1 deletion. The remaining 67 cases were studied using the BRCA1 bar code approach on combed DNA which allows a panoramic view of the BRCA1 region. Three additional rearrangements were detected: a recurrent 23.8 kb deletion of exons 8-13, a 17.2 kb duplication of exons 3-8 and a 8.6 kb duplication of exons 18-20. Thus, in our series, BRCA1 large rearrangements accounted for 3.3% (4/120) of breast-ovarian cancer cases and 9.5% (4/42) of the BRCA1 gene mutation spectrum, suggesting that their screening is an important step that should be now systematically included in genetic testing surveys.

Adult↗

PubMed2002

Variation in breast cancer risk of heterozygotes for ataxia-telangiectasia according to environmental factors.

Ataxia-telangiectasia (AT) is a rare autosomal recessive disorder, characterized by progressive neuronal degeneration, immunological deficiency, radio-sensitivity and an increased risk of cancer. Although several studies have confirmed that AT heterozygosis increases the risk of breast cancer (BC), we do not know how exogenous factors affect this risk. We performed an epidemiological study on the cancer risks associated with AT heterozygosis in France and explored the variation in BC risk according to environmental factors, such as reproductive factors and exposure to ionizing radiation. Information on the amount of ionizing radiation received by an individual in their lifetime and on their reproductive life was collected from the living relatives of 34 AT children (175 female relatives). Consistent with previous reports and with our previous estimate on the entire retrospective cohort, we found that the risk of developing BC is 3.6-fold higher among ATM heterozygous women. An increased risk was associated with an early age at menarche, a late age at first childbirth, nulliparity, premenopausal status and increasing periods of breast cell mitotic activity (BCMA) prior to the first childbirth. Age at menarche, age at 1st childbirth and BCMA seemed to have a stronger effect in ATM heterozygotes than in non-ATM heterozygotes. However, the tests were not all statistically significant (only age at 1st childbirth). Surprisingly, the risk of BC decreased when the chest or breasts were irradiated. It is difficult to interpret the data because of the small sample size, but further investigations should provide a biological explanation for the variation in BC risk associated with exogenous factors according to ATM heterozygosis status.

Adolescent↗

PubMed2002

Distinct BRCA1 rearrangements involving the BRCA1 pseudogene suggest the existence of a recombination hot spot.

The 5' end of the breast and ovarian cancer-susceptibility gene BRCA1 has previously been shown to lie within a duplicated region of chromosome band 17q21. The duplicated region contains BRCA1 exons 1A, 1B, and 2 and their surrounding introns; as a result, a BRCA1 pseudogene (PsiBRCA1) lies upstream of BRCA1. However, the sequence of this segment remained essentially unknown. We needed this information to investigate at the nucleotide level the germline deletions comprising BRCA1 exons 1A, 1B, and 2, which we had previously identified in two families with breast and ovarian cancer. We have analyzed the recently deposited nucleotide sequence of the 1.0-Mb region upstream of BRCA1. We found that 14 blocks of homology between the tandemly repeated copies (cumulative length = 11.5 kb) show similarity of 77%-92%. Gaps between blocks result from insertion or deletion, usually of repetitive elements. BRCA1 exon 1A and PsiBRCA1 exon 1A are 44.5 kb apart. In the two families with breast and ovarian cancer mentioned above, distinct homologous recombination events occurred between intron 2 of BRCA1 and intron 2 of PsiBRCA1, leading to 37-kb deletions. Breakpoint junctions were found to be located at close but distinct sites within segments that are 98% identical. The mutant alleles lack the BRCA1 promoter and harbor a chimeric gene consisting of PsiBRCA1 exons 1A, 1B, and 2, which lacks the initiation codon, fused to BRCA1 exons 3-24. Thus, we report a new mutational mechanism for the BRCA1 gene. The presence of a large region homologous to BRCA1 on the same chromosome appears to constitute a hot spot for recombination.

Alleles↗

PubMed2002

A single mutated BRCA1 allele leads to impaired fidelity of double strand break end-joining.

Heterozygosity for mutations in the BRCA1 gene in humans confers high risk for developing breast cancer, but a biochemical basis for this phenotype has not yet been determined. Evidence has accumulated implicating BRCA1, in the maintenance of genomic integrity and the protection of cells against DNA double strand breaks (DSB). Here we present evidence that human cells heterozygous for BRCA1 mutations exhibit impaired DNA end-joining, which is the major DSB repair pathway in mammalian somatic cells. Using an in vivo host cell end-joining assay, we observed that the fidelity of DNA end-joining is strongly reduced in three BRCA1(+/-) cell lines in comparison to two control cell lines. Moreover, cell-free BRCA1(+/-) extracts are unable to promote accurate DNA end-joining in an in vitro reaction. The steady-state level of the wild type BRCA1 protein was significantly lower than the 50% expected in BRCA1(+/-) cells and thus may underlie the observed end-joining defect. Together, these data strongly suggest that BRCA1 is necessary for faithful rejoining of broken DNA ends and that a single mutated BRCA1 allele is sufficient to impair this process. This defect will compromise genomic stability in BRCA1 germ-line mutation carriers, triggering the genetic changes necessary for the initiation of neoplastic transformation.

Alleles↗

PubMed2002

Evaluation of linkage of breast cancer to the putative BRCA3 locus on chromosome 13q21 in 128 multiple case families from the Breast Cancer Linkage Consortium.

The known susceptibility genes for breast cancer, including BRCA1 and BRCA2, only account for a minority of the familial aggregation of the disease. A recent study of 77 multiple case breast cancer families from Scandinavia found evidence of linkage between the disease and polymorphic markers on chromosome 13q21. We have evaluated the contribution of this candidate "BRCA3" locus to breast cancer susceptibility in 128 high-risk breast cancer families of Western European ancestry with no identified BRCA1 or BRCA2 mutations. No evidence of linkage was found. The estimated proportion (alpha) of families linked to a susceptibility locus at D13S1308, the location estimated by Kainu et al. [(2000) Proc. Natl. Acad. Sci. USA 97, 9603-9608], was 0 (upper 95% confidence limit 0.13). Adjustment for possible bias due to selection of families on the basis of linkage evidence at BRCA2 did not materially alter this result (alpha = 0, upper 95% confidence limit 0.18). The proportion of linked families reported by Kainu et al. (0.65) is excluded with a high degree of confidence in our dataset [heterogeneity logarithm of odds (HLOD) at alpha = 0.65 was -11.0]. We conclude that, if a susceptibility gene does exist at this locus, it can only account for a small proportion of non-BRCA1/2 families with multiple cases of early-onset breast cancer.

Australia↗

PubMed2002

Rapid detection of novel BRCA1 rearrangements in high-risk breast-ovarian cancer families using multiplex PCR of short fluorescent fragments.

Recent studies have revealed a significant proportion of BRCA1 exon deletions or duplications in breast-ovarian cancer families with high probability of BRCA1- or BRCA2-linked predisposition, in which mutations of these genes have not been found. The difficulty of detecting such heterozygous rearrangements has stimulated the development of several new screening methods. Quantitative fluorescent multiplex PCR is based on simultaneous amplification of multiple target sequences under conditions that allow rapid and reliable quantitative comparison of the fluorescence of each amplicon in test samples and in controls. The modified method described here, named quantitative multiplex PCR of short fluorescent fragments (QMPSF), is particularly well suited for large genes. All BRCA1 coding exons were analyzed using four multiplexes in 52 families without point mutations in the exons or splice-sites of BRCA1 and BRCA2, and selected because of high probability of a BRCA1- or BRCA2-linked genetic predisposition. Five distinct BRCA1 rearrangements were detected: a deletion of exons 8-13, a duplication of exons 3-8, a duplication of exons 18-20, a deletion of exons 15-16, and a deletion of exons 1-22-which is the largest deletion found so far within the BRCA1 gene. The method described here lends itself to rapid, sensitive, and cost-effective search of BRCA1 rearrangements and may be included into the routine molecular analysis of breast-ovarian cancer predispositions. Hum Mutat 20:218-226, 2002.

BRCA1 Protein↗

PubMed2002

Effects of genetic consultation on perception of a family risk of breast/ovarian cancer and determinants of inaccurate perception after the consultation.

The aim of this study was to assess the effects of cancer genetic consultations on women's perception of their family risk of breast/ovarian cancer, and to determine which factors were associated with an inaccurate perception after the consultation. A multicenter prospective survey was carried out on women (n = 397) attending cancer genetic clinics in France for the first time, in which the perceived family risk was measured both before and after the consultation, using self-administered questionnaires. The effects of the consultation on risk perception were significant among low (P <.001) and moderate risk women (P <.05). However, after the consultation, 76.3% of the "low"-risk women did not perceive their family as "low"-risk families, and 21.9% of the moderate-risk women were still definitely sure there was a genetic risk running in their family. The consultation did not affect the family risk perception of the high risk women (n = 171): the risk was thought to be very high both before (87.7%) and after (89.5%) the consultation (NS); however 10.5% of this group still perceived their family as being unlikely to be at risk after the consultation. In the low- and moderate-risk groups after multivariate adjustment, the inaccurate perceptions varied, depending on the clinics and on the psychosocial context of the consultation: they increased when the consultee was personally affected by cancer, and decreased when the consultee had a health occupation. Cancer genetic consultations had only marginal effects on the perception of family risk on the whole, although they were significant in the case of low- and moderate-risk women. The question arises as to whether a more comprehensive approach should be implemented and how to go about providing efficient cancer risk information in the context of health care systems.

Adult↗

PubMed2002

Testing for BRCA1 mutations: a cost-effectiveness analysis.

Breast carcinoma is the most common type of cancer affecting women in the Western world. The hereditary forms, which amount from 5 to 10% of all the cases of breast cancer, mainly involve BRCA1 or BRCA2 mutations. Due to the diagnostic strategy used by the patent owner, Direct DNA sequencing (DS) may become the only BRCA1/2 test procedure available, although there exist several alternative strategies. A cost-effectiveness study was carried out using BRCA1 testing as a model. The main techniques available for performing mutation searches were assessed: DS, denaturing high performance liquid chromatography (DHPLC), single-strand conformation polymorphism (SSCP), denaturing gradient gel electrophoresis (DGGE), heteroduplex analysis (HA), fluorescent assisted mismatch analysis (FAMA) and the protein truncation test (PTT). Twenty strategies involving the use of one or more techniques were then devised for performing the complete genetic analysis. DS was adopted as the 'gold standard' for effectiveness. All the strategies except for DS involved a two-step procedure. The first step consisted of pre-screening the 22 coding exons of BRCA1. The second step consisted of performing DS only on the variations detected in the coding sequence. The cost of the strategies tested, including a pre-screening stage, turned out to be 30 to 90% lower than that of DS, whatever annual use was made of the equipment. The most cost-effective strategy, ie, that corresponding to the lowest cost per mutation detected, was found to be a combination between PTT on exon 11 (60% of the coding sequence) and HA on the remaining 21 exons (PTT(11)+ HA(21)). However, since a high false negative rate is associated with this strategy, at least four other strategies are worth mentioning: PTT(11)+ DHPLC(21), DHPLC alone, FAMA(11)+ DHPLC(21) and FAMA alone. Our results on genetic testing for breast cancer show that DS is not the most cost-effective method available. The monopolist approach of the firm which owns the patents on the BRCA1/2 genes, may, therefore limit the use of the most cost-effective strategies.

Cost-Benefit Analysis↗

PubMed2002

[CHEK2 and breast cancer risk].

The BRCA1 et BRCA2 genes are involved in 2/3 of genetic predisposition with major risk of breast cancer. One or more genes remain to be identified. The CHEK2 gene is a good candidate. The CHEK2 gene mutation 1100delC is associated with a moderate increase of breast cancer risk (RR = 2). CHEK2 is probably one of the genetic factors associated with moderate risk, but however when associated with other variants, it could explain some familial breast cancer and sporadic cancer cases. It seems to be too early to include CHEK2 in genetic counselling at the present time.

Breast Neoplasms↗

PubMedDate not supplied

[Opposition to Myriad Genetics patents and their total or partial revocation in Europe: early conclusions].

The proceedings instituted against three European patents held by the US company Myriad Genetics, on the BRCA1 gene and the breast cancer diagnosis gene, resulted in the total or partial revocation of these patents. These decisions put an end to the legal monopoly claimed by Myriad Genetics on the BRCA1 gene and on breast cancer gene tests, and left the field open to European geneticists to develop and implement their test methods within the framework of a clinical not-for-profit organization. The opposition procedure, through which any actor is allowed to challenge European patents, was used by geneticists doctors in Europe to refuse the emergence of an industrial monopoly on a medical service offered in a clinical context. The decision to revoke or strongly limit these patents was based on the European Patent Office's refusal to establish an invention priority on a sequence that had errors at the time the application was filed by the patent holder, in September 1994. The patent holder was granted an invention priority only on 24 March 1995, when it filed an application for a corrected sequence of the gene. But by then the BRCA1 gene sequence had already been divulged in a public data base, Genbank, from October 1994, notably by Myriad. Myriad Genetics' patents were thus victims of the patent race that prompted the firm to file multiple patent applications on insufficiently validated sequences, and of the conflict between diffusion in the public domain and the novelty requirement. Opposition to the patents, undertaken by a coalition of medical institutions, human genetic societies, two States, Holland and Austria, an environmental protection organization (Greenpeace), and the Swiss Labour Party, made it possible to preserve and develop the clinical economy of genetic tests in Europe. It resulted in amendments to intellectual property laws in France and thus extended the possibility of using compulsory licences for public health purposes to in vitro diagnosis.

Animals↗

PubMedDate not supplied

Lack of HIN-1 methylation defines specific breast tumor subtypes including medullary carcinoma of the breast and BRCA1-linked tumors.

Medullary carcinoma is a poorly differentiated breast cancer with a high histologic grade and a paradoxically good prognosis. It accounts for only 3 percent of all breast cancers except in BRCA-1 families, in which it can account for as many as 13 percent of cancers. To date, only histologic criteria have been used to define this tumor type. In an attempt to more clearly define the genetic pathway leading to this subtype of cancer, we recently demonstrated that nearly 100 percent of these carcinomas display p53 mutations. In the present study, we extended our analysis to include HIN-1, a candidate tumor suppressor that has been shown to be silenced by methylation in the majority of breast tumors. In striking contrast to unselected sporadic invasive ductal carcinoma, we show that medullary carcinomas do not display a high frequency of HIN-1 methylation (p less than 0.001). This feature is also found in BRCA-1 associated tumors that shared several histologic characteristics with medullary carcinomas of the breast. Medullary carcinoma of the breast should therefore be considered to be a unique entity defined by specific histologic and molecular traits.

Breast Neoplasms↗