Association of PHB 1630 C>T and MTHFR 677 C>T polymorphisms with breast and ovarian cancer risk in BRCA1/2 mutation carriers: results from a multicenter study.

BACKGROUND: The variable penetrance of breast cancer in BRCA1/2 mutation carriers suggests that other genetic or environmental factors modify breast cancer risk. Two genes of special interest are prohibitin (PHB) and methylene-tetrahydrofolate reductase (MTHFR), both of which are important either directly or indirectly in maintaining genomic integrity. METHODS: To evaluate the potential role of genetic variants within PHB and MTHFR in breast and ovarian cancer risk, 4102 BRCA1 and 2093 BRCA2 mutation carriers, and 6211 BRCA1 and 2902 BRCA2 carriers from the Consortium of Investigators of Modifiers of BRCA1 and BRCA2 (CIMBA) were genotyped for the PHB 1630 C>T (rs6917) polymorphism and the MTHFR 677 C>T (rs1801133) polymorphism, respectively. RESULTS: There was no evidence of association between the PHB 1630 C>T and MTHFR 677 C>T polymorphisms with either disease for BRCA1 or BRCA2 mutation carriers when breast and ovarian cancer associations were evaluated separately. Analysis that evaluated associations for breast and ovarian cancer simultaneously showed some evidence that BRCA1 mutation carriers who had the rare homozygote genotype (TT) of the PHB 1630 C>T polymorphism were at increased risk of both breast and ovarian cancer (HR 1.50, 95%CI 1.10-2.04 and HR 2.16, 95%CI 1.24-3.76, respectively). However, there was no evidence of association under a multiplicative model for the effect of each minor allele. CONCLUSION: The PHB 1630TT genotype may modify breast and ovarian cancer risks in BRCA1 mutation carriers. This association need to be evaluated in larger series of BRCA1 mutation carriers.

The TP53 Arg72Pro and MDM2 309G>T polymorphisms are not associated with breast cancer risk in BRCA1 and BRCA2 mutation carriers.

BACKGROUND: The TP53 pathway, in which TP53 and its negative regulator MDM2 are the central elements, has an important role in carcinogenesis, particularly in BRCA1- and BRCA2-mediated carcinogenesis. A single nucleotide polymorphism (SNP) in the promoter region of MDM2 (309T>G, rs2279744) and a coding SNP of TP53 (Arg72Pro, rs1042522) have been shown to be of functional significance. METHODS: To investigate whether these SNPs modify breast cancer risk for BRCA1 and BRCA2 mutation carriers, we pooled genotype data on the TP53 Arg72Pro SNP in 7011 mutation carriers and on the MDM2 309T>G SNP in 2222 mutation carriers from the Consortium of Investigators of Modifiers of BRCA1/2 (CIMBA). Data were analysed using a Cox proportional hazards model within a retrospective likelihood framework. RESULTS: No association was found between these SNPs and breast cancer risk for BRCA1 (TP53: per-allele hazard ratio (HR)=1.01, 95% confidence interval (CI): 0.93-1.10, P(trend)=0.77; MDM2: HR=0.96, 95%CI: 0.84-1.09, P(trend)=0.54) or for BRCA2 mutation carriers (TP53: HR=0.99, 95%CI: 0.87-1.12, P(trend)=0.83; MDM2: HR=0.98, 95%CI: 0.80-1.21, P(trend)=0.88). We also evaluated the potential combined effects of both SNPs on breast cancer risk, however, none of their combined genotypes showed any evidence of association. CONCLUSION: There was no evidence that TP53 Arg72Pro or MDM2 309T>G, either singly or in combination, influence breast cancer risk in BRCA1 or BRCA2 mutation carriers.

The contribution of germline rearrangements to the spectrum of BRCA2 mutations.

BACKGROUND: Few germline BRCA2 rearrangements have been described compared with the large number of germline rearrangements reported in the BRCA1 gene. However, some BRCA2 rearrangements have been reported in families that included at least one case of male breast cancer. OBJECTIVE: To estimate the contribution of large genomic rearrangements to the spectrum of BRCA2 defects. METHODS: Quantitative multiplex PCR of short fluorescent fragments (QMPSF) was used to screen the BRCA2 gene for germline rearrangements in highly selected families. QMPSF was previously used to detect heterozygous deletions/duplications in many genes including BRCA1 and BRCA2. RESULTS: We selected a subgroup of 194 high risk families with four or more breast cancers with an average age at diagnosis of < or = 50 years, who were recruited through 14 genetic counselling centres in France and one centre in Switzerland. BRCA2 mutations were detected in 18.6% (36 index cases) and BRCA1 mutations in 12.4% (24 index cases) of these families. Of the 134 BRCA1/2 negative index cases in this subgroup, 120 were screened for large rearrangements of BRCA2 using QMPSF. Novel and distinct BRCA2 deletions were detected in three families and their boundaries were determined. We found that genomic rearrangements represent 7.7% (95% confidence interval 0% to 16%) of the BRCA2 mutation spectrum. CONCLUSION: The molecular diagnosis of breast cancer predisposition should include screening for BRCA2 rearrangements, at least in families with a high probability of BRCA2 defects.

Does nonsense-mediated mRNA decay explain the ovarian cancer cluster region of the BRCA2 gene?

BRCA2 (BReast CAncer susceptibility gene 2) germline mutation carriers are at increased risk for breast and ovarian cancers. Mutations occurring in the ovarian cancer cluster region (OCCR) are linked to higher ovarian cancer and/or lower breast cancer risk(s) than mutations occurring elsewhere in BRCA2. Most BRCA2 germline mutations introduce premature termination codons (PTCs), making their mRNAs likely targets of nonsense-mediated mRNA decay (NMD), a mechanism that eliminates PTC-bearing transcripts to prevent expression of truncated proteins. Contradictory evidence exists regarding whether NMD can be triggered by PTCs located far upstream of the nearest exon-exon junction (EEJ). Since the OCCR comprises a major portion of the 4.9 kb exon 11 of BRCA2, we investigated if transcripts bearing PTCs in this large exon are unable to trigger NMD, and if this might contribute to the phenotypic difference associated with the OCCR. We examined cDNA from 18 carriers of PTC-introducing germline mutations located throughout BRCA2, and found that PTC-bearing transcripts were 1.4-3.3-fold less prevalent than their nonmutated counterparts irregardless of PTC position. We conclude that NMD can recognize PTCs up to 4.5 kb upstream of the nearest EEJ, demonstrating that a general inability of NMD to recognize PTCs in exon 11 is unlikely to explain the genotype-phenotype correlation associated with the OCCR.

Ratio of male to female births in the offspring of BRCA1 and BRCA2 carriers.

A recent report based on 68 families, including 17 with mutations in BRCA1, suggested that there was an excess of female offspring born to BRCA1 mutation carriers. We have examined the gender ratio among offspring of 511 mutation carriers from 116 BRCA1 families, 77 and 39 from Australia and the United States, respectively. We found no evidence for a significant deviation from the expected proportion of female offspring in the Australian pedigrees, but there was an excess of female offspring in pedigrees from the USA. Ascertainment bias probably explains this bias, rather than a link with X-chromosome inactivation as previously suggested, because the families from the USA were ascertained for the purposes of linkage studies whereas those from Australia were ascertained through Familial Cancer Clinics to which they had been referred for clinical genetic counseling and mutation testing.

Germline brca2 sequence variants in patients with ocular melanoma.

On the basis, chiefly, of anecdotal reports of cases of ocular melanoma (OM) occurring in families with inherited susceptibility to breast cancer due to brca2 germline mutations, we examined the frequency of brca2 alterations in a series of 62 ocular melanoma cases. These cases were preferentially selected on the basis of reported family history of breast or ovarian cancer, or OM, although the series also included a randomly selected set of cases without family history of cancer. A total of 7 germline alterations were found, of which 3 were likely to be associated with disease. While all 3 deleterious mutations were found in patients who also had a personal history of breast cancer, only 1 of the 3 families had a family history of breast/ovarian cancer or OM. Although germline brca2 mutations may account for a small proportion of all OM cases, there may be additional loci that contribute to familial aggregation of OM and to the familial association between OM and breast cancer.

Regulation of dauer larva development in Caenorhabditis elegans by daf-18, a homologue of the tumour suppressor PTEN.

The tumour suppressor gene PTEN (also called MMAC1 or TEP1) is somatically mutated in a variety of cancer types [1] [2] [3] [4]. In addition, germline mutation of PTEN is responsible for two dominantly inherited, related cancer syndromes called Cowden disease and Bannayan-Ruvalcaba-Riley syndrome [4]. PTEN encodes a dual-specificity phosphatase that inhibits cell spreading and migration partly by inhibiting integrin-mediated signalling [5] [6] [7]. Furthermore, PTEN regulates the levels of phosphatidylinositol 3,4,5-trisphosphate (PIP3) by specifically dephosphorylating position 3 on the inositol ring [8]. We report here that the dauer formation gene daf-18 is the Caenorhabditis elegans homologue of PTEN. DAF-18 is a component of the insulin-like signalling pathway controlling entry into diapause and adult longevity that is regulated by the DAF-2 receptor tyrosine kinase and the AGE-1 PI 3-kinase [9]. Others have shown that mutation of daf-18 suppresses the life extension and constitutive dauer formation associated with daf-2 or age-1 mutants. Similarly, we show that inactivation of daf-18 by RNA-mediated interference mimics this suppression, and that a wild-type daf-18 transgene rescues the dauer defect. These results indicate that PTEN/daf-18 antagonizes the DAF-2-AGE-1 pathway, perhaps by catalyzing dephosphorylation of the PIP3 generated by AGE-1. These data further support the notion that mutations of PTEN contribute to the development of human neoplasia through an aberrant activation of the PI 3-kinase signalling cascade.

Screening for germ-line rearrangements and regulatory mutations in BRCA1 led to the identification of four new deletions.

Most previous BRCA1 mutation screening studies conducted on breast cancer families were aimed at identifying mutations in the coding sequence and splice sites. Mutations in the promoter and untranslated regions, and large rearrangements are missed by standard mutation detection strategies. To look specifically for such germ-line mutations in the BRCA1 gene, we have analyzed a series of 27 American and 51 French breast cancer families in which no BRCA1 mutation was identified by classical techniques. No mutations were detected in either the promoter or untranslated regions, and we did not find any deletion of the whole gene. Four families were found to carry distinct deletions. Two of them, probably generated by Alu-mediated homologous recombination, were internal deletions of 3 and 23.8 kb, encompassing exon 15 and exons 8-13, respectively. These alterations both lead to a frameshift in the mutant mRNA and to premature stop codon-mediated mRNA decay. The other two deletions encompass exons 1 and 2. On the basis of previous and present analyses, rearrangements represent 8% (3/37) of all mutations in our set of BRCA1 American families. Consequently, the search for rearrangements appears mandatory in BRCA1 mutation screening studies.

A 1-kb Alu-mediated germ-line deletion removing BRCA1 exon 17.

Although more than 100 different BRCA1 germ-line mutations have already been identified in breast and/or ovarian cancer families, we report for the first time a deleterious genomic rearrangement in BRCA1. A 1-kb deletion comprising exon 17 was found in a large breast and ovarian cancer family, leading to a frameshift in the mutant mRNA due to the absence of exon 17. This deletion is probably the result of a recombination between two closely related Alu sequences. It was not detected by conventional PCR-based methods involving the genomic screening of the 22 coding exons or reverse transcription-PCR because the transcript without exon 17 is unstable in lymphoblastoid cell lines. Therefore, rearrangements in the BRCA1 gene should be sought in breast/ovarian cancer families in which no mutations have been found by PCR-based methods in the coding region or in the splice sites.