8 research outputs found
Origin and distribution of the BRCA2-8765delAG mutation in breast cancer
Background: The BRCA2-8765delAG mutation was firstly described in breast cancer families from
French-Canadian and Jewish-Yemenite populations; it was then reported as a founder mutation in
Sardinian families. We evaluated both the prevalence of the BRCA2-8765delAG variant in Sardinia
and the putative existence of a common ancestral origin through a haplotype analysis of breast
cancer family members carrying such a mutation.
Methods: Eight polymorphic microsatellite markers (D13S1250, centromeric, to D13S267,
telomeric) spanning the BRCA2 gene locus were used for the haplotype analysis. Screening for the
8765delAG mutation was performed by PCR-based amplification of BRCA2-exon 20, followed by
automated sequencing.
Results: Among families with high recurrence of breast cancer (≥ 3 cases in first-degree relatives),
those from North Sardinia shared the same haplotype whereas the families from French Canadian
and Jewish-Yemenite populations presented distinct genetic assets at the BRCA2 locus. Screening
for the BRCA2-8765delAG variant among unselected and consecutively-collected breast cancer
patients originating from the entire Sardinia revealed that such a mutation is present in the
northern part of the island only [9/648 (1.4%) among cases from North Sardinia versus 0/493
among cases from South Sardinia].
Conclusion: The BRCA2-8765delAG has an independent origin in geographically and ethnically
distinct populations, acting as a founder mutation in North but not in South Sardinia. Since BRCA2-
8765delAG occurs within a triplet repeat sequence of AGAGAG, our study further confirmed the
existence of a mutational hot-spot at this genomic position (additional genetic factors within each
single population might be involved in generating such a mutation)
Microsatellite instability and mutation analysis of candidate genes in unselected Sardinian patients with endometrial carcinoma
BACKGROUND. Microsatellite instability (MSI) is due mostly to a defective DNA
mismatch repair (MMR). Inactivation of the two principal MMR genes, hMSH1and
hMSH2, and the PTEN tumor suppressor gene seems to be involved in endometrial
tumorigenesis. In this study, Sardinian patients with endometrial carcinoma (EC)
were analyzed to assess the prevalence of both the mutator phenotype (as defined
by the presence of MSI and abnormal MMR gene expression at the somatic level)
and the hMSH2, hMSH1, and PTEN germline mutations among patients with MSI
positive EC.
METHODS. Paraffin embedded tissue samples from 116 consecutive patients with
EC were screened for MSI by polymerase chain reaction-based microsatellite
analysis. Immunohistochemistry (IHC) with anti-hMLH1 and anti-hMSH2 antibodies
was performed on MSI positive tumor tissue sections. Germline DNA was
used for mutational screening by denaturing high-performance liquid chromatography
analysis and automated sequencing.
RESULTS. Thirty-nine patients with EC (34%) exhibited MSI; among them, 25 tumor
samples (64%) showed negative immunostaining for hMLH1/hMSH2 proteins
(referred to as IHC negative). No disease-causing mutation within the coding
sequences of the hMSH1/hMSH2 and PTEN genes was found in patients with EC
who had the mutator phenotype (MSI positive and IHC negative), except for a
newly described hMSH1 missense mutation, Ile655Val, that was observed in 1 of 27
patients (4%). Although MSI was more common among patients with advancedstage
EC and increased as the tumor grade increased, no significant correlation
with disease free survival or overall survival was observed among the two groups
(MSI positive or MSI negative) of patients with EC.
CONCLUSIONS. In patients with MSI positive EC, epigenetic inactivations rather
than genetic mutations of the MMR genes seem to be involved in endometrial
tumorigenesis. No prognostic value was demonstrated for MSI in patients with EC
High-resolution methylation analysis of the hMLH1 promoter in sporadic endometrial and colorectal carcinomas
BACKGROUND. Microsatellite instability (MSI) has been reported in endometrial
carcinoma (EC) and in colorectal carcinoma (CRC), primarily as a result of defective
DNA mismatch repair (MMR). The MMR gene hMLH1 commonly is inactivated
in both EC and CRC. In the current study, epigenetic mechanisms involved
in hMLH1 inactivation have been investigated to further elucidate the role of these
mechanisms in the pathogenesis of EC and CRC.
METHODS. Polymerase chain reaction (PCR)-based microsatellite analysis performed
on paraffin-embedded tissues was used to select 42 sporadic carcinomas
(21 ECs and 21 CRCs) with MSI. Immunohistochemistry (IHC), using the antihMLH1
antibody, and mutation analysis, using denaturing high-performance liquid
chromatography and automated sequencing, were performed on unstable
carcinoma samples. Methylation analysis, using modified protocols for bisulfite
treatment and methylation-specific PCR (MSP), was performed on DNA from
archival tissue samples.
RESULTS. No MSI-positive tumor samples with normal hMLH1 immunostaining (π =
7) exhibited hMLH1 promoter methylation, whereas 8 of 35 unstable cases with
loss of hMLH1 expression (23%) exhibited MSP amplification. Among analyzed
cases, germ-line mutations of hMLH1 were found in 4 of 20 unmethylated samples
(20%) and in 0 of 8 methylated samples. Bisulfite sequencing of amplification
products from methylated samples demonstrated that almost all CpG dinucleotides
within the hMLH1 promoter elements underwent methylation.
CONCLUSIONS. Although an MMR gene other than hMLH1 may be responsible for
genetic instability in MSI-positive/IHC-positive tumors, the presence of MSP amplification
and allelic deletions within the hMLH1 locus in subsets of MSI-positive/
IHC-negative cases strongly suggests that hMLH1 promoter methylation may contribute
to the inactivation of both hMLH1 alleles. Bisulfite analysis suggests that the
mechanisms of hMLH1 silencing may depend on CpG density rather than sitespecific
methylation
Spectrum and prevalence of <i>BRCA</i>1 and <i>BRCA</i>2 germline mutations in Sardinian patients with breast carcinoma through hospital-based screening
BACKGROUND.Factors that are predictive of carrying BRCA1 and BRCA2 germline mutations in patients with breast carcinoma are awaited widely. The genetically homogeneous Sardinian population may be useful for defining the role of such genetic alterations further through a clinical evaluation program.
METHODS. One hundred two of 659 patients with breast carcinoma (15.5%) who were collected consecutively had a family history of breast carcinoma and were screened for BRCA1/2 mutations by denaturing high-performance liquid chromatography and DNA sequencing.
RESULTS. Three deleterious germline BRCA1/2 mutations were detected in 15 of 102 families (14.7%), including 13 families (86.7%) with BRCA2 mutations and 2 families (13.3%) with BRCA1 mutations. A single variant, BRCA2-8765delAG, was the most recurrent mutation in the series and was found in 12 of 102 families (11.8%) and in 18 of 657 patients (2.7%). The average age at diagnosis was significantly younger in families with BRCA1/2 mutations (48.6 yrs) compared with the age of patients who had no detectable mutation (52.9 yrs; P 0.039). Moreover, BRCA1/2 mutations were found at a significantly higher rate in families who had at least 1 member with ovarian carcinoma or male breast carcinoma (5 of 12 families; 41.7%) than in families without such an association (10 of 90 families; 11.1%; P 0.003).
CONCLUSIONS. BRCA2 mutations were approximately 6 times more prevalent than BRCA1 mutations. A diagnosis of breast carcinoma before age 50 years, ovarian carcinoma, male breast carcinoma, and 3 affected generations all were associated significantly with BRCA1/2 mutations. Although the current findings provided further support for the hypothesis that additional breast carcinoma susceptibility genes remain to be identified, such indicators of the presence of BRCA1/2 mutations may be useful in counseling patients about undergoing genetic testing.</br
Origin and distribution of the BRCA2-8765delAG mutation in breast cancer
Abstract Background The BRCA2-8765delAG mutation was firstly described in breast cancer families from French-Canadian and Jewish-Yemenite populations; it was then reported as a founder mutation in Sardinian families. We evaluated both the prevalence of the BRCA2-8765delAG variant in Sardinia and the putative existence of a common ancestral origin through a haplotype analysis of breast cancer family members carrying such a mutation. Methods Eight polymorphic microsatellite markers (D13S1250, centromeric, to D13S267, telomeric) spanning the BRCA2 gene locus were used for the haplotype analysis. Screening for the 8765delAG mutation was performed by PCR-based amplification of BRCA2-exon 20, followed by automated sequencing. Results Among families with high recurrence of breast cancer (≥ 3 cases in first-degree relatives), those from North Sardinia shared the same haplotype whereas the families from French Canadian and Jewish-Yemenite populations presented distinct genetic assets at the BRCA2 locus. Screening for the BRCA2-8765delAG variant among unselected and consecutively-collected breast cancer patients originating from the entire Sardinia revealed that such a mutation is present in the northern part of the island only [9/648 (1.4%) among cases from North Sardinia versus 0/493 among cases from South Sardinia]. Conclusion The BRCA2-8765delAG has an independent origin in geographically and ethnically distinct populations, acting as a founder mutation in North but not in South Sardinia. Since BRCA2-8765delAG occurs within a triplet repeat sequence of AGAGAG, our study further confirmed the existence of a mutational hot-spot at this genomic position (additional genetic factors within each single population might be involved in generating such a mutation).</p
Identification of a novel candidate gene, CASC2, in a region of common allelic loss at chromosome 10q26 in human endometrial cancer
Allelic deletions, which are suggestive for the presence of tumor suppressor genes, represent a common event in endometrial cancer (EC). Previous loss-of-heterozygosity studies for human chromosome 10q identified a candidate deletion interval at 10q25-q26, which we further narrowed to a 160-kb region at 10q26, bounded by markers D10S1236 and WIAF3299. Using a positional candidate approach, we identified three alternative transcripts of a novel human gene, CASC2 (cancer susceptibility candidate 2; formely C10orf5). One of such transcripts, CASC2a, encodes a short protein of 102 amino acids with no similarity to any other known gene product. Three (7%) CASC2a mutations were identified in tumor DNA from 44 EC patients. While c.-156G>T and c.22C>T (p.Pro8Ser) are sequence variants with unknown functional significance, c.84delA is a mutation with a truncation effect on the predicted protein (p. Asn28fsX50). Expression studies by real-time RT-PCR on several normal and tumor cells revealed that CASC2a mRNA is downregulated in cancer, suggesting that it may act as a potential tumor suppressor gene. The very low mutation rate seems to also indicate that inactivation of CASC2a might probably be due to mechanisms different from genetic alteration
Molecular alterations at chromosome 9p21 in melanocytic naevi and melanoma
Background The chromosome 9p21 and its CDKN locus, with the p16 tumour suppressor gene (CDKN2A), are recognized as the genomic regions involved in the pathogenesis of melanoma.
Objectives To elucidate further the role of such regions during the different phases of melanocytic tumorigenesis.
Methods Tissue sections from naevi, primary and metastatic melanomas were investigated by fluorescence in situ hybridization for allelic loss at the 9p21 chromosome and by immunochemistry for p16CDKN2A expression.
Results Dysplastic naevi and primary or secondary melanomas were found to carry hemizygous deletions within the entire 9p21 region at similar frequencies (varying from 55% to 62%). Allelic deletion spanning the CDKN locus was observed at significantly increased rates moving from early (7%) to advanced (28%) primary melanomas and to secondary melanoma lesions (37%) (P = 0·018). Also, inactivation of the p16 gene (CDKN2A) was absent in naevi and present at steadily increasing rates moving from primary melanomas (7% early lesions to 17% advanced lesions) to melanoma metastases (62%) (P = 0·004).
Conclusions Our findings indicate that, in a model of sequential accumulation of genetic alterations, 9p21 deletions may play a role in melanocytic transformation and tumour initiation whereas rearrangements at the CDKN locus, and p16 gene (CDKN2A) inactivation may contribute to tumour progression