[show abstract][hide abstract] ABSTRACT: Mismatch repair mutations are the cause of generalized genomic instability and are particularly evident at microsatellite loci, which is known as microsatellite instability (MSI). MSI is present in 85% to 90% of colorectal cancers and occurs in hereditary non-polyposis colorectal cancer (HNPCC). The National Cancer Institute recommends the "Bethesda panel" for MSI screening. Recently, a novel T(25) mononucleotide marker was described, termed CAT25. This microsatellite marker displays a quasi-monomorphic pattern in normal tissues. The aim of our study was to evaluate the performance of CAT25 in HNPCC patients and to compare its reliability with the results of the Bethesda panel. We tested 55 tumor tissues from HNPCC patients using both the Bethesda panel and the CAT25 mononucleotide marker. One hundred healthy blood donors were used as controls. The CAT25 microsatellite was found to be altered in all 13 colorectal cancers classified as MSI-H using the standard Bethesda panel. Colorectal tumors that showed a stable Bethesda pattern did not show altered CAT25 repeats. Additionally, CAT25 showed a monomorphic allele pattern in all tissue samples. In our series, the concordance between the Bethesda panel and CAT25 in identifying colorectal cancers with high MSI reached 100%. Our results suggest that the CAT25 microsatellite represents a sensitive and specific marker for MSI and could be, at least, included in the panel of markers for the identification of HNPCC patients.
The Journal of molecular diagnostics: JMD 04/2009; 11(3):248-52. · 3.48 Impact Factor
[show abstract][hide abstract] ABSTRACT: Hereditary Non-Polyposis Colorectal Cancer (HNPCC) is associated with germline mutations in one of several MisMatch Repair (MMR) genes. An increasing proportion (20-25%) of the reported MSH2 variants consists of single amino-acid substitution with uncertain disease-causing significance. The present study was undertaken to functionally characterize 3 MSH2 nontruncating variants: p.Gly162Arg (c.484G>C), p.Asp167His (c.499G>C) and p.Arg359Ser (c.1077A>T). Missense alterations, were assessed in a human system for expression/stability and for the ability to heterodimerize with MSH6 and correctly localize into the nucleus. Functional assays results were correlated with clinical and genetic features indicative of HNPCC as MicroSatellite-Instability (MSI), abnormalities of MMR gene expression in tumour tissue (IHC) and familial history. p.Gly162Arg and p.Arg359Ser variants showed a clearly decreased expression level of the MutSá complex and were associated with an abnormal subcellular localization pattern, which can be suggestive of an incorrect MSH2/MSH6 heterodimerization. Functional analysis results were supported by MSI and IHC data and by familial cancer history. The subcellular localization assay, performed in a human expression system, classifies as pathogenetic two MSH2 nontruncating alterations providing a useful tool in genetic testing programs.
Human Mutation 09/2008; 29(11):E296-309. · 5.21 Impact Factor
[show abstract][hide abstract] ABSTRACT: The prognostic significance of KIT or platelet-derived growth factor receptor alpha (PDGFRalpha) mutations in gastrointestinal stromal tumors (GISTs) is still controversial.
In all, 104 patients were diagnosed with GISTs by KIT immunoreactivity; tumor DNA was sequenced for the presence of mutations in KIT exons 9, 11, 13 and 17 and in PDGFRalpha exons 12 and 18. Disease-free survival (DFS) was analyzed in 85 radically resected patients.
KIT mutations occurred in exon 11 (69), in exon 9 (11) and in exon 17 (1). PDGFRalpha mutations were detected in exon 18 (10) and in exon 12 (3). Ten GISTs were wild type. Exon 11 mutations were as follows: deletions in 42 cases and point mutations in 20 cases and insertions and duplications, respectively, in 2 and 5 cases. A better trend in DFS was evident for duplicated and point-mutated exon 11 KIT GISTs. There was a significant association between PDGFRalpha mutations, gastric location and lower mitotic index. Moreover, PDGFRalpha-mutated GISTs seemed to have a better outcome.
Point mutations and duplications in KIT exon 11 are associated with a better clinical trend in DFS. PDGFRalpha-mutated GISTs are preferentially localized in the stomach and seem to have a favorable clinical behavior.
Annals of Oncology 05/2008; 19(4):706-10. · 7.38 Impact Factor
[show abstract][hide abstract] ABSTRACT: Subjects affected by hereditary non-polyposis colorectal cancer exhibit a high susceptibility to colon and extracolonic tumours, due to MMR gene defects. Revised Bethesda criteria are used to select patients as candidates for genetic tests. Recently, the CRCAPRO model has been developed, based on family history of colorectal and endometrial cancers. Our study aims to evaluate the reliability of CRCAPRO in identifying mutation carriers. We used the CRCAPRO program to evaluate carrier probability risk in 99 patients fulfilling Amsterdam or Bethesda guidelines. MLH1 and MSH2 were studied by direct sequencing in all the 99 patients, and the study of microsatellite instability and of MMR proteins expression was performed. Nine MLH1 and nine MSH2 germline mutations were identified. Five out of the nine patients with MLH1 mutation showed a CRCAPRO risk evaluation of less than 20%. The same happened for four out of nine patients with MSH2 mutation. Of the 17 patients with an estimated risk of more than 80%, only four harboured a mutation, all in the MSH2 gene. The highest risk calculated by the CRCAPRO system in the nine carriers of a MLH1 mutation has been 31.7%. In our experience, the CRCAPRO program sensitivity and specificity appears to be low but needs to be further evaluated in larger samples.
[show abstract][hide abstract] ABSTRACT: Hereditary Non-Polyposis Colorectal Cancer (HNPCC) is an autosomal dominant inherited disease predisposing to the development of colorectal cancers and several other malignancies (endometrium, ovaries, stomach, small bowel, hepatobiliary and urinary tract). HNPCC is caused by germline mutations in any of the MisMatch Repair (MMR) genes. Mutations in MLH1 and MSH2 account for almost 90% of all identified ones. About 15% of mutations identified in MSH2 are missense ones.
We studied one family, fulfilling Amsterdam II criteria, referred to our Center for genetic counselling. The proband, and some of her relatives, have been investigated for microsatellite instability (MSI), immunohistochemical MMR protein staining and by direct sequencing and Multiplex Ligation-dependent Probe Amplification (MLPA).
All patients carried the same novel MSH2 germline missense mutation (R359S) in exon 7, which determines the substitution of an Arginine, which is a basic amino acid, with a polar Serine residue (R359S). The mutation was associated with lack of expression of MSH2 protein and high microsatellite instability in tumour tissues. The same mutation has been detected in one healthy relative.
The mutation here reported shows a high correlation with phenotype. The mutation is located in an evolutionary conserved domain. Taken together, our findings suggest evidence that the amino acid substitution can be interpreted as pathogenetic.
Familial Cancer 02/2007; 6(1):97-102. · 1.94 Impact Factor
[show abstract][hide abstract] ABSTRACT: Hereditary nonpolyposis colorectal cancer (HNPCC) is an inherited syndrome of cancer susceptibility caused by germ line mutations of genes participating in mismatch repair (MMR). Carriers of MMR gene mutations have an increased risk of colorectal cancers and cancer of other organs. Tumors of the endometrium represent the most frequent extracolonic malignancies in HNPCC. It has been suggested that women harboring MMR gene mutations have a higher risk of endometrial cancer than of colon cancer. Here, we describe an HNPCC patient with early-onset endometrial cancer and a strong familial history of endometrial tumors who harbored a germ line MSH2 splice site mutation (IVS9_2A>G). This mutation was responsible for abnormal messenger RNA processing, leading to the introduction of a premature stop signal and to the expression of a truncated MSH2 protein. In addition, the same mutation was associated with loss of MSH2 protein expression, high microsatellite instability, and PTEN inactivation. Although a direct relationship between the endometrial cancer susceptibility and the MSH2 mutation we found cannot be established, our observations, consistent with the work of other authors, suggest the involvement of germ line MSH2 abnormalities in endometrial tumor development and support the case for endometrial cancer screening in women from HNPCC families.
International Journal of Gynecological Cancer 06/2006; 16(3):1419-23. · 1.94 Impact Factor