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R Hamelin

Publications and source records attributed to R Hamelin.

At least 37 records · Page 2Linked to original sources
PubMed1999

Mutational analysis of the PTEN gene in gliomas: molecular and pathological correlations.

The PTEN gene, recently identified on chromosome 10q23, has been proposed to be a candidate tumor suppressor gene inactivated in multiple cancers including glial tumors. We investigated 47 glioblastomas (GBM), 14 anaplastic astrocytomas (AA), 6 non-pilocytic low-grade astrocytomas (LGA), 21 low-grade and anaplastic oligodendrogliomas (O) and oligoastrocytomas (OA), and 3 ependymomas (E) for mutation of the PTEN gene using denaturing gradient gel electrophoresis (DGGE) followed by DNA sequencing. These tumors have been previously screened for loss of heterozygosity (LOH) on chromosome 10q, p53 mutations and EGFR amplification. Overall, PTEN mutations, detected in 14 of 91 tumors, were present in 13 of 47 GBM and 1 of 14 AA. In contrast, mutations were absent in other glioma subtypes (0/30). In all informative cases, PTEN mutations occurred in tumors showing LOH on chromosome 10q, confirming the inactivation of this gene by a 2-hit mechanism. No correlation was observed between the presence of PTEN mutation and p53 mutation and EGFR amplification. Our results indicate that biallelic PTEN inactivation plays an important role in the pathogenesis of high-grade astrocytomas as a late event. Moreover, they suggest that PTEN alterations are equally involved in the 2 glioblastoma pathways defined by the presence of EGFR amplification and p53 mutation. Finally, correlation analysis with clinical data did not show that PTEN mutation was linked to survival of the patients.

Brain Neoplasms↗
PubMed1999

Gastric carcinomas with microsatellite instability: clinical features and mutations to the TGF-beta type II receptor, IGFII receptor, and BAX genes.

The replication error phenotype (RER+) represents an important new form of genetic alteration characterized by widespread instability in repetitive nucleotide sequences. The aim of this study was to compare the features of RER+ gastric tumours with those of RER+ colonic tumours. RER status was determined by analysis of size alterations in the BAT-26 mononucleotide repeat microsatellite. Twelve of 121 (10 per cent) gastric carcinomas from a low-incidence region were found to be RER+. BAT-26 instability was associated with tumours showing an absence of nodal invasion ( p=0.009) and with a trend for improved prognosis. These tumours were more frequent in older, female patients. Frameshift mutations in mononucleotide repeat sequences within the transforming growth factor-beta receptor II (RII), insulin-like growth factor II receptor (IGFIIR), and BAX genes were observed in 83, 33, and 25 per cent, respectively, of RER+ tumours. Only 1/12 (8 per cent) RER+ tumours contained a p53 gene mutation compared with 29/109 (27 per cent) RER- tumours. RER+ gastric carcinomas therefore share several important features with RER+ colonic tumours, including less frequent nodal invasion, improved prognosis, a similar frequency of mutation in growth control genes containing repetitive nucleotide sequences, and a low frequency of mutation of the p53 tumour suppressor gene.

Aged↗
PubMed1999

Germline mutations of p53 but not p16/CDKN2 or PTEN/MMAC1 tumor suppressor genes predispose to gliomas. The ANOCEF Group. Association des NeuroOncologues d'Expression Française.

Constitutional DNA from 44 selected patients suspected of being genetically predisposed to develop astrocytic tumors was analyzed for germline mutations of the p53, p16, and PTEN genes. Six constitutional missense mutations of the p53 gene were identified (13.6%), but no mutations of the p16 and PTEN genes were found, suggesting that (1) germline p53 mutations contribute to a small portion of astrocytic tumors, (2) inherited mutations of the p16 and PTEN gene do not predispose to the development of gliomas, and (3) other genes are involved in glioma predisposition.

Amino Acid Substitution↗
PubMed1999

TP53 status and gene amplification in human colorectal carcinomas.

Gene amplification is one of the characteristics of cancer cells. In vitro studies suggested that alterations of the TP53 gene might be responsible for gene amplification. We have examined the presence of TP53 mutations and looked for cytogenetic evidence of gene amplification in a series of 79 primary colorectal carcinomas. Other parameters such as the pattern of cytogenetic alterations, microsatellite instability, tumor site, and histological staging were also considered. A multiparametric study supported by statistical analyses suggests the existence of two major pathways of colorectal carcinogenesis. No relationships could be established between the presence of TP53 alterations and gene amplification.

Aged↗
PubMed1999

p53 abnormalities in adenocarcinoma of the gastric cardia and antrum.

AIM: To compare the frequency and type of p53 alterations (gene mutation and/or protein overexpression) in a consecutive series of surgically resected adenocarcinomas arising in the gastric cardia and gastric antrum, and to evaluate associations with clinicopathological findings (age, sex, and tumour histology, grade, and stage). METHODS: The series comprised 50 patients with adenocarcinoma of the cardia and 20 patients with adenocarcinoma of the antrum. p53 gene mutations (exons 5-8) were detected by denaturing gradient gel analysis and DNA sequencing. Nuclear p53 overexpression was detected by immunohistochemistry with the DO7 antibody. RESULTS: p53 gene mutations were found in 21 of 50 and five of 20 adenocarcinomas of the cardia and the antrum, respectively. Base transitions occurring at CpG dinucleotides were frequent in both types of tumour. p53 protein overexpression was seen in 32 of 50 and seven of 20 adenocarcinomas of the cardia and of the antrum, respectively. p53 gene mutation and/or protein overexpression were significantly more frequent in adenocarcinomas of the cardia (37 of 50) than in adenocarcinomas of the antrum (seven of 20). There were no differences in the clinicopathological characteristics of the tumours between p53 positive and p53 negative cases in both types of cancer. CONCLUSIONS: This study shows that p53 alterations are more frequent in adenocarcinoma of the cardia than in adenocarcinoma of the antrum. This feature is consistent with the clinical and epidemiological characteristics of these cancers, which suggest that adenocarcinoma arising in the gastric cardia might be related to oesophageal adenocarcinoma, and unrelated to adenocarcinomas of the gastric body and antrum.

Adenocarcinoma↗
PubMed1998

Quantification and characterization of total cellular p53 protein in colorectal cancer.

Immunochemical methods were developed for the optimal detection and characterization of total cellular p53 protein expression, both in the nuclear-attached and soluble fractions of colorectal cancers, in order to improve the correlation between protein deregulation and gene status. Seventy colorectal carcinomas were studied using 3 monoclonal antibodies in a sensitive analyzing system combining flow cytometry (nuclear-bound fraction) and enzyme-linked immunosorbent assay (ELISA; soluble fraction). DNA indices were calculated on the DNA histograms and mutations of the p53 gene were searched for in a subset of 41 cases. Three p53 expression patterns were found: 35 tumors were classified as pattern "A," characterized by high p53 expression including "mutant" conformation and missense mutations of the gene (16/17 cases tested), pattern "B" consisted of 15 tumors with total absence of p53 expression corresponding to nonsense mutations of the gene (8/9 cases tested), and pattern "C" of 20 tumors presenting low or undetectable nuclear-bound p53 but intermediate p53 protein content (pAb (1801+) in the soluble fraction. The latter pattern was associated with wild-type genes (14/15 cases tested), and with tumors that were often localized in the right colon compared to pattern "A" and "B" tumors (45% versus 8%, P < 0.009) and were frequently near-diploid (80% versus 29%, P < 0.0002). No correlation was found between tumor stage and the patterns of p53 expression. The results indicate that both flow cytometry (FCM) and ELISA seem necessary for the proper characterization of the p53 expression pattern, thus achieving a high degree of concordance with molecular analysis of gene mutations.

Adenoma↗
PubMed1998

Mutation of the transforming growth factor-beta type II receptor gene in right-sided colorectal cancer: relationship to clinicopathological features and genetic alterations.

The presence of inactivating mutations in the transforming growth factor-beta (TGF-beta) type II receptor (RII) gene in the colon cancer suggests that it may behave like a tumour suppressor gene. RII is mutated in the majority of colon tumours exhibiting widespread microsatellite instability, a characteristic generally referred to as the replication error phenotype (RER+). We investigated the association between RII mutations and various clinicopathological variables and genetic alterations in a large series of sporadic adenocarcinomas arising in the proximal colon. RII mutations were found in 17 per cent (36/210) of right-sided tumours and in 86 per cent (32/37) of those displaying RER+. They were associated with the absence of lymph node invasion (P = 0.04), poor histological differentiation (P = 0.006), and with a trend for improved patient survival. Tumours with an RII mutation also showed non-significant trends for a lower incidence of p53 protein overexpression and of p53, K-ras, and APC gene mutation compared with tumours with normal RII. These results indicate that right-sided colorectal tumours containing RII mutations resemble those with the RER+ phenotype in terms of their clinicopathological features and genetic alterations.

Adenocarcinoma↗
PubMed1998

Rapid and nonisotopic SSCP-based analysis of the BAT-26 mononucleotide repeat for identification of the replication error phenotype in human cancers.

Microsatellite instability is an important new form of genetic alteration that characterizes tumors of the replication error phenotype (RER+). The RER status of tumors has been determined until now by analyzing several microsatellite loci for size variations compared with matching normal DNA. This has been done by separating isotopically labeled PCR products on denaturing gels. We recently showed that deletions within BAT-26, a polyadenine tract within the hMSH2 gene, could be used to establish the RER status of tumors without the need for matching normal DNA. We now propose a rapid and nonisotopic method of determining RER status based on PCR-SSCP analysis of the BAT-26 poly(A) tract. Compared with conventional means of examining RER+, this method reduces the PCR and gel manipulations involved by 10-fold. Size variations of as little as 2 bp in the BAT-26 sequence were readily detected using mini-sized, silver-stained SSCP gels run for 2.5 h. The incidence of RER+ detected in 183 right-sided colonic, 121 gastric, and 123 endometrial carcinomas was 20%, 10%, and 5% respectively, using this method. Frameshift mutations in the mononucleotide repeat of the TGF-beta RII gene were found in 86% (42/49) of RER+ but less than 1% (1/237) of RER- colon and gastric tumors identified by BAT-26 analysis. Our results demonstrate that nonisotopic PCR-SSCP of the poly(A) BAT-26 tract is a specific, sensitive and rapid method for determining the RER status of human tumors.

DNA Replication↗
PubMed1998

Determination of the replication error phenotype in human tumors without the requirement for matching normal DNA by analysis of mononucleotide repeat microsatellites.

Microsatellite instability (MI) characterizing tumors with replication errors (RER+ tumors) was first described in colorectal tumors from hereditary non-polyposis colorectal cancer (HNPCC) patients as well as in sporadic cases. It has also been observed in subgroups of extracolonic sporadic tumors, but there is no consensus as to the number of microsatellite loci to examine, and the threshold percentage of unstable loci required to classify a tumor as RER+. We have recently shown that BAT-26, a mononucleotide repeat microsatellite, was quasi-monomorphic in DNA from normal individuals and from colorectal RER- samples, and showed important size variations in RER+ samples. In the present work, we analyzed BAT-26 allelic profiles in tumors of the breast (n = 107), brain (n = 78), stomach (n = 59), prostate (n = 49), esophagus (n = 36), thyroid (n = 31), endometrium (n = 12), and cervix (n = 10) whose RER status was already known, thus extending BAT-26 analysis to a total of 542 human solid tumors. BAT-26 alleles were quasi-monomorphic in RER- samples (475/481) and shortened in RER+ tumors (57/61), including four tumors shown to have been misclassified on the basis of dinucleotide repeat microsatellite analysis. In 3/481 RER- and 4/61 RER+ cases, BAT-26 size variation was important enough to attract attention, but not sufficient to establish the RER status of the corresponding tumors. In these cases, the analysis of BAT-25 and BAT-34C4, two other mononucleotide repeat microsatellites, was necessary to resolve the ambiguity. There were only 3 false positive cases. In conclusion, BAT-26 was able to identify the RER status of 539 out of 542 tumors from various origins (99.5% efficiency) in a single-step experiment without the requirement for matching normal DNA.

Breast Neoplasms↗
PubMed1998

Analysis of the p21 gene in gliomas.

The p21 gene encodes a cyclin dependent kinase inhibitor protein (p21) which has a tumor suppressive activity in a variety of tumor cell lines. Since, the p21 gene is up-regulated by the p53 tumor suppressor gene, which is frequently mutated in gliomas, acting therefore in the same control pathway, it constitutes a good candidate gene to be also inactivated in these tumors. To test this hypothesis, DNAs from 81 gliomas (48 glioblastomas, 11 anaplastic astrocytomas, 10 low-grade astrocytomas, 12 oligodendrogliomas and mixed gliomas), were investigated for mutations in the p21 coding sequence by denaturant gradient gel electrophoresis followed by sequencing. All these tumors have been previously screened for p53 mutations. Three different DNA variants were identified on codon 31 (17 cases), 27 (1 case) and 117 (1 case) and shown to be also present in matching constitutional DNA, suggesting they were polymorphisms. None of the tumors demonstrated a somatic mutation. No significant correlation between the presence of a p21 variant and the p53 mutation tumor status was observed. In conclusion, mutation in the p21 gene unlikely contributes to the development of gliomas.

Alleles↗
PubMed1997

Alternative genetic pathways in colorectal carcinogenesis.

The comparative typing of matched tumor and blood DNAs at dinucleotide repeat (microsatellite) loci has revealed in tumor DNA the presence of alleles that are not observed in normal DNA. The occurrence of these additional alleles is possibly due to replication errors (RERs). Although this observation has led to the recognition of a subtype of colorectal cancer with a high incidence of RERs (caused by a deficiency in DNA mismatch repair), a thorough analysis of the RER frequency in a consecutive series of colorectal cancers had not been reported. It is shown here that the extensive typing of 88 colorectal tumors reveals a bimodal distribution for the frequency of RER at microsatellite loci. Within the major mode (75 tumors, RER- subtype), the probability that a locus exhibited instability did not differ significantly among loci and tumors, being 0.02. The subsequent development of a statistical test for an operational discrimination between the RER- and RER+ subtypes indicated that the probability of misclassification did not exceed 0.001 in this series. The frequency of K-ras mutation was found to be equivalent in the two subtypes. However, in the RER+ tumors, the p53 gene mutation was less frequently detected, the adenomatous polyposis coli (APC) mutation was rare, and the biallelic inactivation of either of these genes was not observed. Furthermore, the concomitant occurrence of APC and tumor growth factor beta receptor type II gene alterations was found only once. These data suggest that the repertoires of genes that are frequently altered in RER+ and RER- tumors may be more different than previously thought.

Aged↗
PubMed1997

Allelic profiles of mononucleotide repeat microsatellites in control individuals and in colorectal tumors with and without replication errors.

We have recently shown that analysis of BAT-26, was sufficient to establish the Replication Error status of colorectal tumors and cell lines without the need for matching normal DNA. BAT-26, a poly(A) tract in the 5th intron of the hMSH2 gene, does not present significant size variation either between the alleles of one individual or between alleles of different individuals. In colorectal tumors without defects in the replication error system (RER- phenotype), BAT-26 is also quasi-monomorphic. On the contrary, in RER+ colorectal tumors, BAT-26 shows unstable shortened alleles. In order to see whether this behaviour was specific for BAT-26, or was a more general phenomenon for mononucleotide repeat microsatellites, we analysed eight other mononucleotide repeats. In control individuals (72 samples) and in RER- colorectal tumors and cell lines (55 samples), these microsatellites were polymorphic, dimorphic, quasi-monomorphic or monomorphic, indicating that the quasi-monomorphic nature of BAT-26 was not a general rule. All of them showed a tendency to be shorter in RER+ colorectal tumors and cell lines (19 samples), but only quasi-monomorphic and monomorphic mononucleotide repeats could be used to determine the RER status of tumors without matching normal DNA, although with a lower efficiency than BAT-26 due to either a smaller range of shortening in RER+ tumors or to a larger number of false negative cases.

Alleles↗
PubMed1997

Loss of heterozygosity on chromosome 9 and p16 (MTS1, CDKN2) gene mutations in esophageal cancers.

Loss of heterozygosity on chromosome 9 has been reported in a variety of human cancers. The cyclin-dependent kinase inhibitor p16 gene, mapped on chromosome 9p21, is presumed to be the tumor-suppressor gene localized in this chromosome. The aim of our study was to determine, in 26 Barrett's adenocarcinomas and 20 squamous-cell carcinomas of the esophagus, the prevalence of loss of heterozygosity on chromosome 9 by typing of microsatellite loci and mutation of p16 by direct sequencing of exons 1 and 2. Allelic losses were found in 69% of adenocarcinomas, but only a microdeletion in exon 1 of p16 occurred in 1 tumor. Among squamous-cell carcinomas, 65% had allelic losses and 5 tumors were mutated on the p16 gene (1 deletion, 3 nucleotide substitutions and 1 insertion). The relatively low rate of p16 mutation observed here coupled with the high frequency of loss of heterozygosity on chromosome 9 suggests that one or several tumor-suppressor gene(s) distinct from p16 may be the target(s) of allelic deletion in most esophageal cancers or that p16 is inactivated in another way.

Adenocarcinoma↗
PubMed1997

BAT-26, an indicator of the replication error phenotype in colorectal cancers and cell lines.

Instability of microsatellites is a hallmark of the DNA replication error phenotype (RER+) due to the inactivation of mismatch repair genes. In humans, microsatellite instability has first been described in colorectal tumors developing in either hereditary nonpolyposis colorectal cancer or sporadic patients. Colorectal tumorigenesis in RER+ and RER- tumors is probably due to distinct mechanisms, and RER+ tumors have a better prognosis than RER- tumors. The study of the RER status of a tumor may thus be important in the future to determine biological prognosis factors and investigate therapeutical strategies. The RER status of 134 primary tumors and 26 cell lines derived from colorectal cancers was established by PCR amplification and analysis of a minimum of 32 microsatellite loci. This characterization allowed us to unambiguously classify 35 primary tumors and 7 cell lines as RER+. Typing of a single poly(A) tract, BAT-26, was sufficient to confirm the RER status of 159 of these 160 tumors and cell lines. Moreover, in DNA from unaffected individuals, normal tissues of a subset of the RER+ patients, and all RER- tumors or cell lines, BAT-26 was quasi-monomorphic, showing only minor size variations. BAT-26 alleles showing shortening from 4 to 15 bp were observed in all but 1 of the RER+ tumors and cell lines. The size difference between the range of normal large alleles and unstable small alleles was sufficient to be detected by electrophoresis on conventional polyacrylamide gels stained with ethidium bromide. We thus propose a simple, low-cost, and rapid method to screen for the RER status of colorectal cancer primary tumors and cell lines, even in the absence of matching normal DNA and, in most cases, without the need for radioactivity. This method could easily be set up in routine laboratories.

Alleles↗
PubMed1997

Infrequent microsatellite instability in oesophageal cancers.

Alterations of microsatellites have been found at relatively high frequency in hereditary and sporadic colorectal cancer and gastric and pancreatic cancers and at lower frequency in some other cancers. We determined the frequency of instability at 39 poly-CA microsatellite loci in 20 squamous cell carcinomas and 26 Barrett's adenocarcinomas of the oesophagus. None of the tumours presented instability for a high percentage of the tested loci. Four squamous cell carcinomas and six Barrett's adenocarcinomas showed microsatellite instability at one locus, and three Barrett's adenocarcinomas showed microsatellite instability at two loci. The presence of few loci showing microsatellite instability could be due to an instability background. We conclude that genetic defects in the DNA mismatch repair system do not play an important role in oesophageal cancers.

Adenocarcinoma↗
PubMed1997

[p53 and colorectal cancer].

Mutations of the p53 gene have been found in 380 of the 768 tumors (49.5%) included in the eight largest published series of colorectal cancer. Most point mutations of p53 change the conformation of the gene, and by stabilizing it make it detectable by immunohistochemistry. However, studies using both tests for p53 mutations and immunohistochemical methods found that the results of these two approaches were concordant in only 68% of cases. Conflicting data have been reported regarding the prognostic significance of positive p53 staining. Presence of a mutation is generally believed to indicate a poor prognosis.

Colorectal Neoplasms↗