hMSH2 Protein Expression Research Articles

DNA Mismatch Repair Enzyme Activity and Gene Expression in Prostate Cancer

Microsatellite instability (MSI) of short repetitive sequences in human chromosomal DNA can result from defective DNA mismatch repair function in tumor cells. We hypothesize that DNA mismatch repair (MMR) activity is down-regulated during prostatic carcinogenesis. To test this hypothesis, MMR activities and mismatch repair-related genes were analyzed in five different prostate cancer cell lines. Our results demonstrate that MMR activities were decreased as compared to MMR proficient HeLa cells. Interestingly, LNCaP, PC-3 and DU145 had much lower MMR activities as compared to DUPro and TSUPr1. The MMR-related genes (hMLH1, hPMS1, hPMS2, hMSH2, hMSH3, hMSH6) showed mRNA transcripts in all prostate cancer cell lines. However, Western blotting showed decreased or absent hMLH1 protein expression in PC-3, DU145, DUPro and TSUPr1 cells. Similarly, the hMSH2 protein expression was low or absent in DU145 and LNCaP cells. This is the first report that demonstrates decreased MMR activities is associated with low expression of hMLH1, hMSH2 and other MMR-related proteins in prostate cancer.

Assessment of markers for the identification of microsatellite instability phenotype in gastric neoplasms

We tested three mononucleotide, 45 dinucleotide, and five tetranucleotide repeats in 30 gastric adenomas and 30 gastric carcinomas for microsatellite instability (MSI) in order to evaluate which microsatellites might indicate the MSI status in gastric neoplasms. Along with the increase in tested markers, the proportion of low-frequency MSI (MSI-L) tumors increased. On immunohistochemistry, MSI-L gastric neoplasms did not show any alteration in hMLH1 or hMSH2 protein expression, while most of the high-frequency MSI (MSI-H) tumors did show alterations in the above mismatch repair proteins. The above findings suggested that MSI-L tumors cannot be distinguished from microsatellite stable tumors. Two mononucleotides, BAT25 and BAT26, were sufficient for the screening of MSI. An additional three dinucleotides, D17S786, D6S105 and D19S188, were also highly sensitive and specific in identifying MSI phenotype tumors.

Prise en charge multi-disciplinaire du cancer colorectal héréditaire

The aim of this study was to assess the feasibility and success of multidisciplinary approach for the management of hereditary colorectal cancer. From November 1998 to November 2000, 32 individuals with putative familial/hereditary predisposition to colorectal cancer were investigated for adenomatous polyposis (attenuated or classical familial adenomatous polyposis coli, FAP) or for hereditary nonpolyposis colorectal cancer (HNPCC). Amsterdam criteria (I and II) and Bethesda guidelines were used to select putative HNPCC kindreds. Clinical data including endoscopy, pathological and operative reports as well as family history were collected. Pre- and post-test genetic counseling was offered to at-risk individuals. Genetic testing included microsatellite instability (MSI) and search for germline mutations in the APC, hMSH2 and hMLH1 genes. Immunohistochemistry (IHC) of hMSH2 and hMLH1 protein expression in tumour samples was also performed. 11 APC mutations were characterized, whereas four mutations in HNPCC genes were found in hMSH2 (2) and in hMLH1 (2). MSI and IHC correlated completely for cases with identified pathogenic mutation (100%). A thorough evaluation and management of hereditary colorectal requires a multidisciplinary approach. Thus, more mutation carriers can be identified and benefit from appropriate genetic counselling, while non-carrier individuals are relieved from unnecessary surveillance.

Clinicopathological and molecular genetic analysis of 4 typical Chinese HNPCC families.

To study the clinicopathological and molecular genetic characteristics of typical Chinese hereditary nonpolyposis cotorectal cancer (HNPCC) families. Four typical Chinese HNPCC families were analyzed using microdissection, microsatellite instability analysis, immunostaining of hMSH2 and hMLH1 proteins and direct DNA sequencing of hMSH2 and hMLH1 genes. All five tumor tissues of 4 probands from the 4 typical Chinese HNPCC families showed microsatellite instability at more than two loci (MSI-H or RER+ phenotype). Three out of the 4 cases lost hMSH2 protein expression and the other case showed no hMLH1 protein expression. Three pathological germline mutations (2 in hMSH2 and 1 in hMLH1), which had not been reported previously, were identified. The same mutations were also found in other affected members of two HNPCC families,respectively. Typical Chinese HNPCC families showed relatively frequent germline mutation of mismatch repair genes. High-level microsatellite instability and loss of expression of mismatch repair genes correlated closely with germline mutation of mismatch repair genes. Microsatellite instability analysis and immunostaining of mismatch repair gene might serve as effective screening methods before direct DNA sequencing. It is necessary to establish clinical criteria and molecular diagnostic strategies more suitable for Chinese HNPCC families.

Microsatellite instability and hMLH1/hMSH2 expression in young endometrial carcinoma patients: associations with family history and histopathology.

Endometrial cancer is the second most common malignancy in patients with hereditary nonpolyposis colorectal cancer (HNPCC). The age at diagnosis of HNPCC-associated endometrial cancer is approximately 15 years younger than for sporadic endometrial cancer. Our current study was undertaken to determine the frequency of microsatellite instability (MSI) and absence of hMLH1 or hMSH2 protein expression in young patients with endometrial carcinoma and to correlate these findings with histopathologic and clinical features. Endometrial carcinoma from 62 women (23-52 years, median age 46) were assessed for MSI. Twenty-one of the 62 (34%) tumors demonstrated MSI. Of the 21 tumors demonstrating MSI, 12 showed an absence of hMLH1 expression, 4 showed an absence of hMSH2 expression, and 5 demonstrated normal expression of both proteins. All 41 tumors without MSI demonstrated normal hMLH1 and hMSH2 expression. Two patients with MSI tumors fulfilled the Amsterdam criteria for HNPCC, while 2 had histories suggestive of HNPCC. None of the patients with tumors without MSI had a personal or family cancer history suggestive of HNPCC. The MSI phenotype was associated (p < 0.05) with high FIGO stage and grade, cribriform growth pattern, mucinous differentiation and necrosis. Our findings suggest that the frequency of HNPCC in young endometrial cancer patients is relatively low when compared with the frequency of HNPCC in young colorectal cancer patients. Defects of the MMR proteins hMSH2 or hMLH1 account for MSI in most but not all endometrial cancers from young patients.

Altered expression of hMLH1 and hMSH2 protein in endometrial carcinomas with microsatellite instability

Microsatellite instability (MI) has been observed in approximately 20% of presumably sporadic cases of uterine endometrioid carcinoma (UEC). A previous mutational analysis of the 4 known DNA mismatch repair genes (hMSH2, hMLHI, hPMS1, and hPMS2) on a small number of Ml-positive tumors detected mutations in only 2 of 8 cases, both in hMSH2. To further explore the underlying cause of MI in UEC, we analyzed the protein expression of hMSH2 and hMLHI in UEC of known MI status. Formalin-fixed, paraffin-embedded archival tissue from 21 UECs was analyzed by immunoperoxidase staining with monodonal antibodies against hMLH1 and hMSH2 protein. Tumors were evaluated for presence of nuclear staining by 3 investigators. Lack of nuclear hMLHI staining was found in 7 of 13 carcinomas with MI, but in none of 8 carcinomas without MI (Fischer's exact, 0.018). Lack of nuclear hMSH2 staining was found in 3 of the MI-positive cases, but none of the MI-negative cases (not statistically significant). Taken together, lack of nuclear staining of either hMLH1 or hMSH2 was found in 9 of 13 cases with MI and in none of 8 cases without MI (Fischer's exact, 0.005). We conclude that MI in sporadic UEC appears to be associated with lack of expression of either hMLH1 or hMSH2, suggesting that inactivation of these genes may be responsible for MI in most MI-positive sporadic UECs.

Immunohistochemical analysis of DNA mismatch repair enzyme hMSH-2 in normal human skin and basal cell carcinomas.

We have analysed the expression and distribution of the DNA mismatch repair enzyme hMSH-2 in normal skin and basal cell carcinomas. hMSH-2 protein was investigated immunohistochemically (normal human skin: n = 10; basal cell carcinomas: n = 16) on frozen sections using a highly sensitive streptavidin-peroxidase technique and a specific mouse monoclonal antibody (clone FE11). In normal human skin, we found nuclear immunoreactivity for hMSH-2 in epidermal keratinocytes of the basal and first 1-3 suprabasal cell layers. All basal cell carcinomas analysed revealed strong nuclear imunoreactivity that was pronounced in peripheral tumour cells and cells of the palisade. Expression of hMSH-2 protein was consistently and strongly upregulated in tumour cells of the carcinomas as compared to adjacent unaffected epidermis or epidermis of normal human skin. Twelve of the sixteen carcinomas analysed revealed no visual correlation in comparing the labelling patterns for hMSH-2 with the labelling pattern for the proliferation marker Ki-67. Our findings indicate that (a) hMSH-2 is expressed in human epidermal keratinocytes, predominantly in lower cell layers of the viable epidermis; (b) expression of hMSH-2 protein is strongly upregulated in basal cell carcinomas as compared to unaffected epidermis; (c) the level of hMSH-2 proteins in the carcinomas is not exclusively regulated by the proliferative activity of these tumour cells; (d) inactivating mutations of the hMSH-2 gene may in the carcinomas not be involved in the carcinogenesis or microsatellite instability secondary to replication errors; (e) expression of hMSH-2 may be of importance for the genetic stability of basal cell carcinomas in vivo.

Origin of Microsatellite Instability in Gastric Cancer

Microsatellite instability (MSI) is observed in 13-44% of gastric carcinoma. The etiology of MSI in gastric carcinoma has not been clearly defined. To assess the role of mismatch repair in the development of MSI in gastric cancer, expression of hMSH2 and hMLH1 was explored. We examined 117 gastric carcinomas for MSI and observed instability at one or more loci in 19 (16%) of these tumors. Of the 19 tumors with MSI, nine exhibited low-rate MSI (MSI-L) with instability at <17% of loci, whereas the remaining 10 exhibited high-rate MSI (MSI-H) with instability at >33% of loci examined. Immunohistochemical staining for hMLH1 and hMSH2 was performed on eight of the tumors with MSI-H, five with MSI-L, and 15 tumors without MSI. All eight tumors with MSI-H showed loss of staining for either hMLH1 (n = 5) or hMSH2 (n = 3). In contrast, tumors with MSI-L or without MSI all showed normal hMSH2 and hMLH1 protein expression patterns. Moreover, all eight of the tumors with MSI-H also showed instability at BAT-26, whereas none of the MSI-L tumors or tumors without instability showed instability at BAT-26. These findings suggest that the majority of high-level MSI in gastric cancer is associated with defects of the mismatch repair pathway. Although larger studies are needed, BAT-26 appears to be a sensitive and specific marker for the MSI-H phenotype in gastric carcinoma.

Human mismatch repair gene (hMSH2) product expression in relation to recurrence of transitional cell carcinoma of the urinary bladder

Several convincing studies have shown that the hMSH2 gene plays major roles in mismatch repair by recognizing mismatched bases and preventing mutations during DNA replication. Loss of this function may result in the accumulation of DNA replication errors or even the mutator phenotype (which may be responsible for the multiple mutations required for multistep carcinogenesis), and it has been found to affect the prognosis of patients. Thus, the authors felt that it would be of interest to study the expression patterns of hMSH2 protein in malignant tumors and to assess the correlation of hMSH2 protein to various clinical and pathologic features in these patients. The authors examined the expression patterns of hMSH2 protein in 115 patients with transitional cell carcinoma (TCC) of the urinary bladder by immunohistochemical technique using monoclonal antibody to hMSH2 protein. The group in which hMSH2 was preserved (>20% of nuclei were positive for staining by anti-hMSH2 antibody) included 86 of 115 (75%) of the TCCs, whereas the remaining 29 cases (25%) belonged to the group in which hMSH2 was preserved (< or = 20% of tumor nuclei were positive), including 2 negative cases. Univariate analysis indicated that reduced expression of hMSH2 protein was significantly more frequent in high grade tumors than in low grade tumors (P = 0.04). Furthermore, statistical multivariate analysis revealed that hMSH2 reduction was significantly related to the recurrence (P = 0.02). These results suggest that abnormal expression of hMSH2 protein might be involved in tumorigenic processes and in the progression of some TCCs, and that the loss of hMSH2 protein expression might be a useful predictor of recurrence of TCC.

Immunohistochemical analysis of DNA 'mismatch-repair' enzyme human Mut-S-Homologon-2 in ovarian carcinomas.

The human Mut-S-Homologon-2 (hMSH-2) gene product is a member of a highly conserved family of proteins involved in postreplication mismatch repair. We have analysed hMSH-2 expression in normal ovarian tissue (n = 15) and ovarian carcinomas (n = 40). hMSH-2 protein was investigated immunohistochemically on frozen sections using a highly sensitive streptavidin-peroxidase technique and a specific mouse monoclonal antibody (clone FE11). A hMSH-2-immunoreactivity score (hMSH-2-IRS) for semiquantitative analysis of hMSH-2 expression is presented. In normal ovarian tissue, we only found weak nuclear immunoreactivity for hMSH-2 in 60%, while the remaining 40% were hMSH-2 negative (mean hMSH-2-IRS: 0.73; SD: +/-0.70). All ovarian carcinomas analysed revealed moderate to strong nuclear immunoreactivity (mean hMSH-2-IRS: 8.05; SD: +/-3.65). hMSH-2 staining was heterogeneous, with visual differences between individual tumour cells. Expression of hMSH-2 protein was consistently and strongly upregulated in tumour cells of ovarian carcinomas as compared to normal ovarian tissue. No statistically significant correlation in comparing the labelling patterns for hMSH-2 with the labelling patterns for Ki-67 (mean percentage of Ki-67 positive tumour cells: 25.88%; SD: +/-18.43) was observed in ovarian carcinomas. Furthermore, no statistical significant correlations between hMSH-2-IRS and histological grading (p = 0.47), histological type of carcinoma (p = 0.706) or FIGO-classification (p = 0.054) were found. Our findings indicate that (a) hMSH-2 is expressed in normal human ovarian tissue, (b) expression of hMSH-2 is increased in ovarian carcinomas, (c) expression of hMSH-2 may be of importance for the genetic stability of ovarian carcinomas in vivo, (d) hMSH-2 mutations may not cause microsatellite instability in ovarian carcinomas, (e) hMSH-2 may contribute to mechanisms responsible for resistance to anticancer drugs.

Expression of the mismatch repair gene hMSH2 in sporadic colorectal cancer.

At least four genes involved in DNA mismatch repair (MMR), hMSH2, hMLH1, hPMS1 and hPMS2, have been cloned and characterized. These genes have been demonstrated to be altered in the germline of patients with hereditary non-polyposis colorectal cancer (HNPCC). HNPCC is an autosomal dominant disease characterized by a preponderance of proximal colon, young age of onset, increased multiplicity, and improved stage-specific survival. In this study, we examined the expression of hMSH2 protein in sporadic colorectal cancer (CRC). As a result, the frequency of right-sided CRC and multiple CRCs were significantly higher in the patients with hMSH2-negative CRC than in those with hMSH2-positive CRC. The rate of p53 positivity was significantly lower in the hMSH2-negative tumours than that in the hMSH2-positive tumours. The disease-free survival rate tended to be higher in the patients with hMSH2-negative CRC than in the patients with hMSH2-positive CRC. Our findings suggest that both the clinicopathological and biological features of hMSH2-negative sporadic CRC seemed to be similar to those of HNPCC. To clarify the mechanism of carcinogenesis in HNPCC and sporadic CRC, further investigations of genetic alterations caused by MMR genes will be needed.

Expression of hMSH2 and hMLH1 in Colorectal Carcinomas with Microsatellite Instability

Microsatellite instability (MIN) due to defective mismatch repair (MMR) genes has been reported in a subset of sporadic colorectal carcinomas and in the majority of tumors from patients with hereditary nonpolyposis colorectal cancer (HNPCC) syndrome. Among the known MMR genes, hMSH2 and hMLH1 genes are known to be predominantly altered in HNPCC patients and MIN-positive tumors. In this study, we examined MIN and the protein expression pattern of the hMSH2 and hMLH1 by Western blot and immunohistochemistry from 32 sporadic colorectal carcinomas. MIN was observed in 6 (18%) colorectal carcinomas. Of the 6 MIN-positive tumors, one case showed no expression of either protein, 3 cases showed an absence of hMSH2 protein expression, one case showed an absence of hMLH1 protein expression and one case showed no altered expression of either protein by immunohistochemistry. The decreased expression of the hMSH2 protein in a tumor compared to the normal mucosa was also observed in 5 of the 6 MIN-positive cases by Western blot analysis. All of the MIN-negative tumors showed expression of both proteins by immunohistochemistry. Thus most of the MIN-positive tumors appear to be directly related to the altered expression of these two genes and can be diagnosed by the examination of protein expression.