胃癌微卫星不稳定性和抑癌基因杂合缺失

目的研究微卫星不稳和抑癌基因缺失在胃癌发生中的作用．方法采用ＰＣＲ为基础的方法,检测了５３例胃癌中６个微卫星标记突变及ＡＰＣ／ＭＣＣ和ＤＣＣ基因杂合缺失（ＬＯＨ）．结果胃癌微卫星不稳的检出率为３２１％（１７／５３）．７例（１３２％）为微卫星高频率不稳（３个以上微卫星标志）,１０例（１８９％）为微卫星低频率不稳（１或２个微卫星标记）．肠型胃癌微卫星高频率不稳的发生率（２５０％）显著高于弥漫型胃癌（３４％）（Ｐ＜００５）．高频率不稳组未发现有ＡＰＣ,ＭＣＣ和ＤＣＣ基因ＬＯＨ,微卫星高频率不稳与ＡＰＣ／ＭＣＣ和ＤＣＣ基因ＬＯＨ呈负相关．结论微卫星不稳在部分胃癌,特别是肠型胃癌早期发生中起重要作用,高频率不稳胃癌与遗传性非息肉大肠癌有共同的特点．与此相反,低频率不稳和无不稳胃癌可能通过ＬＯＨ病理途径发生     

Mutation analysis of APC gene in gastric cancer with microsatellite instability

AIM：To evaluate the role of APCmutation in gastric carcinogenesis and to correlate APC mutation with microsatellite instability(MSI)in gastiric carcinomas.METHODS:APC mutation was measured with multiplexPCR,denaturing gradient gel electrophoresis and DNAsequencing;and MSIwas analyzed by PCR-based methods.RESULTS:Sixty-eight cases of sporadis gastric carcinoma were studied for APCmutation at exon15and MSI,APC mutaions were detected in15(22.1%)gastric cancers,Frequence of APCmutation(33.3%)in in testinal type of gastric ancer was significantly higher than that in diffuse type(13.1%,P<0.05).On the contrary.on association was observed dbtween APC mutation and tumor size,differentiation,depth of invasion,metastasis or clinical stages.Using five microsatellite markers.MSIin at least one locus was detected in 17of68(25%)of the tumors analyzed,APC mutations were all detected in MSI-L(only one locus,n=9)orMSS(tumor lacking MSI or stable,n=51),but no mutation was found in MSI-H(≥2loci,n=8).CONCLUSION:APC mutation is involved in carcinogenesis of intestial type of gastric cancer and is independent of MSI phenotype but related to the LOH pathway in gastri cancer.     

Loss of heterozygosity at adenomatous polyposis coli,mutation in colorectal cancer and deleted in colorectal cancer genetic loci in colorectal cancers

AIM: To evaluate the role and analyze the loss of heterozygosity (LOH) of adenomatous polyposis coli (APC), mutation in colorectal cancer (MCC) and deleted in colorectal cancer (DCC) genes in the development and progression of colorectal cancers. METHODS: LOH at APC, MCC and DCC genes was examined in 41 surgically resected specimens of colorectal carcinomas by polymerase chain reaction and restriction fragment length polymorphism analysis technique. RESULTS: LOH of APC and MCC were observed in 7 of 25 (28.0%) and 8 of 22 (36.4%) of informative cases, respectively. When considered as one locus, the LOH frequency for APC/MCC was 14 of 36 (38.9%). LOH at DCC gene locus was detected in 21 of 38 (55.3%) of informative cases. No correlation was found between the LOH at APC or MCC gene and tumor histological types, size, invasion, lymph node metastasis and Dukes’ stages (P > 0.05). However, LOH rates at DCC locus in the group with lymph-node metastasis (80.0%) and in Dukes’ stages III and IV (71.4%) were significantly higher than those without lymph node metastasis (39.1%) and in Dukes’ stages I and II (35.3%) (P < 0.05). CONCLUSION: LOH at APC and/or MCC may occur more frequently in the early stages and plays a role in the initiation of colorectal cancer while LOH at DCC is frequent at late event and associated with the progression and metastasis of colorectal cancer.     

Mutation and methylation of hMLH1 in gastric carcinomas with microsatellite instability

AIM: To appraise the correlation of mutation and methylation of hMSH1 with microsatellite instability (MSI) in gastric cancers.METHODS: Mutation of hMLH1 was detected by Twodimensional electrophoresis (Two-D) and DNA sequencing;Methylation of hMLH1 promoter was measured with methylation-specific PCR; MSI was analyzed by PCR-based methods.RESULTS: Sixty-eight cases of sporadic gastric carcinoma were studied for mutation and methylation of hMLH1 promoter and MSI. Three mutations were found, two of them were caused by a single bp substitution and one was caused by a 2 bp substitution, which displayed similar Two-D band pattern.Methylation of hMLH1 promoter was detected in 11(16.2%)gastric cancer. By using five MSI markers, MSI in at least one locus was detected in 17/68(25%) of the tumors analyzed.Three hMLH1 mutations were all detected in MSI-H (≥2 loci,n=8), but no mutation was found in MSI-L (only one locus,n=9) or MSS (tumor lacking MSI or stable, n=51). Methylation frequency of hMLH1 in MSI-H (87.5%, 7/8) was significantly higher than that in MSI-L (11.1%, 1/9) or MSS (5.9%, 3/51)(P&lt;0.01-0.001), but no difference was found between MSI-L and MSS (P&gt;0.05).CONCLUSION: Both mutation and methylation of hMLH1 are involved in the MSI pathway but not related to the LOH pathway in gastric carcinogenesis.     

Is gastric cancer part of the tumour spectrum of hereditary non-polyposis colorectal cancer? A molecular genetic study

BACKGROUND: Gastric cancer is the second most common extracolonic malignancy in individuals with hereditary non-polyposis colorectal cancer (HNPCC)/Lynch syndrome. As gastric cancer is relatively common in the general population as well, it is not clear whether or not gastric cancer is a true HNPCC spectrum malignancy. AIM: To determine whether or not gastric cancer is a true HNPCC spectrum malignancy. Subjects and METHODS: The molecular and clinicopathological profiles of gastric cancers (n = 13) from HNPCC mutation carriers were evaluated and compared with the profiles of sporadic gastric cancers (n = 46) stratified by histology and microsatellite instability (MSI) status. RESULTS: This study on sporadic and HNPCC gastric cancers revealed several important universal associations. Loss of heterozygosity in the adenomatous polyposis coli (APC) region was associated with intestinal histology regardless of the MSI (p = 0.007). KRAS-mutations (p = 0.019) and frameshift mutations in repeat tracts of growth-regulatory genes (p<0.001) were associated with MSI tumours being absent in microsatellite stable (MSS) tumours. The average number of methylated tumour suppressor gene loci among the 24 genes studied (methylation index) was higher in MSI than in MSS tumours regardless of histology (p<0.001). Gastric cancers from HNPCC mutation carriers resembled sporadic intestinal MSI gastric cancers, except that MLH1 promoter methylation was absent (p<0.001) and the general methylation index was lower (p = 0.038), suggesting similar, but not identical, developmental pathways. All these lacked the mismatch repair protein corresponding to the germline mutation and displayed high MSI. CONCLUSION: The present molecular evidence, combined with the previous demonstration of an increased incidence relative to the general population, justify considering gastric cancers as true HNPCC spectrum malignancies.     

Loss of heterozygosity： An independent prognostic factor of colorectal cancer

AIM: Colorectal cancers result from the accumulation of several distinct genetic alterations. This study was to investigate the frequency and prognostic value of loss of heterozygosity (LOH) and microsatellite instability (MSI) at 14 genetic loci located near or within regions containing important genes implicated in colorectal tumorigenesis. METHODS: We studied colorectal cancers with corresponding normal mucosae in 207 patients (139 males and 68 females, mean age at the time of tumor resection 66.2±12.4 years, range 22-88 years). There were 37 right-sided colonic tumors, 85 left-sided colonic tumors and 85 rectal tumors. The distribution of tumor staging was stage Ⅰ in 25, stage Ⅱ in 73, stage Ⅲ in 68, and stage Ⅳ in 41. We analyzed the LOH and MSI of HPC1, hMSH2, hMLH1, APC, MET, P53, NH23-H1, DCC, BAT25, BAT26, D17S250, MYCL1 and D8S254 with fluorescent polymerase chain reaction and denatured gel electrophoresis. High-frequency LOH was determined to be greater than three, or more than 50% of the informative marker with LOH. High-frequency MSI (MSI-H) was determined as more than four markers with instability (>30%). Correlations of LOH and MSI with clinical outcomes and pathological features were analyzed and compared. RESULTS: The occurrence of MSI-H was 7.25%, located predominantly in the right colons (7/15) and had a higher frequency of poor differentiation (6/15) and mucin production (7/15). LOH in at least one genetic locus occurred in 78.7% of the tumors and was significantly associated with disease progression. Of the 166 potentially cured patients, 45 developed tumor recurrence within 36 mo of follow-up. Clinicopathological factors affecting 3-year disease-free survival (DFS) were TNM staging, grade of differentiation, preoperative CEA level, and high LOH status. Patients with high LOH tumors had a significantly lower DFS (50%) compared with patients with low LOH tumors (84%). Of the patients developing subsequent tumor recurrence, the number and percentage of LOH were 2.97 and 46.8% respectively, similar to the stage IV disease patients. TNM staging had the most significant impact on DFS, followed by high LOH status. CONCLUSION: Clinical manifestations of LOH and MSI are different in colorectal cancer patients. High-frequency LOH is associated with high metastatic potential of colorectal cancers.     

Genetic aberrations common in gastric high-grade large B-cell lymphoma

Genetic aberrations associated with the development of extranodal high-grade large B-cell lymphoma originating in the stomach have not been fully identified yet. We analyzed 31 such lymphomas using 73 microsatellite markers for allelic imbalance and microsatellite instability. The highest frequency (42%) of loss of heterozygosity (LOH) was found on the long arm of chromosome 6. We identified 2 LOH hot spots on 6q21-22.1 and 6q23.3-25, flanked by markers D6S246-D6S261 and D6S310-D6S441, respectively, containing putative tumor suppressor genes (TSGs). These 6q aberrations were found to be the sole allelic imbalance in 1 patient only; they were mostly accompanied by additional abnormalities. Several known TSGs, namely, the APC, p15/p16, p53, and DCC genes, were found to suffer frequent LOH during lymphomagenesis. LOH was also detected in regions containing putative TSGs on 7q and 13q14. Frequent amplification of genomic material was found in the 2p, 3q27 at the BCL-6 gene locus, 6p, 7q, 11q23-24 at the MLL gene locus, and 18q regions. Analysis of the pattern of occurrence of these aberrations revealed an association of the amplification of the MLL gene region with LOH at the p53 locus (P =.02). Only low frequency of microsatellite instability (MSI) was detected in these lymphomas and MSI incidence increased with age (P =.01). Karyotypic instability thus plays the main role in the development of gastric high-grade large B-cell lymphoma. Common genetic aberrations responsible for lymphomagenesis are deletions of 6q, loss of p53, and amplification of the 3q27 and the MLL gene regions. (Blood. 2000;95:1180-1187)     

Relationship between grade of microsatellite instability and target genes of mismatch repair pathways in sporadic colorectal carcinoma

Microsatellite instability (MSI) induces carcinoma through the alteration of target genes; TGF- RII, BAX, IGFIIR, hMSH3, and hMSH6. The grade of MSI is classified into two categories according to the number of positive microsatellite markers; MSI-H and MSI-L. To elucidate the relationship between the propriety of grading MSI, target genes and clinical features of sporadic colorectal cancers, MSI and frameshift mutation of target genes were examined in 132 colorectal cancer patients. Fourteen and 12 cases showed MSI-H and MSI-L, respectively. Of the 14 MSI-H cases, 13 cases showed alteration of target genes. However, of the 12 MSI-L cases, only one case showed alteration of one target gene. Furthermore, all 14 MSI-H patients had cancers of the right colon and were typically older in age than the MSI-negative patients. These results imply a strong relationship between MSI-H and carcinogenesis by the frameshift mutation of target genes.     

大肠癌组织APC/MCC基因杂合的缺失

目的研究大肠癌ＡＰＣ／ＭＣＣ基因杂合缺失的作用。方法采用ＰＣＲ技术，并配合限制性片段长度多态现象（ＲＦＬＰ）分析，对４１例外科手术切除大肠癌组织ＡＰＣ／ＭＣＣ基因杂合缺失（ＬＯＨ）进行检测。结果在大肠癌４１例中ＡＰＣ基因属信息个体者２５例，检出ＬＯＨ７例，占２８．０％；ＭＣＣ基因属信息个体者２２例，检出ＬＯＨ８例，占３６．４％。若将ＡＰＣ和ＭＣＣ基因进行综合分析，则信息个体者３６例，检出ＬＯＨ１４例，占３８．９％。ＡＰＣ和ＭＣＣ基因的ＬＯＨ与肿瘤大小、组织学类型、浆膜浸润、淋巴结转移及Ｄｕｋｅｓ分期无关（Ｐ＞０．０５）。结论ＡＰＣ和ＭＣＣ基因ＬＯＨ是大肠癌的常见改变     

Two types of sporadic multiple colorectal cancers with and without HNPCC-like genetic instability

BACKGROUND/AIMS: Sporadic multiple colorectal cancers (MCCs) potentially have similar genetic alterations to hereditary nonpolyposis colorectal cancers (HNPCCs), but genetically unstable MCCs other than HNPCCs are not well characterized. We report the frequency of HNPCC-like sporadic multiple colon cancers and their characterization as for HNPCC-related gene mutations. METHODOLOGY: Microsatellite instability (MSI) at 12 microsatellite loci was examined by a polymerase chain reaction in 19 cases each of MCC and single colorectal cancer (SCC). The target sequences of MSI, transforming growth factor beta type II receptor (TGF beta RII) gene, and the HNPCC genes, hMSH2 and hMLH1, were amplified and analyzed for mutations by sequencing. RESULTS: In 5 of 19 cases with MCC, MSI was observed at more than 4 microsatellite loci, and the other cases including all SCCs showed no MSI or MSI(+) at a single locus. In 4 of the 5 severe MSI(+) cases, a 10-bp adenine tract at codons 125-128 of the TGF beta RII gene was mutated. In terms of the hMSH2 and hMLH1 genes, only silent mutations and non-critical amino acid substitutions were found. CONCLUSIONS: We found severe MSI in 26% of sporadic multiple colorectal cancers. Mutations of the TGF beta RII gene are closely associated with severe MSI(+) MCCs as observed in HNPCC, suggesting that these MCCs develop by the similar carcinogenic process to HNPCC.     

The CpG island methylator phenotype and chromosomal instability are inversely correlated in sporadic colorectal cancer

BACKGROUND & AIMS: The CpG island methylator phenotype (CIMP) is one of the mechanisms involved in colorectal carcinogenesis (CRC). Although CIMP is probably the cause of high-frequency microsatellite instability (MSI-H) sporadic CRCs, its role in microsatellite stable (MSS) tumors is debated. The majority of MSS CRCs demonstrate chromosomal instability (CIN) with frequent loss of heterozygosity (LOH) at key tumor suppressor genes. We hypothesized that the majority of sporadic CRCs without CIN would be associated with CIMP. METHODS: We tested 126 sporadic CRCs for MSI and LOH and categorized tumors into MSI, LOH, or MSI-/LOH- subgroups. Methylation status was evaluated using 6 CIMP-related markers (MINT1, MINT2, MINT31, p16(INK4alpha), p14(ARF), and hMLH1) and 6 tumor suppressor genes (PTEN, TIMP3, RUNX3, HIC1, APC, and RARbeta2). BRAF V600E mutation analysis was performed using allele-specific polymerase chain reaction and DNA sequencing. RESULTS: We observed frequent methylation at all 12 loci in all CRCs. BRAF V600E mutations correlated with the MSI (P < .0001) and MSI-/LOH- (P = .03) subgroups. MSI and MSI-/LOH- tumors exhibited more promoter methylation than CRCs with LOH (P < .0001). We also found an inverse correlation between the frequencies of methylation and LOH (rho = -0.36; P < .0001). CONCLUSIONS: The associations between methylation frequencies at CIMP-related markers and MSI or MSI-/LOH- sporadic CRCs suggest that the majority of these tumors evolve through CIMP. These findings suggest that CIN and CIMP represent 2 independent and inversely related mechanisms of genetic and epigenetic instability in sporadic CRCs and confirm that MSI cancers arise as a consequence of CIMP.     

Microsatellite instability in patients with multiple primary cancers of the gastrointestinal tract

BACKGROUND: Little is known about the genetic alterations in multiple primary cancers of the gastrointestinal tract. Microsatellite instability (MSI) is frequently observed in hereditary non-polyposis colorectal cancer (HNPCC), and multiple primary cancers is a feature of this syndrome. AIMS: To identify MSI incidence, target gene mutation, and mismatch repair (MMR) protein status in patients with multiple primary cancers of the gastrointestinal tract. SUBJECTS: Fifty seven cancers from 22 Japanese patients with multiple primary cancers of the stomach, duodenum, colon, and rectum. METHODS: MSI was examined at 5-7 microsatellite loci. Mutation analysis for TGFbetaRII, IGFIIR, and BAX was performed in cancers with MSI. MMR protein status was examined by immunohistochemical analysis using a monoclonal antibody against hMSH2 and hMLH1. RESULTS: MSI was observed in 16 of 22 patients (73%) and in 29 of 57 lesions (51%). High frequency MSI (MSI-H) was found often in patients with multiple cancers in the same organ (p = 0.042), especially in multiple gastric cancer patients (p = 0.038). In contrast, patients with multiple cancers in different organs had a tendency to show low frequency MSI (MSI-L) or microsatellite stable (MSS) phenotype. Both target gene mutation and decreased expression of MMR protein were found only in seven lesions of three patients with MSI at more than four microsatellite loci. CONCLUSIONS: These results suggest that genetic instability may play an important role in the development of multiple gastrointestinal cancers but there may be different genetic alterations between multiple gastrointestinal cancers of the same and different organs.     

Genetic characterization of colorectal cancers in young patients based on chromosomal loss and microsatellite instability

BACKGROUND: In recent studies a high frequency of microsatellite instability among colorectal cancers in young patients has been reported, but the frequency of microsatellite instability (MSI) and chromosomal instability among colorectal cancers in young patients has not yet been fully elucidated. Only one report showed an increased loss of heterozygosity (LOH) ratio at 9p locus, which harbors tumor suppressor genes p16. The LOH and MSI status among colorectal cancers in young patients was examined. METHODS: Twenty-five patients under 40 years of age diagnosed with colorectal cancer were examined for MSI and LOH using 17 microsatellite markers, and also p16 expression patterns were evaluated by immunohistochemistry and methylation status of the p16 gene was assessed by methylation-specific PCR. RESULTS: MSI was observed in only one case (4%). LOH at 2p, 5q, 9p, 11q, 17p, and 18q was observed in 41%, 59%, 42%, 35%, 46%, and 56% of cases, respectively. Eighty-three percent of patients showed p16-positive expression patterns. Fifty percent of colorectal cancers in young patients exhibited p16 methylation (3/6). CONCLUSIONS: Our study demonstrated that colorectal cancers in young patients without MSI showed a high frequency of LOH at the 9p locus. However, LOH status at 9p and p16 expression pattern did not show a significant correlation. Other tumor suppressor genes on the 9p, with the exception of p16, may play an important role in the carcinogenesis of colorectal cancers in young patients.     

Evolution of colorectal cancer: change of pace and change of direction

This review compiles evidence for an alternative to the classical adenoma-carcinoma sequence in the evolution of colorectal cancer. It is suggested that between 30 and 50 of colorectal cancers are not initiated by mutation of the tumor suppressor gene APC, but through the epigenetic silencing of genes implicated in the control of differentiation, cell cycle control and DNA repair proficiency. The precursor polyps are often characterized by a serrated architecture, and include hyperplastic polyps, admixed polyps and serrated adenomas. The alternative pathway is heterogeneous and may culminate in cancers showing low or high level DNA microsatellite instability (MSI-L and MSI-H, respectively), and in cancers that are microsatellite stable (MSS). Cancers showing DNA MSI may be characterized by an accelerated evolution. Cancers in hereditary non-polyposis colorectal cancer show features of both classical (adenoma and APC mutation) and alternative pathways (rapid evolution, MSI-H and lack of chromosomal instability).     

Loss of heterozygosity and HIV infection in patients with anal squamous-cell carcinoma

PURPOSE: This study was designed to determine whether loss of heterozygosity and/or microsatellite instability correlate with HIV infection and tumor recurrence after chemoradiation therapy in patients with squamous-cell carcinoma of the anus. BACKGROUND: The molecular mechanisms leading to the progression of HIV-related squamous-cell carcinoma of the anus are poorly understood. In particular, genetic alterations responsible for resistance to chemoradiation have important clinical and functional implications. METHODS: In a case-control study, we analyzed normal and tumor DNA samples of four patients with squamous-cell carcinoma of the anus who were successfully treated with chemoradiotherapy and four patients with radio-resistant squamous-cell carcinoma of the anus who required abdominoperineal resection for local recurrence. To determine the presence of microsatellite instability, we used the reference panel of five pairs of microsatellite primers recommended for colorectal cancer specimens. These include the microsatellite markers BAT25, BAT26, D5S346 (APC), D2S123 (hMSH2), and D17S250 (P53). In addition, we used microsatellite markers for loss of heterozygosity analyses that were tightly linked to tumor suppressor genes. These included D3S1611 (hMLH1), D17S513 (P53), D18S46 and 18qTA (DCC/SMAD4), D5S107 (APC), and CA5 (hMSH2). RESULTS: There were two HIV-positive and two HIV-negative patients in each group. Three HIV-positive patients (one in the chemoradiotherapy group and two in the nonchemoradiotherapy group) demonstrated loss of heterozygosity. In the chemoradiotherapy group, one HIV-positive patient demonstrated loss of heterozygosity at the hMLH1 locus. In the nonchemoradiotherapy group, two HIV-positive patients exhibited a total of four instances of loss of heterozygosity. One tumor had loss of heterozygosity at hMSH2 and DCC/SMAD4; another tumor demonstrated loss of heterozygosity at hMSH2 and APC. Microsatellite instability-low was found in two HIV-positive patients. No instances of loss of heterozygosity and microsatellite instability were detected in HIV-negative patients. CONCLUSION: Loss of heterozygosity and microsatellite instability, which reflect inactivation of tumor-suppressor genes and genomic instability, occur with increased frequency in HIV-associated squamous-cell carcinoma. These data demonstrate for the first time evidence of loss of heterozygosity at the APC and DCC/SMAD4 gene loci in anal carcinoma. Although the findings presented here need to be expanded in a larger study, the recurrent loss of heterozygosity at D2S123, which was demonstrated in HIV-positive patients with radio-resistant squamous-cell carcinoma of the anus, is notable.Supported by a grant from the SICPA foundation and an educational grant from the Eleanor Naylor Dana Charitable Trust Fund.     

APC and CTNNB1 mutations in a large series of sporadic colorectal carcinomas stratified by the microsatellite instability status

BACKGROUND: Adenomatous polyposis coli (APC) and beta-catenin (encoded by CTNNB1) are important components in the WNT signalling pathway, a pathway altered in nearly all colorectal tumours. Conflicting results are reported on whether APC mutations are less common in tumours with a high degree of microsatellite instability (MSI-H) than in microsatellite stable (MSS) ones, and whether mutations in the regulatory domain of CTNNB1 substitute for APC mutations in the MSI-H tumours. METHODS: A consecutive series of 218 primary colorectal carcinomas, stratified by MSI status, were analysed for mutations in the APC gene (by the protein truncation test) and in the CTNNB1 gene (by single-strand conformation polymorphism). RESULTS: APC mutations detected in 66% of the patients were significantly more frequent in the MSS and MSI-L (low) tumours than in the MSI-H tumour group (P < 0.001). The MSI-H tumours tended to have more frameshift mutations than the MSS/MSI-L tumours. The majority of the APC mutations were located in the mutation cluster region (MCR). Patients that had lost all beta-catenin binding sites of the APC gene showed a shorter survival time than patients who retained some or all of these binding sites (P = 0.045). Two mutations were found in the CTNNB1 gene, but neither of them was located in the regulatory domain in exon 3. CONCLUSION: This study confirms that APC mutations are less frequent in MSI-H tumours than in MSS and MSI-L tumours. However, CTNNB1 mutations do not substitute for APC mutations in MSI-H tumours in these Norwegian patients.     

IL-1β and IL-8, matrix metalloproteinase 3, and pepsinogen secretion before and after H. pylori eradication in gastroduodenal phenotypes

Background: Adenomatous polyposis coli (APC) and β -catenin (encoded by CTNNB1 ) are important components in the WNT signalling pathway, a pathway altered in nearly all colorectal tumours. Conflicting results are reported on whether APC mutations are less common in tumours with a high degree of microsatellite instability (MSI-H) than in microsatellite stable (MSS) ones, and whether mutations in the regulatory domain of CTNNB1 substitute for APC mutations in the MSI-H tumours. Methods: A consecutive series of 218 primary colorectal carcinomas, stratified by MSI status, were analysed for mutations in the APC gene (by the protein truncation test) and in the CTNNB1 gene (by single-strand conformation polymorphism). Results: APC mutations detected in 66% of the patients were significantly more frequent in the MSS and MSI-L (low) tumours than in the MSI-H tumour group ( P < 0.001). The MSI-H tumours tended to have more frameshift mutations than the MSS/MSI-L tumours. The majority of the APC mutations were located in the mutation cluster region (MCR). Patients that had lost all β -catenin binding sites of the APC gene showed a shorter survival time than patients who retained some or all of these binding sites ( P = 0.045). Two mutations were found in the CTNNB1 gene, but neither of them was located in the regulatory domain in exon 3. Conclusion: This study confirms that APC mutations are less frequent in MSI-H tumours than in MSS and MSI-L tumours. However, CTNNB1 mutations do not substitute for APC mutations in MSI-H tumours in these Norwegian patients.     

DNA microsatellite instability and mismatch repair protein loss in adenomas presenting in hereditary non-polyposis colorectal cancer

BACKGROUND AND AIM: Hereditary non-polyposis colorectal cancer (HNPCC), as its name implies, is associated with few adenomas, and the early evolution of colorectal neoplasia is poorly understood. In this study our aim was to clarify the genetic profiles of benign polyps in subjects with HNPCC using a combined molecular and immunohistochemical approach. METHODS: Thirty adenomas and 17 hyperplastic polyps were obtained from 24 affected HNPCC subjects. DNA was extracted from paraffin embedded tissue by microdissection and analysed for the presence of microsatellite instability (MSI) and mutations in five genes known to be targets in mismatch repair deficiency (TGFbetaRII, IGF2R, BAX, hMSH3, and hMSH6). Serial sections were stained by immunohistochemistry for hMLH1 and hMSH2. RESULTS: Twenty four (80%) of 30 adenomas showed MSI. Of MSI positive adenomas, 66.7% showed MSI at more than 40% of markers (high level of MSI (MSI-H)). Two of 17 hyperplastic polyps revealed MSI at one marker (low level of MSI (MSI-L)). A significant association was found between MSI-H and high grade dysplasia in adenomas (p=0.004). Eight of nine adenomas with mutations of coding sequences revealed high grade dysplasia and all nine were MSI-H. Four of the nine ranged in size from 2 to 5 mm. The presence of the hMSH6 mutation was significantly correlated with high levels of MSI (80% of markers) (p<0.02). Twenty four adenomas gave evaluable results with immunohistochemistry. One of six (17%) microsatellite stable, six of seven (86%) MSI-L, and 11 of 11 (100%) MSI-H adenomas showed loss of either hMLH1 or hMSH2. CONCLUSIONS: Most adenomas in subjects with a definite diagnosis of HNPCC show MSI (80%). The finding of MSI-L is usually associated with loss of expression of hMLH1 or hMSH2, unlike the situation in MSI-L sporadic colorectal cancer. The transition from MSI-L to MSI-H correlated with the finding of high grade dysplasia and mutation of coding sequences and may be driven by mutation of secondary mutators such as hMSH3 and hMSH6. Advanced genetic changes may be present in adenomas of minute size.     

MSI is frequently recognized among gastric cancer patients with a family history of cancer

BACKGROUND/AIMS: A family history of cancer raises the risk of gastric cancer. Microsatellite instability (MSI) is frequently observed in hereditary non-polyposis colorectal cancer (HNPCC), and gastric cancer is the most frequent extra-colonic cancer among HNPCC patients. The relationship between gastric cancer and MSI is controversial. The purpose of this study was to identify the relationship between MSI incidence and gastric cancer in patients with a family history of cancer. METHODOLOGY: MSI incidence was examined at 5 microsatellite loci and hMLH1 and hMSH2 protein expression was examined by immunohistochemical analysis in 30 gastric cancer patients with family histories of cancer (familial group) and 37 gastric cancer patients without any family history of cancer (sporadic group). RESULTS: The incidence of high-frequency MSI (MSI-H) in the familial group was 33.3% (10/30) and in the sporadic group it was 5.4% (2/37) (p < 0.05). The incidence of lack of at least one mismatch repair (MMR) protein was 66.7% (8/12) in MSI-H cases, which was significantly higher than in low-frequency MSI (MSI-L) cases and microsatellite stable (MSS) cases. CONCLUSIONS: MSI may play an important role in the development of gastric cancer in individuals with a family history of cancer and it is induced by a deficiency in MMR protein expression.