Heterozygous mutations in PMS2 cause hereditary nonpolyposis colorectal carcinoma (Lynch syndrome)

BACKGROUND & AIMS: The role of the mismatch repair gene PMS2 in hereditary nonpolyposis colorectal carcinoma (HNPCC) is not fully clarified. To date, only 7 different heterozygous truncating PMS2 mutations have been reported in HNPCC-suspected families. Our aim was to further assess the role of PMS2 in HNPCC. METHODS: We performed Southern blot analysis in 112 patients from MLH1-, MSH2-, and MSH6-negative HNPCC-like families. A subgroup (n = 38) of these patients was analyzed by denaturing gradient gel electrophoresis (DGGE). In a second study group consisting of 775 index patients with familial colorectal cancer, we performed immunohistochemistry using antibodies against MLH1, MSH2, MSH6, and PMS2 proteins. In 8 of 775 tumors, only loss of PMS2 expression was found. In these cases, we performed Southern blot analysis and DGGE. Segregation analysis was performed in the families with a (possibly) deleterious mutation. RESULTS: Seven novel mutations were identified: 4 genomic rearrangements and 3 truncating point mutations. Three of these 7 families fulfill the Amsterdam II criteria. The pattern of inheritance is autosomal dominant with a milder phenotype compared with families with pathogenic MLH1 or MSH2 mutations. Microsatellite instability and immunohistochemical analysis performed in HNPCC-related tumors from proven carriers showed a microsatellite instability high phenotype and loss of PMS2 protein expression in all tumors. CONCLUSIONS: We show that heterozygous truncating mutations in PMS2 do play a role in a small subset of HNPCC-like families. PMS2 mutation analysis is indicated in patients diagnosed with a colorectal tumor with absent staining for the PMS2 protein.     

Germline mutations of EXO1 gene in patients with hereditary nonpolyposis colorectal cancer (HNPCC) and atypical HNPCC forms

BACKGROUND & AIMS: Germline mutations in one of four mismatch repair genes have been found in the majority of families with hereditary nonpolyposis colorectal cancer (HNPCC), but only in a small part of families with atypical HNPCC. The recently cloned EXO1 gene might be involved in the pathogenesis of HNPCC because the EXO1 protein strongly interacts with the MSH2 protein. To determine its role in HNPCC, EXO1 was scanned for germline mutations. METHODS: All 14 exons of EXO1 were scanned for mutations in index patients from 33 families with HNPCC fulfilling the Amsterdam criteria and in 225 index patients suspected of HNPCC. RESULTS: Germline variants of EXO1 were detected in 14 patients, including one splice-site mutation in a family with HNPCC and 13 missense mutations in patients with atypical HNPCC. These variants did not occur in more than 200 control individuals. From 13 of these 14 patients, tumors were available for analysis of microsatellite instability and loss of heterozygosity. Six of the tumors showed microsatellite instability. Heterozygosity analysis showed one case without EXO1 allelic loss and 12 tumors with loss of the mutant allele and retention of the normal one. CONCLUSIONS: The results indicate a possible association of germline EXO1 variants with HNPCC and atypical HNPCC.     

Aetiology of colorectal cancer and relevance of monogenic inheritance

BACKGROUND AND AIMS: Although diet and lifestyle are associated with the development of colorectal malignancies, the only clearly identified aetiological factors in colorectal cancer are inheritance (hereditary non-polyposis colorectal cancer (HNPCC) and familial polyposis), inflammatory bowel diseases, papillomavirus, and acquired immunodeficiency syndrome (AIDS). Our aim was to determine what proportion of colorectal neoplasms could be attributed to these specific factors. PATIENTS AND METHODS: Data from a colorectal cancer registry were analysed over a 15 year period, during which nearly 2500 cases were recorded. In patients with suspected HNPCC, microsatellite instability and immunohistochemical expression of proteins encoded by the main DNA mismatch repair genes were assessed. In families with unstable neoplasms, constitutional mutations of the mismatch repair genes hMSH2, hMLH1, and hMSH6 were evaluated by single strand conformation polymorphism analysis and sequencing. RESULTS: Inflammatory bowel diseases, familial polyposis, and AIDS were rare causes of colorectal cancer (three, three, and one case, respectively). Anal squamous carcinoma developed in 27 patients (1.0%) and could be attributed to papillomavirus infection. In 58 patients (from 34 families) a clinical diagnosis of HNPCC was established (2.4%). In total, cases with a known aetiology were 92 (3.7% of all patients). Microsatellite instability was detected in 15 cancers from HNPCC families, and germline mutations in six families (12 patients, 0.5% of the total). Families with unstable tumours, with or without mutations, were clinically similar, suggesting the involvement of the mismatch repair system even when mutations were not detected. CONCLUSIONS: The study suggests that the aetiology of colorectal malignancies remains elusive in the large majority of cases. Among specific causes, HNPCC represents the most frequent. However, with a population based approach, constitutional mutations of the main genes involved in HNPCC can be detected in only 20% of cases.     

First report of a de novo germline mutation in the MLH1 gene

Hereditary non-polyposis colorectal carcinoma (HNPCC) is an autosomal dominant disorder associated with colorectal and endometrial cancer and a range of other tumor types. Germline mutations in the DNA mismatch repair (MMR) genes, particularly MLH1, MSH2, and MSH6, underlie this disorder. The vast majority of these HNPCC-associated mutations have been proven, or assumed, given the family history of cancer, to be transmitted through several generations. To the best of our knowledge, only a single case of a de novo germline MMR gene mutation (in MSH2) has been reported till now. Here, we report a patient with a de novo mutation in MLH1. We identified a MLH1 Q701X truncating mutation in the blood lymphocytes of a male who had been diagnosed with rectal cancer at the age of 35. His family history of cancer was negative for the first- and second-degree relatives. The mutation could not be detected in the patient' parents and sibling and paternity was confirmed with a set of highly polymorphic markers. Non-penetrance and small family size is the common explanation of verified negative family histories of cancer in patients with a germline MMR gene mutation. However, in addition to some cases explained by non-paternity, de novo germline mutations should be considered as a possible explanation as well. As guidelines that stress not to restrict MMR gene mutation testing to patients with a positive family history are more widely introduced, more cases of de novo MMR gene germline mutations may be revealed.     

No Major Difference in K-ras and p53 Abnormalities in Sporadic and Hereditary Nonpolyposis Colorectal Adenomas

K-ras and p53 gene mutations are known to occur in high frequencies in sporadic colorectal cancers, but findings are inconsistent in hereditary nonpolyposis colorectal cancer (HNPCC). We compared K-ras codon 12 and 13 gene mutations and p53 protein overexpression in 48 HNPCC (positive for Amsterdam criteria) and 59 sporadic colorectal adenomas, to examine whether they may represent similar or different molecular pathways to cancer. In sporadic adenomas K-ras mutations were detected in 32% and p53 overexpression in 31% of the cases. Similarly, K-ras mutations and p53 overexpression were both found in 25% of HNPCC adenomas. The frequencies of these abnormalities were not significantly different between HNPCC and sporadic adenomas. When taking differences in adenoma size into account, the frequencies were even more similar. In conclusion, these results suggest a similar molecular pathway to adenomas in HNPCC and sporadic carcinogenesis, with respect to involvement of K-ras and p53.     

Mutator phenotype in a subset of chronic lymphocytic leukemia

The replication error phenotype (RER+), characterized by widespread microsatellite instability, is an important feature of tumors from patients with hereditary nonpolyposis colorectal carcinoma (HNPCC). This widespread instability affects repeat tracts of all lengths and is usually attributed to mutations of critical mismatch repair genes. Recently, several reports described occasional microsatellite alterations in tumors not associated with HNPCC. However, a true mutator phenotype (RER+) is very rare outside of HNPCC-associated malignancies. We examined 29 cases of chronic lymphocytic leukemia (CLL), the most common leukemia in the Western world for evidence of microsatellite instability. We identified a mutator phenotype in (2/29) 7% of the cases studied. These data suggest that the mismatch repair pathway may be altered in at least a subset of patients with CLL.     

Hereditary non-polyposis colorectal cancer: clinical and molecular evidence for a new entity of hereditary colorectal cancer

BACKGROUND: Hereditary non-polyposis colorectal cancer (HNPCC) is clinically defined by familial clustering of colorectal cancer and other associated tumours. METHODS: By thorough molecular and clinical evaluation of 41 families, two different groups were characterised: group 1, 25 families with truncating mutations in MLH1 or MSH2 (12 novel mutations); and group 2, 16 Amsterdam positive families without mutations in these genes and without microsatellite instability in their corresponding tumours. RESULTS: Significant clinical differences between these two groups were found. Firstly, earlier age of onset for all colorectal cancers (median 41 v 55 years; p < 0.001) and all tumours (median 43 v 56 years; p = 0.022) was observed, comparing groups 1 and 2. Secondly, 68% of the index colorectal cancers were localised proximally of the splenic flexure in group 1 compared with 14% in group 2 (p < 0.010). Thirdly, more synchronous and metachronous colorectal (p = 0.017) and extracolorectal tumours (p < 0.001) were found in group 1. Fourthly, a higher colorectal adenoma/carcinoma ratio (p = 0.030) and a tendency towards more synchronous or metachronous adenomas in group 2 (p = 0.084) was observed, indicating a slower progression of adenomas to carcinomas. As three mutation negative tumours revealed chromosomal instability after comparative genomic hybridisation, these tumours may be caused by one or more highly penetrant disease alleles from the chromosomal instability pathway. CONCLUSION: These data show that HNPCC includes at least two entities with clinical and molecular differences. This will have implications for surveillance programmes and for cancer research.     

Does Mutation of Transforming Growth Factor-β Type II Receptor Gene Play an Important Role in Colorectal Polyps?

Mutations in the transforming growthfactor- type II receptor (RII) gene that remainuncorrected due to mutation and inactivation of mismatchrepair genes play an important role in hereditarynonpolyposis colorectal cancer (HNPCC) and in a subset ofsporadic colorectal cancers. Some colorectal cancersdevelop from colorectal polyps. To elucidate the role ofthe RII gene in the generation of colorectal polyps, we analyzed 137 colorectal polyps from 100patients for RII mutations and microsatelliteinstability (MSI). MSI was detected in three of 36polyps from 25 patients. For one of these three polyps,the mobilities of the PCR products between polyp and nonpolyptissues was different for only one microsatellitemarker, and for the other two polyps the mobilities weredifferent for more than two markers. These two polyps were obtained from one patient with ascendingcolon carcinoma and suspected HNPCC based on hisclinical profile and family history. An RII mutation wasdetected in only one of these two polyps. RII may play a minor role in sporadic colorectalpolyps. RII gene analysis in colorectal polyps may be auseful screening measure for potential HNPCCpatients.     

Screening for hereditary non-polyposis colorectal cancer: a study of 22 kindreds in The Netherlands

PURPOSE: Identification of the hereditary non-polyposis colorectal carcinoma syndrome (HNPCC) is a basis for secondary prevention. The objectives of this study are to investigate the natural history of HNPCC and to assess the effect of screening. PATIENTS AND METHODS: Screening for colorectal carcinoma was performed in 22 HNPCC families (colonoscopy or double-contrast barium enema and sigmoidoscopy). The patients were subdivided into two groups. Group A comprised patients with colorectal cancer who were referred because they were symptomatic. Group B included family members of these patients who were found to have a colorectal lesion by screening. We compared these groups with respect to the stage of tumor growth. RESULTS: Histologic examination of the tumors in Group A (87 patients) revealed Dukes A carcinomas in six patients, Dukes B carcinomas in 37, Dukes C carcinomas in 21, and Dukes D carcinomas in 10 patients (classification unknown in 13 patients). In Group B (20 patients), adenoma was found in 14 and carcinoma in six patients (Dukes A in two and Dukes B in four patients). A total of 93 patients, including those whose tumors were detected by screening, had a colorectal carcinoma. The age at diagnosis ranged from 24 to 81 years (mean age: 46 years). The location of the colonic tumors was proximal in 60 percent. Multiple primary tumors were found in 26 percent. CONCLUSION: These results suggest that screening leads to the early detection of colorectal carcinomas and adenomas in asymptomatic members of HNPCC families. Screening should be initiated at the age of 20 and continued during the life of the individual. Careful examination of the right colon is indicated because of the frequent occurrence of tumors in the proximal colon. A subtotal colectomy is indicated at the time of diagnosis of the initial colon cancer because of the risk of multiple primary tumors.     

Exon 3 beta-catenin mutations are specifically associated with colorectal carcinomas in hereditary non-polyposis colorectal cancer syndrome

BACKGROUND AND AIMS: Activating beta-catenin mutations in exon 3 have been implicated in colorectal tumorigenesis. Although reports to the contrary exist, it has been suggested that beta-catenin mutations occur more often in microsatellite unstable (MSI+) colorectal carcinomas, including hereditary non-polyposis colorectal cancer (HNPCC), as a consequence of defective DNA mismatch repair. We have analysed 337 colorectal carcinomas and adenomas, from both sporadic cases and HNPCC families, to provide an accurate assessment of beta-catenin mutation frequency in each tumour type. METHODS: Direct sequencing of exon 3 of beta-catenin. RESULTS: Mutations were rare in sporadic (1/83, 1.2%) and HNPCC adenomas (1/37, 2.7%). Most of the sporadic adenomas analysed (80%) were small (<1 cm), and our data therefore differ from a previous report of a much higher mutation frequency in small adenomas. No oncogenic beta-catenin mutations were identified in 34 MSI+ and 78 microsatellite stable (MSI-) sporadic colorectal cancers but a raised mutation frequency (8/44, 18.2%) was found in HNPCC cancers; this frequency was significantly higher than that in HNPCC adenomas (p=0.035) and in both MSI- (p<0.0001) and MSI+ (p=0.008) sporadic cancers. Mutations were more common in higher stage (Dukes' stages C and D) cancers (p=0.001). CONCLUSION: Exon 3 beta-catenin mutations are associated specifically with malignant colorectal tumours in HNPCC; mutations appear not to result directly from deficient mismatch repair. Our data provide evidence that the genetic pathways of sporadic MSI+ and HNPCC cancers may be divergent, and indicate that mutations in the HNPCC pathway of colorectal tumorigenesis may be determined by selection, not simply by hypermutation.     

Reduced expression of cyclooxygenase 2 proteins in hereditary nonpolyposis colorectal cancers relative to sporadic cancers.

BACKGROUND & AIMS: Cyclooxygenase (COX) enzymes catalyze the conversion of arachidonic acid to prostaglandins. Evidence suggests that nonsteroidal anti-inflammatory drugs reduce the risk of colorectal cancer (CRC) and that this effect is mediated through COX inhibition. We analyzed and compared expression of the inducible COX-2 isoform in colorectal neoplasms from patients with hereditary nonpolyposis colorectal cancer (HNPCC), familial adenomatous polyposis (FAP), and sporadic CRC. Given that COX-2 is induced by transforming growth factor (TGF)-beta and that TGF-beta type II receptor (RII) mutations are found in HNPCCs, we determined the relationship between RII status and COX-2 expression. METHODS: COX-2 protein expression was determined in colorectal epithelia using immunohistochemistry and Western blotting. Patients with HNPCC had known mutations in hMLH1 or hMSH2 genes and/or met the Amsterdam criteria. In CRCs from HNPCC cases, mutations were sought in the coding region of the RII gene using the polymerase chain reaction. RESULTS: COX-2 was detected in adenomas from 2 of 3 HNPCC, 6 of 7 FAP, and 5 of 8 sporadic cases. In CRCs, COX-2 staining was found in 16 of 24 (67%) HNPCC vs. 24 of 26 (92%) sporadic cases (P = 0.035) and in 2 of 2 FAP cases. Staining intensity was reduced in HNPCCs compared with sporadic CRCs (P = 0.035). Staining localized to the cytoplasm of neoplastic cells; normal epithelial cells were negative for COX-2. Overexpression of COX-2 in CRCs relative to normal mucosa was confirmed by Western blotting. TGF-beta RII mutations were detected in 12 of 14 HNPCCs examined, including 3 of 4 COX-2-negative and 9 of 10 COX-2-positive cancers. CONCLUSIONS: The frequency and intensity of COX-2 expression was significantly reduced in HNPCCs relative to sporadic CRCs, and was not a consequence of RII mutations. Given that many HNPCCs express COX-2, inhibition of this enzyme may be an important strategy to prevent CRC in these patients.     

hMLH1和hMSH2蛋白表达在遗传性非息肉病性大肠癌诊断中的应用

目的　评价错配修复基因hMLH1和hMSH2蛋白表达在筛选遗传性非息肉病性大肠癌(HNPCC)中的价值。方法　收集大肠癌患者 6 6例 ,分为HNPCC患者 (A组 ,n =19)、高度可疑HNPCC患者 (B组 ,n =2 0 )、符合Bethesda指导标准的可疑HNPCC患者 (C组 ,n =14 )及散发性大肠癌患者 (D组 ,n =13)四组 ,用免疫组化方法检测各组错配修复基因hMLH1和hMSH2的蛋白表达。结果　A组hMLH1和hMSH2蛋白表达减低或缺失达 72 .8% ;B组为 6 0 .0 % ;C组为 2 8.4 % ;D组为 7.7%。hMLH1和hMSH2蛋白表达减低或缺失与HNPCC显著相关 (P =0 .0 0 0 8)。此外 ,hMLH1蛋白表达减低或缺失率显著高于hMSH2 (P <0 .0 1)。结论　hMLH1和hMSH2蛋白表达减低或缺失与HNPCC的可能性显著相关 ,免疫组化检测此二种蛋白表达能快速、有效地帮助临床医生评估患者HNPCC的可能性 ,同时提示相应错配修复基因存在突变。中国HNPCC患者中hMLH1基因发生突变的机会可能高于hMSH2基因。     

Analysis of masked mutations in familial adenomatous polyposis

Familial adenomatous polyposis (FAP) is an autosomal-dominant disease characterized by the development of hundreds of adenomatous polyps of the colorectum. Approximately 80% of FAP patients can be shown to have truncating mutations of the APC gene. To determine the cause of FAP in the other 20% of patients, MAMA (monoallelic mutation analysis) was used to independently examine the status of each of the two APC alleles. Seven of nine patients analyzed were found to have significantly reduced expression from one of their two alleles whereas two patients were found to have full-length expression from both alleles. We conclude that more than 95% of patients with FAP have inactivating mutations in APC and that a combination of MAMA and standard genetic tests will identify APC abnormalities in the vast majority of such patients. That no APC expression from the mutant allele is found in some FAP patients argues strongly against the requirement for dominant negative effects of APC mutations. The results also suggest that there may be at least one additional gene, besides APC, that can give rise to FAP.     

Family history and molecular features of children, adolescents, and young adults with colorectal carcinoma

BACKGROUND: Colorectal cancer is extremely rare in childhood. Published case series reporting children and adolescents with colorectal cancer have not focused on the underlying genetic aspects of the tumour or genetic susceptibility of the families. AIMS: We examined a cohort of patients with early onset colorectal cancer to determine whether a specific genetic predisposition could be elucidated. In particular, we focused on whether DNA mismatch repair gene deficiency which causes hereditary non-polyposis colorectal cancer (HNPCC) could be elucidated. METHODS: Patients with colorectal cancer 相似文献   

Hereditary Nonpolyposis Colorectal Cancer (Lynch Syndrome): Criteria for Identification and Management

Hereditary nonpolyposis colorectal carcinoma (HNPCC), or Lynch syndrome, is an autosomal dominant syndrome accounting for 5 to 10% of the total colorectal cancer population. Patients with this syndrome develop colorectal carcinoma at an early age, but disease onset can happen in all age groups. Usually the carcinomas are synchronous or metachronous, and most of them arise proximal to the splenic flexure. The prognosis is better than for the sporadic form of cancer, and there is increased risk for cancer development in certain extracolonic sites, such as the endometrium, ovary, stomach, small bowel, hepatobiliary tract, ureter, and renal pelvis. Most patients with HNPCC have a mutation in one of two DNA mismatch repair genes, hMSH2 or hMLH1. More than 90% of colorectal carcinoma patients with hMSH2 or hMLH1 demonstrate high-frequency microsatellite instability (MSI-H). If a patient is suspected to belong to an HNPCC family, the first screening test should be immunohistochemistry for the detection of hMLH1 and hMSH2 proteins, and if it is indicative, it should be followed by genomic sequencing for the identification of mutations in the mismatch repair genes. Genetic counseling and surveillance for high risk HNPCC family members should begin at age 25. Surveillance includes annual colonoscopy of the entire large bowel, with fecal occult blood testing performed twice a year. Systematic surveillance and individually designed treatment of affected patients may help to detect cancers at an earlier stage and subsequently improve the prognosis of the disease further.     

Prevalence of defective DNA mismatch repair and MSH6 mutation in an unselected series of endometrial cancers

Endometrial cancer is the most common gynecologic malignancy in the United States and the most frequent extracolonic tumor in hereditary nonpolyposis colorectal cancer (HNPCC). HNPCC patients have inherited defects in DNA mismatch repair and the microsatellite instability (MSI) tumor phenotype. Sporadic endometrial cancers also exhibit MSI, usually associated with methylation of the MLH1 promoter. Germ-line MSH6 mutations, which are rare in HNPCC, have been reported in several families with multiple members affected with endometrial carcinoma. We reasoned that MSH6 mutation might account for loss of mismatch repair in MSI-positive endometrial cancers in which the cause of MSI is unknown. We therefore investigated MSI and MLH1 promoter methylation in 441 endometrial cancer patients unselected for age or personal and family history of cancers. MSI and MLH1 promoter methylation status were associated with age of onset and tumor histology. One hundred cases (23% of the entire series) were evaluated for MSH6 defects. Inactivating germ-line MSH6 mutations were identified in seven women with MSI-positive, MLH1 promoter unmethylated cancers. Most of the MSI in these cases was seen with mononucleotide repeat markers. The MSH6 mutation carriers were significantly younger than the rest of the population (mean age 54.8 versus 64.6, P = 0.04). Somatic mutations were seen in 17 tumors, all of which had MSI. Our data suggest that inherited defects in MSH6 in women with endometrial cancer are relatively common. The minimum estimate of the prevalence of inherited MSH6 mutation in endometrial cancer is 1.6% (7 of 441), comparable with the predicted prevalence for patients with colorectal cancer.     

Clinical and genetic characteristics of Chinese hereditary nonpolyposis colorectal cancer families

AIM: To analyze the clinical characteristics of Chinese hereditary nonpolyposis colorectal cancer (HNPCC) families and to screen the germline mutations of human mismatch repair genes hMLH1 and hMSH2 in the probands.METHODS: Thirty-one independent Chinese HNPCC families were collected in Zhejiang Province. All of them met Chinese HNPCC criteria. Clinical data about patient gender, site of colorectal cancer, age of onset, history of multiple colorectal cancer, associated extracolonic cancer were recorded. PCR and denaturing high performance liquid chromatography (DHPLC) were employed to screen the mutations. Sequencing analysis was used to find out the exact mutation site and characteristics of the samples showing abnormal DHPLC profiles.RESULTS: One hundred and thirty-six malignant neoplasms were found in 107 patients including 14 multiple cancers. One hundred and six of the 136 neoplasms (77.9%) were diagnosed as colorectal cancer, with an average age of onset at 48.57 ± 29.00 years. Gastric cancer was the most common extracolonic cancer (10.3%) in these families. Twenty-three different sequence variations in hMLH1 and hMSH2 genes were detected in these 17 families. Fifteen sequence variations were located in the exons, including 5 SNPs, 3 silent mutations, 3 missense mutations, 2 nonsense mutations and 2 frameshift mutations. The latter seven mutations seemed to be pathogenic.CONCLUSION: Germline mutations of hMLH1 and hMSH2 genes are identified in about one-third HNPCC kindreds fulfilling Chinese HNPCC criteria. Chinese HNPCC families have some particular clinical characteristics, such as a left-sided predominance, less synchronous or metachronous colorectal cancer, and frequent occurrence of gastric cancer.     

Frequency of hereditary non-polyposis colorectal cancer in Danish colorectal cancer patients.

BACKGROUND: Hereditary non-polyposis colorectal cancer (HNPCC) is an autosomal dominant cancer syndrome, characterised by familial aggregation of HNPCC related cancers, germline mutations in mismatch repair genes, and/or microsatellite instability (MSI) in tumour tissue. AIM: To estimate the frequency of HNPCC among non-selected Danish patients with colorectal cancer (CRC), and to evaluate the value of MSI analysis as a pre-screen test. METHODS: This was a prospective population based study on consecutive CRC patients. A family history of malignancy was obtained and suspected HNPCC cases were screened for hMLH1/hMSH2 mutations and subjected to MSI analysis. Patients with germline mutations and/or those with Amsterdam criteria I or II families were categorised as HNPCC patients. RESULTS: Among 1328 eligible CRC patients, 1200 (90.4%) completed a questionnaire. A total of 1.7% (95% confidence interval (CI) 1.0-2.4) (20 cases) were categorised as HNPCC patients. Amsterdam criteria I or II were met in 18 cases (1.5%), and in another two cases (0.2%) pathogenic hMLH1/hMSH2 mutations were detected without fulfillment of the Amsterdam criteria I or II. Among 77 patients younger than 50 years of age, 11 cases (14.3%) were categorised as HNPCC. The Amsterdam criteria I or II were met in eight of 10 gene carriers (80%). The MSI-high phenotype was demonstrated in all 10 gene carriers. CONCLUSION: The frequency of HNPCC was approximately 1.7% among all CRC cases and 14.3% among patients younger than 50 years of age. MSI analysis is a reliable pre-screen test for hMLH1/hMSH2 mutations in families suspected of having HNPCC.     

Low prevalence of germline hMSH6 mutations in colorectal cancer families from Spain

AIM: To investigate the prevalence and penetrance of hMSH6 mutations in Spanish HNPCC families that was negative for mutation in hMLH1 or hMSH2. METHODS: We used PCR-based DGGE assay and direct sequencing to screen for hMSH6 gene in 91 HNPCC families. RESULTS: we have identified 10 families with germ-line mutations in the DNA sequence. These mutations included two intronic variation, three missense mutation, one nonsense mutation, and four silent mutations. Among the 10 germ-line mutations identified in the Spanish cohort, 8 were novel, perhaps, suggesting different mutational spectra in the Spanish population. Detailed pedigrees were constructed for the three families with a possible pathogenic hMSH6 mutation. The two silent mutations H388H and L758L, detected in a person affected of colorectal cancer at age 29, produce loss of the wild-type allele in the tumor sample. Immunohistochemical analysis showed that expression of MSH6 protein was lost only in the tumors from the carriers of V878A and Q263X mutations. CONCLUSION: Altogether, our results indicate that disease-causing germ-line mutations of hMSH6 are very less frequent in Spanish HNPCC families.