Mutation in the DNA mismatch repair gene homologue hMLH1 is associated with hereditary non-polyposis colon cancer

The human DNA mismatch repair gene homologue hMSH2, on chromosome 2p is involved in hereditary non-polyposis colon cancer (HNPCC). On the basis of linkage data, a second HNPCC locus was assigned to chromosome 3p21-23 (ref. 3). Here we report that a human gene encoding a protein, hMLH1 (human MutL homologue), homologous to the bacterial DNA mismatch repair protein MutL, is located on human chromosome 3p21.3-23. We propose that hMLH1 is the HNPCC gene located on 3p because of the similarity of the hMLH1 gene product to the yeast DNA mismatch repair protein, MLH1, the coincident location of the hMLH1 gene and the HNPCC locus on chromosome 3, and hMLH1 missense mutations in affected individuals from a chromosome 3-linked HNPCC family.     

The Saccharomyces cerevisiae MLH3 gene functions in MSH3-dependent suppression of frameshift mutations

The Saccharomyces cerevisiae genome encodes four MutL homologs. Of these, MLH1 and PMS1 are known to act in the MSH2-dependent pathway that repairs DNA mismatches. We have investigated the role of MLH3 in mismatch repair. Mutations in MLH3 increased the rate of reversion of the hom3-10 allele by increasing the rate of deletion of a single T in a run of 7 Ts. Combination of mutations in MLH3 and MSH6 caused a synergistic increase in the hom3-10 reversion rate, whereas the hom3-10 reversion rate in an mlh3 msh3 double mutant was the same as in the respective single mutants. Similar results were observed when the accumulation of mutations at frameshift hot spots in the LYS2 gene was analyzed, although mutation of MLH3 did not cause the same extent of affect at every LYS2 frameshift hot spot. MLH3 interacted with MLH1 in a two-hybrid system. These data are consistent with the idea that a proportion of the repair of specific insertion/deletion mispairs by the MSH3-dependent mismatch repair pathway uses a heterodimeric MLH1-MLH3 complex in place of the MLH1-PMS1 complex.     

Evidence for involvement of yeast proliferating cell nuclear antigen in DNA mismatch repair

DNA mismatch repair plays a key role in the maintenance of genetic fidelity. Mutations in the human mismatch repair genes hMSH2, hMLH1, hPMS1, and hPMS2 are associated with hereditary nonpolyposis colorectal cancer. The proliferating cell nuclear antigen (PCNA) is essential for DNA replication, where it acts as a processivity factor. Here, we identify a point mutation, pol30-104, in the Saccharomyces cerevisiae POL30 gene encoding PCNA that increases the rate of instability of simple repetitive DNA sequences and raises the rate of spontaneous forward mutation. Epistasis analyses with mutations in mismatch repair genes MSH2, MLH1, and PMS1 suggest that the pol30-104 mutation impairs MSH2/MLH1/PMS1-dependent mismatch repair, consistent with the hypothesis that PCNA functions in mismatch repair. MSH2 functions in mismatch repair with either MSH3 or MSH6, and the MSH2-MSH3 and MSH2-MSH6 heterodimers have a role in the recognition of DNA mismatches. Consistent with the genetic data, we find specific interaction of PCNA with the MSH2-MSH3 heterodimer.     

Mutation of MSH3 in endometrial cancer and evidence for its functional role in heteroduplex repair

Many human tumours have length alterations in repetitive sequence elements. Although this microsatellite instability has been attributed to mutations in four DNA mismatch repair genes in hereditary nonpolyposis colorectal cancer (HNPCC) kindreds, many sporadic tumours exhibit instability but no detectable mutations in these genes. It is therefore of interest to identify other genes that contribute to this instability. In yeast, mutations in several genes, including RTH and MSH3, cause microsatellite instability. Thus, we screened 16 endometrial carcinomas with microsatellite instability for alterations in FEN1 (the human homolog of RTH) and in MSH3 (refs 12-14). Although we found no FEN1 mutations, a frameshift mutation in MSH3 was observed in an endometrial carcinoma and in an endometrial carcinoma cell line. Extracts of the cell line were deficient in repair of DNA substrates containing mismatches or extra nucleotides. Introducing chromosome 5, encoding the MSH3 gene, into the mutant cell line increased the stability of some but not all microsatellites. Extracts of these cells repaired certain substrates containing extra nucleotides, but were deficient in repair of those containing mismatches or other extra nucleotides. A subsequent search revealed a second gene mutation in HHUA cells, a missense mutation in the MSH6 gene. Together the data suggest that the MSH3 gene encodes a product that functions in repair of some but not all pre-mutational intermediates, its mutation in tumours can result in genomic instability and, as in yeast, MSH3 and MSH6 are partially redundant for mismatch repair.     

MLH1 and MSH2 constitutional mutations in colorectal cancer families not meeting the standard criteria for hereditary nonpolyposis colorectal cancer

Genetic diagnosis of hereditary nonpolyposis colorectal cancer (HNPCC) may have a significant impact on the clinical management of patients and their at-risk relatives. At present, clinical criteria represent the simplest and most useful method for the identification of HNPCC families and for the selection of candidates for genetic testing. However, reports of mismatch repair (MMR) gene mutations in families not fulfilling the minimal diagnostic criteria point out the necessity to identify additional clinical parameters suggestive of genetic predisposition to colorectal cancer (CRC) related to MMR defects. We thus investigated a series of 32 Italian putative HNPCC individuals selected on the basis of one of the following criteria: 1) family history of CRC and/or other extracolonic tumors; 2) early-onset CRC; and 3) presence of multiple primary malignancies in the same individual. These patients were investigated for the presence of MLH1 and MSH2 mutations by single-strand conformation polymorphism analysis. Pathogenetic truncating mutations were identified in 4 (12.5%) cases, 3 of them involving MSH2 and 1 MLH1. In addition, 2 missense MLH1 variants of uncertain significance were observed. All pathogenetic mutations were associated with early age (<40 years) at onset and proximal CRC location. Our results support the contention that constitutional MMR mutations can also occur in individuals without the classical HNPCC pattern. Moreover, evaluation of the clinical parameters associated with MMR mutations indicates that early onset combined with CRC location in the proximal colon can be definitely considered suggestive of MMR-related hereditary CRC and should be included among the guidelines for referring patients for genetic testing.     

Microsatellite instability in early onset and familial colorectal cancer

Hereditary non-polyposis colorectal cancer syndrome (HNPCC) is often considered to be the most common form of inherited colorectal cancer, although its precise incidence is unknown. The clinical diagnosis of HNPCC relies on a combination of family history and young age of onset of colorectal cancer, but as many familial aggregations of colorectal cancer do not fulfil the strict diagnostic criteria, HNPCC might be underdiagnosed. The majority of HNPCC families have germline mutations in mismatch repair (MMR) genes, such as MSH2 or MLH1, so that HNPCC cancers characteristically exhibit DNA replication errors (RERs) at microsatellite loci. Although an RER positive phenotype in tumours can also result from somatic mutations in an MMR gene, the prevalence of RER + tumours should provide a maximum estimate of the incidence of germline MMR gene mutations in patients with early onset and familial colorectal cancer. We investigated colorectal cancers for RERs from (1) a population based study of 33 patients with colorectal cancer aged 45 years or less, (2) 65 kindreds with familial colorectal cancer which only partially fulfilled the criteria for the diagnosis of HNPCC, and (3) 18 cancers from 12 HNPCC kindreds. Seven of 33 patients (21%) with colorectal cancer aged 45 years or less had an RER + cancer, with only two of these having a clear family history of HNPCC. A greater proportion of RER + tumours (5/7) occurred proximal to the splenic flexure than RER - tumours (4/26; chi2 = 6.14, p < 0.025). RERs were detected in all 18 cancers from HNPCC patients but in only six of 65 non-HNPCC familial colorectal cancer kindreds (9%; chi2 = 52.2, p < 0.0005). These findings suggest that most cancers in patients diagnosed at 45 years of age or less and familial aggregations of colorectal cancer which do not fulfil HNPCC diagnostic criteria do not have germline mutations in MSH2 and MLH1. Hence population screening for germline mutations in these genes is unlikely to be an efficient strategy for identifying people at high risk of developing colorectal cancer.     

Measuring quality of life in cardiac rehabilitation clients

OBJECTIVE: To clarify the origin of defective mismatch repair (MMR) in sporadic endometrial cancers with microsatellite instability (MSI), a thorough mutation analysis was performed on the human mismatch repair gene MSH3. METHODS: Twenty-eight MSI-positive endometrial cancers were investigated for mutations in the human mismatch repair gene MSH3 using single-strand conformation variant (SSCV) analysis of all 24 exons. All variants were sequenced. Loss of heterozygosity was investigated at all MSH3 polymorphisms discovered. A subset of tumors were investigated for methylation of the 5' promoter region of MSH3 using Southern blot hybridization. RESULTS: An identical single-base deletion (delta A) predicted to result in a truncated proteins was discovered in six tumors (21.4%). This deletion occurs in a string of eight consecutive adenosine residues (A8). Because simple repeat sequences are unstable in cells with defective MMR, the observed mutation may be an effect, rather than a cause, of MSI. Evidence of inactivation of the second MSH3 allele in tumors with the delta A mutation would strongly support a causal role for these MSH3 mutations. However, there was no evidence of a second mutation, loss of sequences, or methylation of the promoter region in any of the tumors with the delta A mutation. CONCLUSION: Although the delta A mutation is a frequent event in sporadic MSI-positive endometrial cancers, it may not be causally associated with defective DNA MMR.     

BAX frameshift mutations in cell lines derived from human haemopoietic malignancies are associated with resistance to apoptosis and microsatellite instability

Bax suppresses tumorigenesis in a mouse model system and Bax-deficient mice exhibit lymphoid hyperplasia suggesting that BAX functions as a tumour suppressor in human haemopoietic cells. We examined BAX expression in 20 cell lines derived from human haemopoietic malignancies and consistent with a potential tumour suppressor function, identified two cell lines, DG75 (a Burkitt lymphoma cell line) and Jurkat (a T-cell leukaemia line), which lacked detectable BAX expression. Apoptosis of DG75 cells induced by low serum or ionomycin was significantly delayed relative to similar Burkitt lymphoma cell lines with normal BAX levels. Although DG75 and Jurkat cells expressed several BAX RNA species including the prototypical BAX alpha RNA, the absence of BAX protein was due to single base deletions and additions in a polyguanine tract within the BAX open reading frame. These frameshift mutations result in premature termination of translation and have recently also been identified in some colon cancers with microsatellite instability. Although mismatch repair defects are not considered a common feature of haemopoietic malignancies, DG75 and Jurkat cells had widespread microsatellite instability and did not express detectable levels of MSH2. In Jurkat cells, lack of MSH2 expression was due to a point mutation in exon 13 of MSH2 resulting in premature termination of translation. Our results suggest that a pathway linking mismatch repair defects, BAX tumour suppressor frameshift mutations and resistance to apoptosis may be a key feature of some lymphomas and leukaemias.     

Usefulness of electron beam computed tomography scanning for distinguishing ischemic from nonischemic cardiomyopathy

DNA mismatch repair ensures genomic stability by correcting biosynthetic errors and by blocking homologous recombination. MutS-like and MutL-like proteins play important roles in these processes. In Escherichia coli and yeast these two types of proteins form a repair initiation complex that binds to mismatched DNA. However, whether human MutS and MutL homologs interact to form a complex has not been elucidated. Using immunoprecipitation and Western blot analysis we show here that human MSH2, MLH1, PMS2 and proliferating cell nuclear antigen (PCNA) can be co-immunoprecipitated, suggesting formation of a repair initiation complex among these proteins. Formation of the initiation complex is dependent on ATP hydrolysis and at least functional MSH2 and MLH1 proteins, because the complex could not be detected in tumor cells that produce truncated MLH1 or MSH2 protein. We also demonstrate that PCNA is required in human mismatch repair not only at the step of repair initiation, but also at the step of repair DNA re-synthesis.     

Hypermethylation of the hMLH1 promoter in colon cancer with microsatellite instability

Recent studies have demonstrated the presence of microsatellite instability (MSI) in tumors from patients with hereditary nonpolyposis colon cancer and in a subset of patients with sporadic colorectal cancer (CRC). In sporadic CRC, three tumor phenotypes have been defined: microsatellite stable (MSS), low-frequency MSI, and high-frequency MSI (MSI-H). Although defective mismatch repair, consisting primarily of alterations in hMSH2 and hMLH1, is believed to be responsible for the MSI phenotype in the majority of patients with hereditary nonpolyposis colon cancer, the genetic defect responsible for this phenotype in sporadic CRC has yet to be clearly delineated. Somatic or germ-line alterations in these two genes have been identified in only a minority of these cases. Analysis of the protein expression patterns of hMSH2 and hMLH1 in unselected CRC, however, suggests that alterations in hMLH1 may account for a majority of the MSI-H cases. In an effort to explore the underlying molecular basis for these findings, we have examined the methylation status of the presumptive hMLHI promoter region in 31 tumors that vary in regard to their MSI status (MSI-H or MSS), their hMLH1 protein expression (MLH- or MLH+), and their gene mutation (Mut+ or Mut-) status. Hypermethylation of the hMLH1 promoter occurred in all 13 MSI-H/ MLH- tumors that did not have a detectable mutation within the hMLH1 gene. Of those MSI-H tumors containing germ-line or somatic alterations in hMLH1 (n = 7, including 3 frameshift, 1 nonsense, 2 missense mutations, and 1 tumor containing multiple mutations: missense, splice-site alteration, and a frameshift), four had a normal methylation pattern, whereas three others demonstrated hypermethylation of the hMLH1 promoter region. Two of these cases had a missense alteration, the other a frameshift alteration. The single MSI-H/Mut+ tumor that had normal hMLH1 and hMSH2 expression, as well as 9 of the 10 MSS cases, lacked methylation of the hMLH1 promoter. Hypermethylation of the hMSH2 promoter was not observed for any of the cases. These results suggest that hypermethylation of the hMLH1 promoter may be the principal mechanism of gene inactivation in sporadic CRC characterized by widespread MSI.     

DNA mismatch repair and O6-alkylguanine-DNA alkyltransferase analysis and response to Temodal in newly diagnosed malignant glioma

PURPOSE: We evaluated the response to Temodal (Schering-Plough Research Institute, Kenilworth, NJ) of patients with newly diagnosed malignant glioma, as well as the predictive value of quantifying tumor DNA mismatch repair activity and O6-alkylguanine-DNA alkyltransferase (AGT). PATIENTS AND METHODS: Thirty-three patients with newly diagnosed glioblastoma multiforme (GBM) and five patients with newly diagnosed anaplastic astrocytoma (AA) were treated with Temodal at a starting dose of 200 mg/m2 daily for 5 consecutive days with repeat dosing every 28 days after the first daily dose. Immunochemistry for the detection of the human DNA mismatch repair proteins MSH2 and MLH1 and the DNA repair protein AGT was performed with monoclonal antibodies and characterized with respect to percent positive staining. RESULTS: Of the 33 patients with GBM, complete responses (CRs) occurred in three patients, partial responses (PRs) occurred in 14 patients, stable disease (SD) was seen in four patients, and 12 patients developed progressive disease (PD). Toxicity included infrequent grades 3 and 4 myelosuppression, constipation, nausea, and headache. Thirty tumors showed greater than 60% cells that stained for MSH2 and MLH1, with three CRs, 12 PRs, three SDs, and 12 PDs. Eight tumors showed 60% or less cells that stained with antibodies to MSH2 and/or MLH1, with 3 PRs, 3 SDs, and 2 PDs. Eleven tumors showed 20% or greater cells that stained with an antibody to AGT, with 1 PR, 2 SDs, and 8 PDs. Twenty-five tumors showed less than 20% cells that stained for AGT, with 3 CRs, 12 PRs, 4 SDs, and 6 PDs. CONCLUSION: These results suggest that Temodal has activity against newly diagnosed GBM and AA and warrants continued evaluation of this agent. Furthermore, pretherapy analysis of tumor DNA mismatch repair and, particularly, AGT protein expression may identify patients in whom tumors are resistant to Temodal.     

Destabilization of tracts of simple repetitive DNA in yeast by mutations affecting DNA mismatch repair

The genomes of all eukaryotes contain tracts of DNA in which a single base or a small number of bases is repeated. Expansions of such tracts have been associated with several human disorders including the fragile X syndrome. In addition, simple repeats are unstable in certain forms of colorectal cancer, suggesting a defect in DNA replication or repair. We show here that mutations in any three yeast genes involved in DNA mismatch repair (PMS1, MLH1 and MSH2) lead to 100- to 700-fold increases in tract instability, whereas mutations that eliminate the proof-reading function of DNA polymerases have little effect. The meiotic stability of the tracts is similar to the mitotic stability. These results suggest that tract instability is associated with DNA polymerases slipping during replication, and that some types of colorectal cancer may reflect mutations in genes involved in DNA mismatch repair.     

Mismatch repair deficiency leads to a unique mode of colorectal tumorigenesis characterized by intratumoral heterogeneity

In order to determine the effects of mismatch repair (MMR) deficiencies in sporadic colorectal carcinomas, 45 such cancers were examined using a sensitive method called crypt isolation technique. Loss of heterozygosity (LOH) in the MSH2 or MLH1 gene was more frequently observed in replication error (RER) (+) carcinomas than in RER (-) carcinomas, which implied that loss of one normal allele could partly affect repair capacity. MSH2 gene defects at both alleles were observed in two carcinomas, which showed severe repair deficiencies. Interestingly, unlike the situation observed in the p53 gene, the MSH2 and MLH1 genes did not show complete LOH. Novel crypt isolation-based subpopulation (CISP) analysis demonstrated that at least two distinct carcinoma subpopulations existed in most carcinomas that showed incomplete LOH; one with and one without LOH. In one carcinoma that had germline mutation and somatic incomplete LOH of the MSH2 gene, the mutator phenotype was only observed in populations affected in both alleles. Thus, the MSH2 gene appears to possess the two hits mechanism of tumor suppressor genes. However, unlike the tumor suppressor genes, MMR gene defects lead to a unique mode of colorectal tumorigenesis characterized by intratumoral heterogeneity.     

Frequency of ApoB and ApoE gene mutations as causes of hypobetalipoproteinemia in the framingham offspring population

Rearranged immunoglobulin variable genes are extensively mutated after stimulation of B lymphocytes by antigen. Mutations are likely generated by an error-prone DNA polymerase, and the mismatch repair pathway may process the mispairs. To examine the role of the MSH2 mismatch repair protein in hypermutation, Msh2-/- mice were immunized with oxazolone, and B cells were analyzed for mutation in their VkappaOx1 light chain genes. The frequency of mutation in the repair-deficient mice was similar to that in Msh2+/+ mice, showing that MSH2-dependent mismatch repair does not cause hypermutation. However, there was a striking bias for mutations to occur at germline G and C nucleotides. The results suggest that the hypermutation pathway frequently mutates G.C pairs, and a MSH2-dependent pathway preferentially corrects mismatches at G and C.     

Human MLH1 deficiency predisposes to hematological malignancy and neurofibromatosis type 1

Heterozygous germ-line mutations in the DNA mismatch repair genes lead to hereditary nonpolyposis colorectal cancer. The disease susceptibility of individuals who constitutionally lack both wild-type alleles is unknown. We have identified three offspring in a hereditary nonpolyposis colorectal cancer family who developed hematological malignancy at a very early age, and at least two of them displayed signs of neurofibromatosis type 1 (NF1). DNA sequence analysis and allele-specific amplification in two siblings revealed a homozygous MLH1 mutation (C676T-->Arg226Stop). Thus, a homozygous germ-line MLH1 mutation and consequent mismatch repair deficiency results in a mutator phenotype characterized by leukemia and/or lymphoma associated with neurofibromatosis type 1.     

Genetic polymorphisms in carcinogen metabolism and their association to hereditary nonpolyposis colon cancer

BACKGROUND & AIMS: The phenotype of hereditary nonpolyposis colorectal cancer shows interfamilial and intrafamilial variation even in the presence of identical predisposing mutations, suggesting the existence of additional phenotype determinants. The modifying role of genetic polymorphisms in loci involved in carcinogen metabolism was studied. METHODS: We focused on colon cancers from kindreds sharing one of two predisposing mutations (mutation 1 or 2) in the mismatch repair gene MLH1 (78 and 14 tumors, respectively). Polymorphisms in N-acetyltransferase 1 (NAT1) and glutathione S-transferase (GST) M1 and GSTT1 were investigated. RESULTS: The NAT1 allele 10 was associated with lower median age at diagnosis in both groups. In mutation 1 group, the NAT1 allele 10 was a risk factor for distal tumor location, both alone (P = 0.028) and combined with the GSTT1-positive genotype (P = 0.008). On the other hand, the combined null genotype of GSTM1 and GSTT1 was associated with proximal tumors. Associations with tumor location were not observed in patients with mutation 2, probably reflecting a small sample size. CONCLUSIONS: The results suggest that genetic polymorphisms in carcinogen metabolism modify the age of onset and tumor location in individuals with inherited deficiency of DNA mismatch repair.     

ATP-dependent assembly of a ternary complex consisting of a DNA mismatch and the yeast MSH2-MSH6 and MLH1-PMS1 protein complexes

MSH2 and MSH6 proteins exist as a stable complex, as do the MLH1 and PMS1 proteins. To study the mismatch binding properties of the MSH2-MSH6 complex and to examine its functional interaction with the MLH1-PMS1 complex, these protein complexes were purified to near homogeneity from overproducing yeast strains. As has been reported previously, the purified MSH2-MSH6 complex binds DNA substrates containing a G/T mismatch and insertion/deletion mismatches, but the binding affinity for the latter decreases as the size of the extrahelical loop increases. Addition of ATP or the nonhydrolyzable ATPgammaS reduces binding of the MSH2-MSH6 complex to the DNA substrates markedly. Here, we show that MSH2-MSH6 forms a ternary complex with MLH1-PMS1 on a mismatch containing DNA substrate. The formation of this ternary complex requires ATP, which can be substituted by ATPgammaS, suggesting that ATP binding alone is sufficient for ternary complex formation. Thus, it appears that ATP binding by the MSH2-MSH6 complex induces a conformation that is conducive for the interaction with MLH1-PMS1 complex, leading to the formation of the ternary complex.     

Defective expression of the DNA mismatch repair protein, MLH1, alters G2-M cell cycle checkpoint arrest following ionizing radiation

A role for the Mut L homologue-1 (MLH1) protein, a necessary component of DNA mismatch repair (MMR), in G2-M cell cycle checkpoint arrest after 6-thioguanine (6-TG) exposure was suggested previously. A potential role for MLH1 in G1 arrest and/or G1-S transition after damage was, however, not discounted. We report that MLH1-deficient human colon carcinoma (HCT116) cells showed decreased survival and a concomitant deficiency in G2-M cell cycle checkpoint arrest after ionizing radiation (IR) compared with genetically matched, MMR-corrected human colon carcinoma (HCT116 3-6) cells. Similar responses were noted between murine MLH1 knockout compared to wild-type primary embryonic fibroblasts. MMR-deficient HCT116 cells or embryonic fibroblasts from MLH1 knockout mice also demonstrated classic DNA damage tolerance responses after 6-TG exposure. Interestingly, an enhanced p53 protein induction response was observed in HCT116 3-6 (MLH1+) compared with HCT116 (MLH1-) cells after IR or 6-TG. Retroviral vector-mediated expression of the E6 protein did not, however, affect the enhanced G2-M cell cycle arrest observed in HCT116 3-6 compared with MLH1-deficient HCT116 cells. A role for MLH1 in G2-M cell cycle checkpoint control, without alteration in G1, after IR was also suggested by similar S-phase progression between irradiated MLH1-deficient and MLH1-proficient human or murine cells. Introduction of a nocodazole-induced G2-M block, which corrected the MLH1-mediated G2-M arrest deficiency in HCT116 cells, clearly demonstrated that HCT116 and HCT116 3-6 cells did not differ in G1 arrest or G1-S cell cycle transition after IR. Thus, our data indicate that MLH1 does not play a major role in G1 cell cycle transition or arrest. We also show that human MLH1 and MSH2 steady-state protein levels did not vary with damage or cell cycle changes caused by IR or 6-TG. MLH1-mediated G2-M cell cycle delay (caused by either MMR proofreading of DNA lesions or by a direct function of the MLH1 protein in cell cycle arrest) may be important for DNA damage detection and repair prior to chromosome segregation to eliminate carcinogenic lesions (possibly brought on by misrepair) in daughter cells.     

Mutation in the mismatch repair gene Msh6 causes cancer susceptibility

Mice carrying a null mutation in the mismatch repair gene Msh6 were generated by gene targeting. Cells that were homozygous for the mutation did not produce any detectable MSH6 protein, and extracts prepared from these cells were defective for repair of single nucleotide mismatches. Repair of 1, 2, and 4 nucleotide insertion/deletion mismatches was unaffected. Mice that were homozygous for the mutation had a reduced life span. The mice developed a spectrum of tumors, the most predominant of which were gastrointestinal tumors and B- as well as T-cell lymphomas. The tumors did not show any microsatellite instability. We conclude that MSH6 mutations, like those in some other members of the family of mismatch repair genes, lead to cancer susceptibility, and germline mutations in this gene may be associated with a cancer predisposition syndrome that does not show microsatellite instability.