Search

Because the discovery of a link between mismatch repair deficiency and sporadic or inherited human cancers characterized by microsatellite instability (MSI-H tumors), genes containing coding repeat sequences have been found to be mutated at these repeats in MSI-H tumors from different primary sites as reported in the present review. Accumulation of such alterations appears to be the main molecular mechanism by which MSI-H cells accumulate functional changes with putative oncogenic effects. These mutations occur in many genes at variable frequencies. They can affect genes with a putative role in human carcinogenesis involved in different or similar pathways and are thus thought to be inactivating or activating events selected for in these cancers in a recessive or dominant manner. However, because of the high level of instability characterizing these cancers, they are also likely to occur in genes without any expected role in MSI-H carcinogenesis. In light of these recent data, the concept of target genes for instability and their possible role in MSI-H cancers is reconsidered here.

Mutation frequencies at 25 genes containing coding repeats were compared in colorectal, gastric, and endometrial mismatch repair-deficient (MSI-H) tumors. The overall number of mutations was significantly lower in endometrial than in gastrointestinal MSI-H cancers. Using a likelihood statistical method, target genes were divided in each tumor location into two groups likely to represent gene mutations that do or do not provide selective pressures during tumoral progression. Mutation profiles were quite similar in gastric and colorectal MSI-H cancers but were different in endometrial MSI-H tumors. Deletions in Bat-25 and Bat-26 noncoding repeats were also significantly less important in endometrial as compared with gastrointestinal MSI-H tumors. Our results show that the profile of target gene mutations in MSI-H tumors is tissue specific, with both qualitative and quantitative differences between gastrointestinal and endometrial MSI-H cancers.

Length alterations in microsatellite repeats, termed microsatellite instability (MSI), are found in 10-15% of sporadic colon, endometrial, and gastric cancers harboring defects in DNA mismatch repair (MMR) genes We used the microsatellite markers Big Adenine Tract (BAT) 26 and BAT-25 from the reference panel of five markers recommended by the National Cancer Institute to evaluate the incidence of MSI in 206 central nervous system tumors. We screened 102 pediatric and 104 adult cases representing 165 astrocytic and 41 nonastrocytic tumors. The overall incidence of MSI was 8% (16 of 206). All 16 tumors with MSI were found in pediatric rather than adult patients. MSI was associated with two distinct subtypes of pediatric tumors occurring in 27% (12 of 45) of WHO grade III and grade IV astrocytomas and 24% (4 of 17) of gangliogliomas We evaluated the difference in clinicopathological and genetic features among 45 high-grade pediatric astrocytomas by MSI status. The median survival for pediatric patients with MSI (n = 12) was 8 months compared with 15 months for those patients without MSI (n = 33; P = 0.18). The frequency of p53 gene mutations was 13% for pediatric patients with MSI (n = 8) compared with 47% for those patients without MSI (n = 19; P = 0.19). These results revealed a trend between MSI status and prog nosis and MSI status and frequency of p53 gene mutations. Our data suggest that pediatric high-grade astrocytomas can be attributed to two different genetic pathways: a MMR-deficient pathway and a MMR proficient pathway.

The human T cell transcription factor-4 (hTCF-4) interacts functionally with beta-catenin in the Wnt signaling pathway, which regulates many developmental processes. Moreover, inappropriate reactivation of this pathway attributable to APC or beta-catenin mutations has been described in colorectal cancers. Because only the human TCF-4 cDNA sequence was known, we determined its genomic structure. A total of 17 exons, of which 5 were alternative, were identified. Moreover, four alternative splice sites were observed either experimentally or in silico by a BLAST approach in expressed sequence tag databases. The alternative use of three consecutive exons localized in the 3' part of the hTCF-4 gene changes the reading frames used in the last exon, leading to the synthesis of a number of hTCF-4 isoforms with short, medium, or long-size COOH-terminal ends. We next screened the entire hTCF-4 gene for mutations in a series of 24 colorectal cancer cell lines by denaturing gradient gel electrophoresis and/or direct sequencing. Besides an already described deletion of an A in an (A)9 coding repeat in four cases, we found DNA variants in eight cases for a total of 12 variants, of which 8 were coding. These include one frameshift mutation in the beta-catenin binding domain (exon 1), and one missense mutation in exon 4. In the remaining six cases, nonsense or frameshift mutations were localized in the 3' part of the gene. These latter alterations have as a common consequence to decrease the proportion of the long COOH-terminal hTCF-4 isoform, which contains two binding domains for c-terminal binding protein, a protein implicated in the repression of the TCF family transcriptional activity. Thus, loss of the TCF-4 capacity to interact with COOH-terminal binding protein could be an important event during colorectal carcinogenesis by modifying Wnt signaling.

About 15% of sporadic colorectal cancers show microsatellite instability (MSI) due to the inactivation of mismatch repair genes and are termed MSI-H tumors. In these tumors, frameshift mutations in coding repeats have been found within the TGFbeta-RII, BAX, and IGFRII genes that are probably involved in their progression. In the present work, we report frequent mutations in TCF-4, another target gene for instability. TCF-4 codes for a transcription factor that is a crucial member of the adenomatous polyposis coil (APC)/beta-catenin/T-cell factor (TCF) pathway. Fifty percent (4 of 8) of human MSI-H colorectal cell lines and 39% (19 of 49) of MSI-H colorectal primary tumors were found to have a 1-bp deletion in an (A)9 repeat within the coding region of this gene. In contrast, a frameshift mutation was found in only 1 of 56 non-MSI colorectal tumors and in none of 16 non-MSI colorectal cancer cell lines. These results suggest that TCF-4 frameshift mutations are selected for and play a role in colorectal MSI-H tumorigenesis. Depending on different reading frames due to alternatively spliced TCF-4 mRNA, the (A)9 repeat normally codes for several isoforms that could serve as modulators of TCF-4 transcriptional activity. The deletion of one nucleotide in this repeat could change TCF-4 transactivating properties by modifying the respective proportions of the different isoforms.

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