Mutations in two short noncoding mononucleotide repeats in most microsatellite-unstable colorectal cancers.

DNA mismatch repair (MMR)-deficient cells typically accumulate mutations in short repetitive DNA tracts. This microsatellite instability (MSI) facilitates malignant transformation when affecting genes with growth-related and caretaker functions. To date, several putative MSI target genes have been proposed mainly based on high mutation frequency within their coding regions. However, some intronic repeat mutations have also been suggested to associate with MSI tumorigenesis, indicating the need for additional analyses on noncoding repeats. Here we have analyzed an intronic T9 repeat of semenogelin I (SEMG1) and report mutation frequencies of 51% (75 of 146) and 62% (8 of 13) in MMR-deficient primary colorectal cancers and cell lines, respectively. The putative effect of the SEMG1 mutations was assessed by RNA and protein level analyses, but no differences were detected between colorectal cancer cell lines with different SEMG1 status. Subsequently, the general background mutation frequency of MSI colorectal cancers was assessed by screening for intergenic T9 repeat alterations. One of 10 examined repeats was mutated in 70% (102 of 145) of the colorectal cancers evaluated. The frequencies observed here are notably higher than previously published in noncoding repeats shorter than 10 bp in MMR-deficient primary tumors. Our results indicate that high mutation frequencies, similar or higher than those observed in proposed and approved target genes, can be detected in repeat tracts of MSI tumors without any apparent selection pressure. These data call for urgent and thorough large-scale evaluation of mutation frequencies in neutral short repetitive sequences in MMR-deficient tumors.

A novel functionally deficient MYH variant in individuals with colorectal adenomatous polyposis.

Inherited biallelic mutations in the base excision repair gene MYH confer susceptibility to colorectal adenomas and carcinoma. Approximately 85% of Caucasians with MYH mutations carry the (c.494A>G) p.Y165C and (c.1145G>A) p.G382D variants. Only a few other clearly pathogenic mutations have been identified, and mutation analyses tend to focus on the two founder mutations rather than the whole coding region of the gene. We sequenced the entire coding region of MYH in a population-based series of 24 Finnish APC-mutation negative polyposis patients in order to identify novel pathogenic MYH variants. A population-based series of 1,042 Finnish colorectal cancer patients and 85 cancer-free controls were available for further evaluation. A functional cleavage assay was designed to evaluate consequences of possible novel variants to protein function. Three novel MYH variants, (c.270C>T) p.Y90Y, (c.1376C>A) p.A459D, and (c.1389G>C) p.T469T, were observed. p.A459D variant in exon 14 was identified in two patients from the polyposis series, once in homozygosity and once in compound heterozygosity with p.Y165C. In the population-based series of 1,042 colorectal cancer patients, the p.A459D mutation was identified once, in homozygosity (allele frequency 0.1%). No p.A459D mutations were identified in the control individuals. In vitro cleavage assay showed significantly reduced repair activity in p.A459D cells. Interestingly, another variant in the same codon has previously been described in a British study, supporting a key role for the codon 459 in MYH function. We therefore suggest that screening of mutations in MYH exon 14 should be added to the molecular analysis at-risk individuals.

Proportion and phenotype of MYH-associated colorectal neoplasia in a population-based series of Finnish colorectal cancer patients.

Recessively inherited mutations in the base excision repair gene MYH have recently been associated with predisposition to colorectal adenomas and cancer in materials selected for occurrence of multiple adenomas. In particular, variants Y165C and G382D have been shown to play a role in Caucasian patients. To evaluate the contribution of MYH mutations to colorectal cancer burden on the population level, and to examine the MYH-associated phenotype in an unselected series of colorectal cancer patients, we determined the frequencies of Y165C and G382D MYH mutations in a population-based series of 1042 Finnish colorectal cancer patients. Four (0.4%) patients had both MYH alleles mutated. Although all these patients had multiple adenomatous polyps, the phenotypes tended to be less extreme than in previous studies on selected cases. The lowest number of colorectal adenomas at the time of cancer diagnosis was five. Cases with one mutant MYH allele were subjected to sequencing of all exons to detect possible Finnish founder mutations, but no additional changes were detected. The Y165C and G382D variants were not present in 424 Finnish cancer-free controls showing that MYH mutations are not enriched in the population. As evaluated against national Finnish Polyposis Registry data MYH-associated colorectal cancer appears to be as common as colorectal cancer associated with familial adenomatous polyposis.