Mutations in TGFbeta-RII and BAX mediate tumor progression in the later stages of colorectal cancer with microsatellite instability

Abstract

Background: Microsatellite instability (MSI) occurs in 15% of colorectal cancers (CRC). The genetic targets for mutation in the MSI phenotype include somatic mutations in the transforming growth factor beta receptor typeII (TGFbetaRII), BAX, hMSH3 and hMSH6. It is not clear how mutations of these genes mediate tumor progression in the MSI pathway, and the temporal sequence of these mutations remains uncertain. In this study, early stage CRCs were examined for frameshift mutations in these target genes, and compared with late stage tumors and CRC cell lines.

Methods: We investigated 6 CRC cell lines and 71 sporadic CRCs, including 61 early stage cancers and 10 late stage cancers. Mutations of repetitive mononucleotide tracts in the coding regions of TGFbetaRII, BAX, hMSH3, hMSH6, IGFIIR and Fas antigen were identified by direct sequencing.

Results: Thirteen (18.3%) of 71 CRC, including 9/61 (14.7%) early stage cancers and 4/10 (40%) late stage cancers, were identified as MSI and analyzed for frameshift mutations. No mutation in the target genes was observed in any of the 9 early stage MSI CRCs. In contrast, frameshift mutations of TGFbetaRII, BAX, hMSH3 and hMSH6 were present in 3/4 late stage MSI tumors. There is a statistical association (p = 0.014) between mutation in any one gene and tumor stage.

Conclusions: TGFbetaRII, BAX, hMSH3 and hMSH6 mutations are relatively late events in the genesis of MSI CRCs. The frameshift mutations in these target genes might mediate progression from early to late stage cancer, rather than mediating the adenoma to carcinoma transition.

Figure 1

Frameshift mutations in CRC cell lines. Lane a: products from normal control DNA. The products of MSI cell lines are shown in Lanes b (HCT116), c (LoVo), e (LS174-T) and f (DLD1). Products of non-MSI cell lines are shown in Lanes d (SW480) and g (HT29). The genes are indicated at the left. Arrowheads indicate frameshift mutations in cell lines.

Figure 2

Frameshift mutations in CRCs with MSI. The genes are indicated at the left. Arrowheads indicate 1 bp deletion mutations in tumors. T and N refer to DNA from tumors and corresponding normal tissues, respectively.

Figure 3

Sequence analysis of frameshift mutations in late stage CRCs. Single one bp deletions in the mononucleotide repeats were found in each case.