We have investigated frameshift mutations in exonic repeats in the ATR, BRCA1, BRCA2, PTCH, CTCF, Cx26, NuMa and TGFbetaRII genes, using human tumor samples from stomach, esophagus, breast and skin and melanoma, as well as colon cancer and endometrial cancer cell lines (125 samples in total). We developed a sensitive method to detect mutations in the repeats, using the introduction of an artificial restriction site into a repeat. The method detects a single mutant among 10(3) normal genes. Thus, an alteration in a repeated sequence can be detected unambiguously. The (A)(8) repeat of BRCA2 was found mutated in only two of five colon cell lines with microsatellite instability (MI(+)). The ATR gene has an (A)(10) repeat which was altered in two of three MI(+) stomach cancer samples and one of three MI(+) endometrial cell lines. The TGFbetaRII gene [with an (A)(10) repeat] had the maximal frequency of mutations: 10 out of 13 MI(+) samples. At least one sample from all types of cancers, except melanomas, was positive for TGFbetaRII gene mutations. No mutations were found in repeats in the BRCA1, PTCH, CTCF, NuMA and Cx26 genes in any types of tumors examined. In conclusion, our study indicates that repeats were altered only in MI(+) cells and that the mutation frequencies in the genes studied differ among tumor types. Based on these results, we discuss meaningful and meaningless alterations in exonic repeats.