AbstractMounting evidence indicates that deregulation of microRNAs (miRNAs) are involved in development of many human diseases, including cancers. Regulation of miRNA is a complicated process and some components in the regulation are known to be altered in human cancers. Among the miRNA regulation‐related genes, we found that AGO1, AGO2, TNRC6A, TNRC6C, TARBP2 and EXPORTIN5 genes have mononucleotide repeats in their coding sequences. To see whether these genes are mutated in cancers with microsatellite instability (MSI), we analysed the mononucleotide repeats in 27 gastric cancers (GCs) with high MSI (MSI‐H), 18 GC with low MSI (MSI‐L), 45 GC with stable MSI (MSS), 41 colorectal cancers (CRCs) with MSI‐H, 14 CRCs with MSI‐L and 45 CRCs with stable MSI (MSS) by single‐strand conformation polymorphism (SSCP) analysis and DNA sequencing. We found AGO2, TNRC6A, TARBP2, TNRC6C and EXPORTIN5 mutations in 10, six, one, one and one cancer(s), respectively. They were detected in MSI‐H but not in MSI‐L or MSS cancers. The GCs and CRCs with MSI‐H harboured one or more mutations of the genes in 22% and 27%, respectively. We also analysed Ago2 and TNRC6A protein expressions in GCs and CRCs with MSI‐H. In cancers with MSI‐H, loss of Ago2 expression was observed in 40% of GCs and 35% of CRCs, while loss of TNRC6A was observed in 52% of the GCs and 54% of the CRCs. Our data indicate that frameshift mutations in AGO2 and TNRC6A and their losses of expression are common in GCs and CRCs with MSI‐H, and suggest that these alterations may contribute to the cancer development by deregulating miRNA regulation. Copyright © 2010 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.