Aberrantly methylated DNA fragments in a human gastric cancer were searched for by a genome-scanning method, methylation-sensitive-representational difference analysis (MS-RDA). Six DNA fragments flanked by CpG islands (CGIs) and hypermethylated in the cancer were isolated. Four of the 6 fragments possessed genes in their vicinities. Quantitative RT-PCR analysis of the 4 genes showed reduced expression of 2 genes in cancers: Insulin-induced protein 1 (INSIG1/CL-6) and p41 Arp2/3 complex (p41-Arc). As for INSIG1, a DNA fragment was derived from the edge of a CGI in the promoter region. The edge was methylated in 11 of 22 primary gastric cancers, whereas the center was not methylated in any cancer. INSIG1 expression was markedly reduced in 19 cancers, including the 11 cancers with the methylation. By 5-aza-2'-deoxycytidine treatment of 5 cell lines with the methylation of the edge, partial restoration of INSIG1 expression was detected only in 2 of them. These data indicated that, although the reduced INSIG1 expression in cancers was associated with the methylation at the edge of the CGI in the promoter region, the methylation was likely to be a secondary change. As for p41-Arc, a DNA fragment was derived from a CGI overlapping exon 8, and its methylation did not correlate with its expression. However, methylation of a CGI in the promoter region with a marked reduction of its expression was observed in 1 of the 22 primary cancers. INSIG1 and p41-Arc are known to be involved in cellular differentiation and morphology, respectively, and it was suggested that their reduced expressions might be involved in gastric cancer development or progression.
Copyright 2002 Wiley-Liss, Inc.