Frequent loss of heterozygosity (LOH) on the long arm of chromosome 5 (5q) has been reported in many types of human malignancies, including gastric carcinoma. One of the targets of 5q-LOH in colorectal carcinoma is certainly the adenomatous polyposis coli (APC) gene on 5q21. However, other evidence has suggested the presence of another tumor suppressor gene in this region which may be inactivated in gastric carcinoma. In the present study, to determine the location of the putative tumor suppressor gene on 5q, LOH at nine microsatellite loci on 5q were investigated at 38 differentiated adenocarcinomas of the stomach that probably did not carry APC mutations. LOH at any locus on 5q occurred in 37% (14 of 38) of the tumors. Although many tumors exhibited large interstitial deletions on 5q that included the APC locus (5q21), we have identified minimum regions of deletion as the D5S428 locus and the interferon regulatory factor-1 (IRF-1) locus. Thus, at least two putative tumor suppressor genes, which play a crucial role in the genesis of differentiated adenocarcinoma of the stomach and are distinct from the APC gene, lie on 5q.