Mutated ras genes have been found to be conspicuously absent from primary tumors of the esophagus, although high expression of ras p21 oncoprotein in some esophageal squamous cell carcinomas and mutations of the Ki- and Ha-ras genes in esophageal carcinoma cell lines have been reported. In this study, we found amplification of the Ki-ras gene in four of 10 esophageal adenocarcinomas (40%). No such amplification was observed among 61 squamous cell carcinomas, one pseudosarcomatous carcinoma, and eight esophageal cell lines, nor in six adenocarcinomas of the stomach. In two samples on which immunohistochemical analysis could be performed, we found overexpression of Ki-ras proteins when compared with normal samples. This Ki-ras amplification in esophageal tumors did not correlate with any pathological feature of the tumors, with the survival of the patients, or with the presence of other genetic alterations. These findings provide the first evidence for amplification of the Ki-ras gene in human esophageal cancer, which is restricted to adenocarcinomas. We also found that six of eight adenocarcinomas had point mutations in the p53 gene; this is a considerably higher prevalence than that reported for esophageal squamous cell carcinomas. These results strongly suggest that esophageal adenocarcinomas differ from squamous cell carcinomas in their molecular genetic characteristics.