ZNRD1, a transcription-associated gene, was recently found in our laboratory significantly suppress the cell proliferation of stomach cancer cells in vitro and in vivo. In this study, we firstly characterized ZNRD1 expression in a wide spectrum of gastric diseases by immunohistochemistry and RT-PCR. We also investigated its antiproliferative effects and associated molecular alterations in human gastric cancer cell line AGS and mouse fibroblast cell line NIH3T3. Anti-ZNRD1 monoclonal antibody H6 was found to react with 38 (63%) of normal gastric tissues, and 51 (81%) of gastritis. In contrast, no positive expression was found in gastric adenocarcinomas. Thus, the expression of ZNRD1 in normal gastric tissues was significantly higher than that in gastric adenocarcinomas. Compared with the control clones, ZNRD1-transfected cells exhibited significant inhibition of cell growth with G1 cell cycle arrest mediated by the suppression of cyclin D1 expression. These results showed that ZNRD1 may play an important role in the regulation of gastric carcinogenesis and could be used as a new target in treatment of stomach cancer.