The purpose of this study was to investigate the role of NAC1 in the development of endometrial cancer. NAC1 expression and localization were assessed with immunohistochemistry in the normal cyclic human endometrium, hyperplastic endometrium, and endometrial cancer. Expression of NAC1 in the glandular cells was significantly higher in the early and mid proliferative phases than in the other menstrual phases, endometrial hyperplasia, and endometrial carcinoma. NAC1 expression was down-regulated during endometrial carcinogenesis. There were significant correlations between positive NAC1 expression and pathological grade (P=0.037). No significant associations were found between NAC1 expression and the other clinicopathological characteristics including patient age, FIGO staging, depth of myometrial invasion, pelvic lymph node metastasis, lymphovascular space invasion, menopause, or body mass index. NAC1 gene knockdown inhibited cell growth and induced apoptosis in Ishikawa, HHUA, and JHEM2 cell lines, all of which overexpressed NAC1. Ectopic overexpression of the NAC1 gene stimulated cell proliferation in the HEC1B, and JHEM1 endometrial cancer cell lines, which have lower endogenous NAC1 expression. Endometrial carcinomas with NAC1 overexpression are clinically aggressive, high-grade carcinomas. Therefore, detection of NAC1 overexpression in endometrial cancers may identify patients who will benefit from NAC1 targeted therapy.