Introduction: The purpose was to evaluate the chemoprevention efficacy of SHetA2 (NSC 726189) in a rat model of atypical endometrial hyperplasia (AEH). Methods: Female Sprague Dawley rats were ovariectomized, rested for 2 weeks, and subcutaneously implanted with placebo or slow-release estrogen (E2; 1.5 mg/90-day) pellets on the lateral side of the neck. The rats were randomly divided for two studies: 1) AEH prevention, in which SHetA2 treatment was started one day after E2 pellet implantation; 2) AEH regression, in which SHetA2 treatment was started after AEH development (12 weeks after E2 pellet implantation) and continued for 4 weeks. SHetA2 was orally given as a suspension at three different doses of 30, 60 or 90 mg/kg/day. The suspension was prepared by first micronizing SHetA2 powder to < 63 µm and then ultrahomogenizing it in aqueous 30% Kolliphor HS15 until the size of drug particles in suspension was < 20 µm. The presence of AEH in H&E stained uterine sections was evaluated by two pathologists as the study endpoint by the presence of abnormal cells (nuclear enlargement, nuclear membrane irregularities, prominent nucleoli, nuclear hyperchromasia, nuclear pleomorphism, mitotic activity and/or loss of axial polarity) within tissue areas that exhibit at least one of the following features: increased gland to stroma ratio of >1:1, glandular complexity including irregular contours, and cribriform/papillary configurations. Results: E2-treated animals had reduced body weight gain compared to placebo-implanted animals (Repeated Measures ANOVA, p2-treated animals compared to placebo (Repeated Measures ANOVA, p2-implanted animals from 86.7% (13/15) in the untreated control to 53.3% (8/15), 26.7% (4/15) and 26.7% (4/15) in the 30, 60 and 90 mg/kg/day SHetA2 treatment groups, respectively (Chai Square Analysis p2-implanted animals observed by lack of AEH in 18.2% (2/11) of the untreated control group and 83.3% (10/12), 81.8% (9/11) and 75% (9/12) in the 30, 60 and 90 mg/kg/day SHetA2 treatment groups, respectively (Chai Square Analysis pConclusions: AEH can be induced by implantation of slow-release E2 pellets in ovariectomized rats. SHetA2 significantly prevented AEH development and caused AEH regression to normal endometrium histology in a dose-dependent manner. Further study is warranted to explore the mechanistic chemoprevention action of SHetA2 and translate these findings toward clinical trials. Funding: NCI PREVENT Program Contract HHSN26100008. Citation Format: Vishal Chandra, Rajani Rai, Stan S. Lightfoot, Manolya Kukut Hatipoglu, Lucila Garcia-Contreras, Doris M. Benbrook. Chemoprevention and regression of estrogen-induced atypical endometrial hyperplasia by oral SHetA2 in a rat model [abstract]. In: Proceedings of the AACR Virtual Special Conference: Endometrial Cancer: New Biology Driving Research and Treatment; 2020 Nov 9-10. Philadelphia (PA): AACR; Clin Cancer Res 2021;27(3_Suppl):Abstract nr PO040.