Abstract 27: Development of a rat model of atypical endometrial hyperplasia and a vaginal suppository formulation of SHetA2 for chemoprevention studies
Authors: Chandra Vishal , Rai Rajani , Hussain Sanam , Garcia Lucila , Hatipoglu Manolya , Benbrook Doris 
Published: September 2020

Introduction: The purposes were to develop a rat model of atypical endometrial hyperplasia and a vaginal suppository of the drug SHetA2 (NSC 726189) for use in a chemoprevention study.

Methods: Female Sprague Dawley rats (150-250 g) were ovariectomized to prevent production of endogenous estrogen and progesterone. After a 2 wks rest, placebo or slow release estrogen (E2) pellets were implanted subcutaneously on the lateral side of the neck in the ovariectomized rats randomly assigned to 3 groups (N=25): 1) placebo, 2) 0.5 mg E2/90-day, 3) 1.5 mg E2/90-day. Pellets were replaced after ~10 wks. Rats were weighed wkly. Three rats in each group were sacrificed at 2, 4, 6, 8, 10, 12, 14, and 16 wks after pellet implantation. Organs were collected and weighed. The dimensions of the vaginal canals of six rats were measured to design an aluminum mold to prepare vaginal suppositories containing SHetA2. The mold was custom-made to have a conical shape with a 5.5 mm diameter base (average vaginal canal diameter) and 5 mm height (half of the total length of the vaginal canal - 10.10 mm). A pilot batch of 20 SHetA2 suppositories was prepared using a cocoa butter base, stored at 4°C in an air-tight and light-tight container and evaluated for SHetA2 stability at 1, 2 and 3 months.

Results: Rats with placebo implants exhibited continuous body weight gain. In comparison, both E2-treated groups had reduced weight gain (Repeated Measures ANOVA, p&lt;0.0001). At each time point, average uterine horn weights from the E2-treated groups were significantly higher than the placebo group (Repeated Measures ANOVA, p&lt;0.0001). Placebo group rats had normal endometrium with occasional glandular crowding. At 8-10 wks, focal atypia was observed in 1 of 6 rats in the 0.5 mg E2 group and in 4 of 6 rats in the 1.5 mg E2 group. At the 12-wk time point, atypical hyperplasia was observed in 7 of 9 rats in the 0.5 mg E2 group and in 8 out of 9 rats in the 1.5 g E2 group. Repeated measures ANOVA of organ/body weight ratios indicated that the E2-treated animals had significantly lower spleen weights (p=0.0019) and higher liver (p=0.0001), kidney (p&lt;0.0001) and pancreas (p=0.0004) weights. The reduced organ weights may be due to the significant reduction in body weight in the E2-treated rats. The SHetA2 suppositories were stable at -20°C for 3 months. Storage of individual suppositories in wells of 48-well plates prevented the twining together that occurred when stored in bulk, and allowed removal from packaging without damaging the suppositories.

Conclusions: A 1.5 mg slow release E2 pellet in ovariectomized rats caused development of atypical endometrial hyperplasia within a sufficient timeframe and incidence to allow a chemoprevention study. The mold was suitable to prepare rat-sized suppositories and the resulting SHetA2 suppositories can be safely stored for use within 3 months.

Funding: NCI PREVENT Program Contract HHSN26100008

Citation Format: Vishal Chandra, Rajani Rai, Sanam Hussain, Lucila Garcia, Manolya Hatipoglu, Doris Benbrook. Development of a rat model of atypical endometrial hyperplasia and a vaginal suppository formulation of SHetA2 for chemoprevention studies [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 27.