Abstract PR001: Preclinical development of SHetA2 for endometrial cancer therapy

Abstract

Abstract Introduction: SHetA2 (NSC 726189) is a novel drug that is entering first in human trials for gynecologic cancers. Upon binding to three related heat shock protein A chaperones proteins (HSPA5/Grp78, HSPA8/Hsc70, HSPA9/mortalin), SHetA2 causes release of their client proteins. Our objective was to evaluate the potential of SHetA2 for endometrial cancer therapy and to explore its mechanism of action. Methods: Endometrial cancer cell lines, Ishikawa, MFE280, MFE296, Hec1a, Hec1B and AN3CA were evaluated for SHetA2 effects on cell growth with a MTT assay. Ishikawa, Hec1B and AN3CA were also evaluated for SHetA2 effects on cell cycle progression by flow cytometry analysis of PI staining; apoptosis by flow cytometry analysis of annexin/PI staining, anchorage-independent growth using an agar colony formation, invasive potential with an matrigel invasion assay, and protein levels (in Ishikawa only) by mass spec analysis of protein extracts using a 5% false discovery rate. The in vivo efficacy of SHetA2 and paclitaxel alone and in combination were evaluated in athymic nude mice bearing Ishikawa xenografts treated with oral 60 mg/kg/day SHetA2, intraperitoneal injection of 10 mg/kg/week of paclitaxel, the combination or vehicle controls. Results: SHetA2 significantly reduced growth of all endometrial cancer cell lines in time- and dose- dependent manners. This inhibition occurred via G1 cell cycle arrest in Ishikawa and Hec-1b, and G2 arrest in AN3CA. SHetA2 inhibited anchorage-independent growth and invasive behaviors, and induced apoptosis in Ishikawa, Hec-1b and AN3CA. Proteomic analysis identified levels of 48 proteins to be significantly altered by SHetA2 in Ishikawa cells. Of these, 15, 10 and three are known to be bound by Hsc70, Grp78 and mortalin, respectively. Ingenuity analysis identified the top SHetA2-regulated canonical pathways to be EIF2 Signaling, Unfolded Protein Response, Glycolysis, ER Stress Pathway and Gluconeogenesis, and predicted the top upstream regulators of the proteomic changes to be three microtubule associated proteins (MAPT, PSEN1, APP), which also bind Hsc70. Since Hsc70 also binds paclitaxel, we predicted positive interaction between SHetA2 and paclitaxel on endometrial tumor growth. Both SHetA2 and paclitaxel significantly inhibited growth of Ishikawa xenografts, and the combination was more effective than either drug alone (Repeated Measures ANOVA p=0.0025). There were no significant effects of the treatments on mouse body weights. Conclusions: SHetA2 inhibits growth, migration and invasion of endometrial cancer cell lines in association with cell cycle arrest, apoptosis and regulation of its Grp78/Hsc70/mortalin target protein networks. Pathways regulated by SHetA2 are consistent with disruption of chaperone proteins. The in vivo efficacy of SHetA2 alone and in combination with paclitaxel justifies further development of this drug and combination with paclitaxel for treatment of endometrial cancer. Funding: Stephenson Cancer Center. Citation Format: Vishal Chandra, Rajani Rai, Doris M. Benbrook. Preclinical development of SHetA2 for endometrial cancer therapy [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 PR001.