Abstract 5218: Inhibition of γ-secretase decreases Notch signaling and cell migration in uterine carcinosarcoma

Abstract

Abstract Carcinosarcoma (CS) is a rare and aggressive type II endometrial adenocarcinoma (EmCa) accounting for less than 5% of all uterine malignancies, but accounting for a disproportionately high disease-specific mortality rate. Notch signaling is an evolutionarily conserved pathway with oncogenic roles in several cancers. Notch proteins are expressed in Type I and Type II EmCa, cell lines, and human tissue, but Notch function has not been characterized in CS. To determine whether inhibiting Notch signaling may be a therapeutic target in CS we assessed expression of Notch family genes and the impact of γ-secretase inhibition on cellular migration in the CS cell line, CS-99, and compared to lower risk Type I EmCa cell line (HEC-1-A). Expression of Notch receptors, ligands and effectors was determined using RT-PCR. We determined the impact of exposure to DAPT, a γ-secretase inhibitor, vs DMSO (vehicle control) on cell viability with a MTT assay. The ability of DAPT to inhibit Notch signaling was assessed by quantitative (q) RT-PCR for the Notch effector, HES1. The relative expression level of each target gene was normalized to 18s rRNA. Cellular migration after exposure to DAPT was assessed using a scratch assay for CS-99 and HEC-1-A. The extent of cell migration was measured using ImageJ (NIH Bethesda, MD). CS-99 and HEC-1-A, expressed receptors NOTCH1-4, ligands DLL4, JAG1, JAG2, and effectors HES1, HEY1, and NRARP. The MTT assay revealed >88% cellular viability after 24 hr exposure to 10µM in CS-99 and 50µM DAPT in HEC-1-A. Exposure to DAPT significantly decreased expression HES1 in CS-99 and HEC-1-A. Migration was decreased in CS-99 and unchanged in HEC-1-A with 10µM and 50 µM of DAPT, respectively. Inhibition of Notch signaling activity decreased cellular migration in CS-99 cell line suggesting that Notch signaling may impact disease growth in CS. Unchanged HEC-1-A cellular migration suggests that Notch signaling might not impact disease growth in Type I EmCa. Inhibition of Notch signaling may be a promising therapeutic target in CS. Inhibition of Notch signaling in CS-99 and HEC-1-A with a γ-secretase inhibitor (median + IQR)*HEC-1-A + DMSOHEC-1-A + DAPT 50μMP-valueCS-99 + DMSOCS-99 + DAPT 10μMP-valueHES1 (normalized fold change)0.94 (0.83-1.3)0.35 (0.25-0.41)0.0220.99 (0.84-1.2)0.19 (0.16-0.23)0.002% wound closure (0-6h)12.5 (8.5-15.5)24.1 (10.5-33.0)0.1311.4 (9.7-11.5)11.3 (9.1-11.4)0.7% wound closure (0-24h)38.1 (31.4-46.1)63.0 (33.6-100.0)0.2473.3 (66.6-73.9)61.7 (52.6-64.7)0.05 Citation Format: Sofia D. Gabrilovich, Yasmin Abedin, Sreevidya Santha, Tracy Wu, Mark Einstein, Nataki C. Douglas, Jenna Z. Marcus. Inhibition of γ-secretase decreases Notch signaling and cell migration in uterine carcinosarcoma [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 5218.