Combined inhibition of EZH2 and histone deacetylases as a potential epigenetic therapy for human uterine sarcoma cells.

Abstract

e17620 Background: Uterine sarcoma is strongly associated with poor prognosis, high rates of recurrence, and metastasis. However, the molecular mechanisms underlying the origin and development of uterine sarcoma and treatment options are limited. Enhancer of zeste homolog 2 (EZH2) and histone deacetylase (HDACs) play an important role in cancer progression. This study aimed to assess the effect of targeting both EZH2 and HDACs on the phenotype of uterine sarcoma (US) cells. Methods: Tazemetostat (EPZ-6438, E7438), a potent, and selective EZH2 inhibitor, was first approved by the US Food and Drug Administration (FDA) in 2020. Entinostat (MS-275, SNDX-275) is one of the strong inhibitors for HDAC1 and HDAC3, Vorinostat is used to treat cutaneous T-cell lymphoma, Tucidinostat (Chidamide, HBI-8000, CS-055) is a low nanomolar inhibitor of HDAC1, 2, 3, and 10. Uterine sarcoma cell line (MES-SA) was treated with different concentrations (1.56-200 mM) of Tazemetostat and/or Entinostat, Vorinostat, Tucidinostat (Chidamide). for 24, 48, 72 h, respectively. Further mechanistic confirmation was carried out on best combination in both 2D and 3D culturing using IHC and real time PCR. Results: Our data showed that Entinostat synergistically (with a combination index 0.539) augments the cytotoxic activity of Tazemetostat in both 2D and spheroids in vitro model systems. The IC50 values for Tazemetostat were significantly reduced to 4.5 mM after combination treatment with 6.25 mM Entinostat for 48 h. Entinostat enhanced the antiproliferative effect of Tazemetostat with a decrease in the expression of proliferative markers Ki67(P < 0.0001) and PCNA (P < 0.01). Furthermore, combination treatment significantly increased US cell apoptosis, concomitantly with increased both BAX and Caspase-3 levels and decreased Bcl-2 (P < 0.0001) using 2D and spheroid in vitro model (P < 0.001). The finding was confirmed by IHC analysis in the US cell-derived spheroids. Conclusions: Our studies demonstrate that Entinostat, in combination with Tazemetostat proves significantly superior to either single treatment. Dual targeting EZH2 and HDACs may provide a promising treatment option for this aggressive cancer disease.