NTX-301, a Novel Hypomethylating Agent for Ovarian Cancer

Aim 1. To test the effect of NTX-301 on resensitizing chemo-resistant ovarian cancer cells to chemotherapy in vitro and in vivo. DNMT inhibitors have been demonstrated to resensitize platinum resistant ovarian cancer (OC) cells to chemotherapy (1). We hypothesize that NTX-301, a novel and potent DNMT1 inhibitor, can be used in a similar fashion to resensitize chemo-resistant ovarian cancer cells to platinum treatment. To test this hypothesis, we propose the following experiments: 1. Effects on cell viability. We will determine the half maximum inhibitory concentration (IC50) of NTX-301 on cell viability of platinum sensitive and resistant variants of the ovarian cancer cell lines OVCAR5, OVCAR3, SKOV3, and COV362, and cells derived from platinum naïve and platinum resistant OC tumors. The resistant OC cells have been developed by treating parental cells with cisplatin or carboplatin at the IC50, followed by recovery of the cells. The procedure was repeated three times. The resistant cells have an IC50 to platinum 2-4X higher compared to parental cells (Wang, Cancer Research, 2020, in press). Cells will be treated with serial doses of NTX-301, and cell viability will be assessed using a Cell Counting Kit-8 (CCK-8). Decitabine will be used as a control for these experiments. 2. DNMT expression and activity. To determine the lowest dose and duration of NTX-301 treatment necessary to inhibit DNMT1, DNMT expression and activity, we will perform time-course experiments using isogenic platinum resistant and sensitive OC cells described above treated with doses of NTX-301 that do not decrease cell viability (from the IC50 Exp.). Levels of DNMT1, 3A and 3B will be measured by western blot and DNMT activity will be assessed with a kit. The lowest dosage of NTX-301, which is associated with maximum inhibition of DNMT expression and activity will be used in the follow-up study to resensitize OC cells to chemotherapy. Decitabine will be used as a control for these