Abstract 3845: Anti-tumor potential of Hydralazine, a DNMT1 inhibitor, on breast cancer growth and progression

Abstract

Breast cancer (BC) is the second largest diagnosed cancer among women, with approximately 266,120 new cases of BC in the United States. Thus far BC is understood as a disease driven by genetic abnormalities. Specifically, recent studies provide evidence that epigenetic modifications, such as methylation profiles, play a critical role in BC development regarding tumor suppressor genes. Despite there being many DNMT inhibitors, such as 5- azacytidine (Vidaza), they are ineffective versus solid tumors and they elicit a cytotoxic response. Thus, it is vital that a drug that is able to halt BC growth and progression is investigated. This project hopes to test the anti-tumor potential and therapeutic potential of Hydralazine (HDZ) because of its ability to act as a DNA methylation inhibitor, specifically DNMT1 and DNMT3A. In addition, despite HDZ being currently tested in clinical cancer trials for its antihypertensive effect, its mechanism of antitumor action remains undefined.We analyzed the anti-cancer potential of HDZ on two human BC cell lines and a mouse mammary tumor cell line (AT3). Cells were treated with different doses of HDZ (0, 1, 10, 25, and 50 M) for different time points (1, 2, 3, 7, and 14 days) and cell viability was measured by MTT assay. From these experiments, we optimized the time and concentration of HDZ in BC cells. Finally, we treated the cells with the appropriate concentration and time and then prepared RNA and protein for gene expression analysis, measured DNMT1 activity, and analyzed stem cells status using cell surface markers (CD24/CD44). We also tested the anti-tumor potential of HDZ in a mouse model using AT3 cells. AT3 cells were implanted in the mammary fat pads and grown to the tumor size of 250 mm3. Then, we treated the mice with HDZ and monitored the tumor growth. Tumor tissues harvested from these mice will be used for gene expression analysis, measure DNMT1 activity, histological analysis and analyze the stem cell status such as Oct4, Nanog, Gata4, Slug, and Sox2.All 4 BC cell lines demonstrated a slightly lower cell viability from the MTT when treated for 3 days and used at the highest dosage. In addition, through the use of RT-PCR, it has been shown that the levels of DNMT1 and CD24 expressions have been slightly downregulated. Western Blot analysis showed, the protein levels of DNMT1 and CD24 also decreased slightly. We also found that HDZ treatment significantly reduced tumor growth and volume in the mouse model. These findings suggest that the drug HDZ has a slight anti-tumor property on breast cancer growth and progression in the cell and mouse models. Although not as significant as we originally thought, the future implications of this project would be to complement HDZ with another drug such as a stem cell inhibitor. We believe that this dual therapy would synergistically work to halt BC progression and growth much more efficiently than any monotherapy for the treatment of triple-negative breast cancer. Citation Format: Parth B. Patel. Anti-tumor potential of Hydralazine, a DNMT1 inhibitor, on breast cancer growth and progression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3845.