Abstract 5855: Radiosensitization of cervical cancer cells by HDAC inhibitors

Abstract

Abstract Introduction: While histone deacetylase (HDAC) inhibitors have demonstrated in vitro and in vivo radiosensitization in several human cancer cell lines, the efficacy and mechanism of action of HDAC inhibitors in HPV-positive cancers are poorly understood. We hypothesize that HDAC inhibitors can be used to treat HPV-positive squamous cell carcinomas by enhancing radiation sensitivity. Methods: The cellular cytotoxicity of ten HDAC inhibitors was examined in HPV-positive (CaSki, SiHa, HeLa) and HPV-negative (C33a) cervical cancer cell lines by the WST-8 [2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium, monosodium salt] assay, and the drug with the highest cytotoxicity (pan-HDAC inhibitor SP-1-161) was selected for further analysis. SAHA was used as a control. Evaluation of the cell cycle phase distribution was performed by flow cytometry using FACS analysis following treatment with drugs at 24, 48, and 72 hours as well as in combination with radiation in CaSki cells (HPV-16 positive). The extent of radiation sensitization was confirmed by cellular clonogenic assays in CaSki cells. Results: Of all drugs tested, cytotoxicity was demonstrated in a dose-dependent manner. The values of IC50s were at 1.071, 1.567, 1.71, and 0.929 μM (SAHA) and 0.054, 0.3554, 0.0994, and 0.095 μM (SP-1-161) for CaSki, SiHa, HeLa, and C33a cell lines, respectively. SP-1-161 conferred significant cytotoxicity (0.054 μM) in CaSki cells. Cellular radiation sensitivity is tightly associated with cell cycle phases and varies as a function of the position in the cell cycle. To assess the effects of HDAC inhibitors on the cell cycle, CaSki cells were treated with IC50 dosage (SP-1-161 at 0.054 μM, SAHA at 1 μM) for 24 hours, 48 hours, and 72 hours and then treated with radiation. The drugs alone arrested cells primarily in the G1 phase of cell cycle (77.3% for SP-1-161, 84% for SAHA), a relatively radiation sensitive phase. Pre-treatment of SP-1-161 for 24 hours prior to radiation (5 Gy) increased cell cycle arrest in the G1 phase as well. To determine the radiosensitization property of SP-1-161, the radiation clonogenic survival assay was performed following pre-treatment of SP-1-161 for 24 hours. Cells were exposed to graded doses of γ-radiation, and the data were fitted using the single-hit multitarget and the linear-quadratic models. Comparisons of survival curves revealed smaller D0 (D0 ratio 1.43) for cells irradiated in the presence of SP-1-161 (D0 = 1.4) than for controls (D0 = 2.0). Interestingly, treatment with SAHA did not impact radiosensitization (D0 = 1.9, D0 ratio 1.05). Conclusion: SP-1-161 conferred significant toxicity in CaSki cells and primarily arrested cells in the G1 phase of cell cycle. Furthermore, treatment with SP-1-161 enhanced radiosensitization, while SAHA showed little change despite cell cycle arrest primarily in the G1 phase. Together, our results support SP-1-161 as a new therapeutic agent with potential for cervical cancer. Citation Format: Ima Paydar, Alfredo Velena, Scott Grindrod, Mira Jung. Radiosensitization of cervical cancer cells by HDAC inhibitors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5855. doi:10.1158/1538-7445.AM2017-5855