Abstract 4030: Epigenetic induction of MHC-II pathway expression in murine ovarian cancer cell line
Authors: Taylor B. Turner, Rebecca C. Arend, Mei Li, Troy D. Randall, Andres Forero-Torres, Albert F. LoBuglio, J Michael Straughn, Donald J. Buchsbaum
Journal: Cancer Research (2016)

Abstract INTRODUCTION This study evaluated the potential of epigenetic treatments to induce the expression of the MHC-II antigen presentation pathway in ovarian cancer. Ovarian cancer escapes immune response allowing it to spread in the peritoneal cavity; however, patients with greater immune response to their tumors at baseline have improved survival. Typically, exogenous antigens from tumors are processed and presented via MHC-II to CD4 T cells by antigen-presenting cells. However, if ovarian cancer cells could be induced to express the MHC-II pathway, they could be converted into antigen presenting cells and stimulate an anti-tumor response. METHODS Murine epithelial ovarian cancer cells (ID8) were treated for 72 h with the histone deacetylase inhibitors (HDACi): entinostat (MS-275) (1.25, 2.5, 5, 10 μM) and quisinostat (20, 40, 80, 160 nM). Cells were treated with 5-azacytidine (5-aza), a DNA methytransferase inhibitor, alone and in combination with MS-275 (5 μM) at 17.25, 37.5, 75, and 150 nM. After treatment, the expression of CD74 (an MCH-II pathway protein) and MHC-II was evaluated by flow cytometry. Reverse transcription polymerase chain reaction (RT-PCR) analysis was performed to measure CD74 and CIITA, a transcriptional coactivator that controls MHC-II gene expression. RNA was extracted using RNEasy Mini Kit (Qiagen), cDNA was created with SuperScript VILO Master Mix (ThermoFischer), and a 109bp amplicon was created using JumpStart REDTaq ReadyMix (Sigma-Aldrich). For statistical analysis, a Welch ANOVA was performed using JMP Pro 12. Tukey-Kramer analysis evaluated individual differences. The mean fluorescence intensities (MFI) for the combination treatments were compared using an unequal variance F-test. RESULTS Increased protein expression of CD74 and MHC-II was seen after treatment with MS-275, quisinostat, and 5-aza compared to untreated control. Increased expression of CD74 and MHC-II was seen with the combination treatment of 5-aza (17, 37.5, 75, 150 nM) and MS-275 (5 μM) compared to individual treatments (p &amp;lt; .0001). A dose dependent increased expression of CIITA was demonstrated by RT-PCR with 5-aza treatment alone and in combination with MS-275. Increased mRNA expression of CD74 was also seen for all treatments compared to a negative control. CONCLUSION Treatment with HDAC inhibitors and a DNA methyltransferase inhibitor induces expression of MHC-II in murine epithelial ovarian cancer cells. The conversion of ovarian cancer cells into antigen presenting cells provides a potential therapeutic model to augment the immune response against epithelial ovarian cancer, a disease known to suppress anti-tumor immunity. Citation Format: Taylor B. Turner, Rebecca C. Arend, Mei Li, Troy D. Randall, Andres Forero-Torres, Albert F. LoBuglio, J Michael Straughn, Donald J. Buchsbaum. Epigenetic induction of MHC-II pathway expression in murine ovarian cancer cell line. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4030.