Abstract 3801: Functional relevance of A-to-I RNA editing on the immune response in breast cancer

Abstract

Abstract INTRODUCTION: RNA editing is a post-transcriptional modification that changes double stranded RNA sequences (dsRNA). The most common way of editing in humans is the Adenosine-to-Inosine type, which is catalyzed by members of the ADAR enzymes family. A-to-I RNA editing has been shown to be essential for normal tissue development but also plays a crucial role in many cancers. We have notably shown that in breast cancer the editing frequency is increased in tumors as compared to normal tissue and correlates with ADAR expression. We also showed that type I IFN response and ADAR DNA copy number together explain 53% of ADAR expression in breast cancers. Moreover, the implication of ADAR-mediated RNA editing in preventing the sensing of endogenous dsRNA leading to inhibition of the immune response has been recently reported. ADAR-mediated RNA editing may play a crucial role in tumor progression by modulating immune response through the dsRNA sensing pathway. Here, we aim to assess the influence of ADAR-mediated RNA editing in the sensing of endogenous dsRNA and the immune response in breast cancer. MATERIAL & METHODS: ADAR expression was modulated in MDAMB231 breast cancer cell line using shRNA lentiviral transduction and lentiviral plasmid transduction for ADAR inhibition and ADAR wild-type and editing-deficient mutant overexpression respectively. ADAR expression was assessed by RTqPCR/Western blotting. dsRNA level was evaluated by immunofluorescence using the dsRNA-specific J2 antibody (Scicons). RESULTS: Poly I:C transfection, mimicking dsRNAs, as well as IFN Type I exposure induced the immune response in BC cell lines, in particular it increases the expression of RIG-I, MDA5 and IRF7 involved in the dsRNA sensing pathway. DNMTi treatment, known as an inducer of dsRNA level, also activated cytosolic dsRNA response in these cells in a dose dependent manner. In order to assess the role of ADAR-mediated RNA editing in the activation of the dsRNA sensing pathway, we exposed MDAMB231 cell lines in which ADAR expression was modulated to DNMTi. A wide range of doses of DNMTi, from 0.3 to 50μM, and time of exposure, from 3h to 72h, were tested. The cytosolic dsRNA pathway was induced in a dose dependent manner after DNMTi exposure. However, no effect of ADAR modulation on the activation of the pathway was observed. No difference in dsRNA levels was observed after activation of the immune pathway by DNMTi or IFN exposure, nor after ADAR modulation. CONCLUSION & PERSPECTIVES: dsRNA pathway activation by DNMTi, poly I:C transfection or IFN Type I treatment is not dependent of ADAR in MDAMB231 cell lines. Moreover, in contrast to data in colorectal cell lines, our data suggest that DNMTi exposure does not increase cytosolic dsRNA level. Citation Format: Floriane Dupont, Florian Clatot, David Venet, Véronique Kruys, Debora Fumagalli, Vincent Detours, Françoise Rothé, Christos Sotiriou. Functional relevance of A-to-I RNA editing on the immune response in breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3801.