Purpose
The double strand breaks elicited by sapacitabine, a clinically active nucleoside analog prodrug are repaired by Rad51 and the homologous recombination repair (HR) pathway which could potentially limit its toxicity. We investigated the mechanism by which HDAC inhibitors targeted Rad51 and HR to sensitize AML cells to sapacitabine.
Experimental Design
Chromatin immunoprecipitation identified the role of HDACs in silencing microRNA-182 in AML. Immunoblotting, gene expression, overexpression or inhibition of miR-182 and luciferase assays established that miR-182 directly targeted Rad51. HR reporter assays, apoptotic assays and colony forming assays established that the miR-182, as well as the HDAC inhibition-mediated decreases in Rad51 inhibited HR repair and sensitized cells to sapacitabine.
Results
The gene repressors, HDAC1 and HDAC2, became recruited to the promoter of miR-182 to silence its expression in AML. HDAC inhibition induced miR-182 in AML cell lines and primary AML blasts. miR-182 targeted Rad51 protein both in luciferase assays and in AML cells. Overexpression of miR-182, as well as HDAC inhibition-mediated induction of miR-182 were linked to time- and dose-dependent decreases in the levels of Rad51, an inhibition of HR, increased levels of residual damage and decreased survival after exposure to double strand damage inducing agents.
Conclusions
Our findings define the mechanism by which HDAC inhibition induces miR-182 to target Rad51 and highlights a novel pharmacological strategy that compromises the ability of AML cells to conduct HR, thereby sensitizing AML cells to DNA damaging agents that activate HR as a repair and potential resistance mechanism.