M R Heerma van Voss, F Vesuna, G M Bol, J Afzal, S Tantravedi, Y Bergman, K Kammers, M Lehar, R Malek, M Ballew, N ter Hoeve, D Abou, D Thorek, C Berlinicke, M Yazdankhah, D Sinha, A Le, R Abrahams, P T Tran, P J van Diest, V Raman

DDX3 is a DEAD box RNA helicase with oncogenic properties. RK-33 is developed as a small-molecule inhibitor of DDX3 and showed potent radiosensitizing activity in preclinical tumor models. This study aimed to assess DDX3 as a target in breast cancer and to elucidate how RK-33 exerts its anti-neoplastic effects. High DDX3 expression was present in 35% of breast cancer patient samples and correlated with markers of aggressiveness and shorter survival. With a quantitative proteomics approach, we identified proteins involved in the mitochondrial translation and respiratory electron transport pathways to be significantly downregulated after RK-33 or DDX3 knockdown. DDX3 localized to the mitochondria and DDX3 inhibition with RK-33 reduced mitochondrial translation. As a consequence, oxygen consumption rates and intracellular ATP concentrations decreased and reactive oxygen species (ROS) increased. RK-33 antagonized the increase in oxygen consumption and ATP production observed after exposure to ionizing radiation and reduced DNA repair. Overall, we conclude that DDX3 inhibition with RK-33 causes radiosensitization in breast cancer through inhibition of mitochondrial translation, which results in reduced oxidative phosphorylation capacity and increased ROS levels, culminating in a bioenergetic catastrophe.Oncogene advance online publication, 4 September 2017; doi:10.1038/onc.2017.308.