The DNA Damage Response as a target for Senolytic Therapies

Abstract

Cellular senescence is a phenomenon by which normal cells cease to divide but stay transcriptionally and metabolically active while their cell cycle is durably arrested. Although senescence functions as tumour suppressor mechanism during early tumourigenesis, accumulating evidence suggests that it poses a risk for tumour progression in pre-existing tumour cells. Ongoing research is done to identify agents that eliminate unwanted senescent cells selectively, so called senolytics. One of the features described for several types of senescence is an activated DNA damage response (DDR). In this work, we used murine NrasG12V p19-/- HCC cells to analyse DDR activation in a novel type of senescence induced by the RNA polymerase I inhibitor CX-5461 that has been recently characterised by the Zender laboratory (K. Wolter) and termed ribosomal checkpoint induced senescence (RCIS). Although we observed pronounced DDR activation and permanent G2 cell cycle arrest upon CX-5461 treatment, we neither found a synergistic effect when treating cells with CX-5461 and DDR inhibitors simultaneously nor a senolytic effect by DDR inhibition in RCIS cells. Quite the contrary, cells became totally resistant against DDR inhibitors when nearing the senescent state. Further, we discovered that CX-5461 affects genomic stability, and that p53 wild type status serves as a key feature in preserving genomic integrity upon CX-5461 treatment. Last but not least, we observed DNA:RNA hybrid formation upon CX-5461 treatment, thus offering new insights in the molecular working mechanism of CX-5461. Based on that, I proposed a novel hypothetic mode of action for CX-5461 focusing on clashes between transcription and replication machineries. This new approach raises the question if DNA:RNA hybrids or such clashes might be critical in senescence induction. Further, targeting proteins involved in DNA:RNA hybrid repair displays a promising starting point for new combination therapy studies aiming to increase the effectiveness of CX-5461 for tumour therapy.