The Myc protein is a multifunctional transcription factor able to regulate several cellular processes. Frequently upregulated in cancerous lesions, Myc overexpression not only causes aberrant cell growth and proliferation, but also triggers a strong DNA Damage Response (DDR), partially caused by replication stress and the consequent accumulation of DNA damage at the level of stalled replication forks. Although the therapeutic targeting of Myc in cancer is highly studied, Myc-targeted drug efficacy and development still have many structural and functional challenges to be overcome. For these reasons, the identification of novel regulators of Myc activities can be exploited for therapeutic purposes.
In order to dissect the Myc-induced DDR and to uncover potential druggable regulators of this response, we set up a high-throughput RNA Interference screen based on the detection of the DNA damage marker gH2A.X at the single cell level. In order to control Myc activity, we took advantage of a conditional cell line in which Myc is fused to the Estrogen Receptor (ER) and can be quickly activated upon 4-hydroxy-tamoxifen (OHT) treatment. Through this screen, we identified both positive and negative regulators of Myc-induced DDR, and several of the candidates that, once inactivated synergized with Myc in inducing DNA damage, showed also synthetic lethality. Among these candidates that may protect Myc-overexpressing cells from broad replicative stress, we focused our attention on factors with a critical role in maintaining genomic stability. In particular, through loss of function experiments, genome wide transcriptional analysis, replication forks monitoring, and the usage of DSBs reporter systems at single cell level, we are interested in defining how these factors help Myc-overexpressing cells to overcome replication stress and genomic instability.
Defining the interplay between Myc and these novel factors will help us to unveil new potential therapeutic targets, whose inhibition could be the basis of specific therapeutic approaches for Myc-driven cancers.