MECHANISTIC INVESTIGATION OF BRD4 INHIBITION IN MYC DEPENDENT TUMORS [Tesi di dottorato]

Università degli Studi di Milano, 2016-03-18

The c-myc gene encodes for a transcription factor involved in the regulation of different cellular mechanisms, ranging from cell cycle control and apoptosis to cellular metabolism. Myc is frequently altered in human cancer either by genomic rearrangement or by alteration of upstream regulatory pathways. Myc crucial role both in tumor formation and maintenance makes it an attractive molecular target for cancer therapy. Unfortunately, Myc is intrinsically resilient to direct pharmacological targeting using small molecules. To overcome this issue, alternative therapeutic avenues have been explored. In the last years, independent groups showed that BET proteins inhibition leads to a strong Myc downregulation in Multiple Myelomas and Acute Myeloid Leukemias, with consequent cell cycle arrest and tumor regression. To support the hypothesis of a direct and specific effect on Myc levels mediated by BET proteins, two different working models were proposed depending on c-myc location (translocated versus endogenous). In order to extend these observations and improve our understanding of the mechanism of action of BETs inhibitors, we evaluated global transcriptional alteration and chromatin profiles in Burkitt’s Lymphomas in response to JQ1. Our results demonstrate that BETs inhibitors efficacy is dependent on global alteration of RNA PolII dynamics, due to the role of BRD4 in regulating elongation. Yet, despite a pervasive eviction of BRD4 from chromatin and the global effect on RNA PolII observed following BETs inhibition, the transcriptional alterations are limited to a subset of genes. These genes are characterized by promoter regions heavily marked by H3K27Ac, high binding of BRD4 and Transcription Factors (Myc and E2F1) and RNA PolII. These JQ1 sensitive genes are consistent among different cell lines and characterized by high expression levels. Prominent promoter saturation and high RNA PolII pausing render their expression rate-limited by transcriptional elongation. Indeed the same genes are selectively targeted by pharmacological treatments affecting components of the elongation machinery. Thus, selective transcriptional effects following JQ1 treatment are linked to BETs role in regulating transcriptional elongation. These observations highlight the role of BETs protein in regulating gene expression and provide a rationale to explain how broad inhibition of elongation may lead to a selective transcriptional response..

Settore MED/04 - - Patologia Generale