Sir Peter MacCallum Department of Oncology - Theses
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ItemBET bromodomain inhibition as combined apoptotic and immunomodulatory therapy for the treatment of MYC-driven lymphoma( 2017)Bromodomain and Extra-Terminal (BET) proteins are a conserved family of ‘epigenetic readers’ that bind to acetylated lysine residues on histone and non-histone proteins to modulate transcription. BET proteins are enriched at promoter and enhancer regions and recruit the positive transcription elongation factor b (P-TEFb) complex to activate RNA polymerase II. Anti-tumour responses elicited by BET inhibitors have been associated with the suppression of genes required for cellular proliferation and survival, including oncogenic transcription factors. Suppression of the proto-oncogene MYC was initially reported as a key mechanistic property of BET inhibitors, however more recent evidence suggests that additional target genes are mechanistically implicated. In this thesis, the Eμ-Myc model of aggressive B-cell lymphoma was utilized to investigate the full repertoire of genes modulated by JQ1 and their functional significance in mediating therapeutic responses. JQ1 did not suppress the expression of transgenic Myc in this model, allowing the determinants of apoptosis induction to be assessed, independently of changes in Myc expression. This apoptotic response was p53-independent and associated with modulation in the ratio of pro- and antiapoptotic Bcl-2 family members to favor activation of the intrinsic mitochondrial apoptotic pathway. Therapeutic administration of JQ1 to mice bearing Eμ-Myc lymphomas led to robust clinical responses, however, universal treatment failure was observed despite ongoing therapy. Using RNA-Seq, disease progression and secondary JQ1 resistance was found to be associated with RAS pathway activation and Bcl-2 upregulation. In addition, the efficacy of JQ1 was found to be dependent on an intact host immune system, where a 50% reduction in the survival advantage was observed upon transplantation into immune-deficient mice. Using RNA-Seq, the immune checkpoint ligand Cd274 (Pd-l1) was found to be potently suppressed by JQ1. Mechanistically, BET inhibition decreased Brd4 occupancy at the Cd274 promoter, leading to promoter-proximal pausing of RNA polymerase II, and loss of Cd274 mRNA production. Rapid epigenetic remodeling of the Cd274 locus in response to interferon gamma (IFN-γ) stimulation led to recruitment of Irf1, Brd4, RNA polymerase II, as well as increased local histone acetylation. Accordingly, BET inhibition suppressed constitutive and IFN-γ-induced PD-L1 expression in genetically diverse tumour models. Ectopic expression of PD-L1 in Eμ-Myc lymphomas was sufficient to reduce the efficacy of JQ1, demonstrating the significance of PD-L1 suppression to the observed therapeutic responses associated with BET inhibition. Finally, treatment of mice bearing Eμ-Myc lymphomas with JQ1 in combination with a checkpoint inhibitor (anti-PD-1) or immune stimulating antibody (anti-4-1BB/CD137) led to improved therapeutic responses. The results presented herein demonstrate the importance of MYC-independent apoptotic signaling to therapeutic responses associated with BET inhibition, as well as acquired drug resistance. In addition, these results demonstrated the ability of BET inhibitors to directly engage the host immune response during anti-cancer therapy. Finally, BET inhibitors can suppress oncogenic PD-L1 transcription for therapeutic gain, leading to augmented anti-tumour immunity. These studies establish a strong rationale for clinical investigation of BET inhibitors in combination with immune modulating therapies.