Targeting Distinct Promoter- and Enhancer-Driven Dependencies in Multiple Myeloma

Abstract

Uncontrolled proliferation is a hallmark of tumorigenesis and is associated with perturbed transcriptional profiles. The proliferative program in Multiple myeloma (MM), a complex disease with heterogeneous genetic changes, is controlled by transcription factors (TFs) and chromatin-associated factors. The dependency on these transcriptional regulators, leading to the dysregulated proliferation, is not predicted by genetic changes, making the tumor cells more sensitive to inhibition of these regulators than normal cells.The relationship between promoter proximal transcription factor-associated gene expression and super-enhancer-driven transcriptional programs is not well-defined. However, their distinct genomic occupancy suggests a mechanism for specific and separable gene control. Our genome-wide epigenomic profile in myeloma has identified the existence of two non-overlapping regulatory axes controlled by promoter and enhancer-driven processes, governing distinct biological functions. We have utilized E2F1 as promoter proximal transcription factor, and evaluated its transcriptional and functional interrelationship with enhancer-associated factors, such as BET bromodomain transcriptional co-activators. We identified that the transcription factor E2F1 and its heterodimerization partner DP1 represent a dependency in MM cells. Global chromatin analysis revealed two distinct regulatory axes, with E2F and MYC predominantly localized to active gene promoters of growth/proliferation genes and CDK9 and BETs disproportionately at enhancer-regulated tissue-specific genes. This divergent BRD4 enhancer and E2F promoter axes is also observed in diffuse large B-cell lymphoma, suggesting a more broader transcriptome control process.Dual inhibition of E2F and BETs displays a superior activity against MM cell growth and viability, both in vitro and in vivo, compared to single perturbation alone providing an important molecular mechanism for combination therapy. Moreover, at low doses of BRD4 inhibitor JQ1, the addition of E2F1 depletion down-regulates the promoter controlled proliferation gene expression axis.As for many TFs, direct pharmacologic inhibition of E2F remains a difficult challenge in drug discovery. However, E2F is not entirely “undruggable” as inhibitors of upstream regulators of the pRB-E2F axis are available. For example, a number of Cyclin dependent kinases (CDK) 4/6 inhibitors, including Palbociclib are now being investigated in clinical trials in in fact approved by the FDA in select malignancies. CDKs are serine threonine kinases that modulate cell cycle progression. CDK4 and CDK6 together with D-type cyclins and cyclin E/CDK2 complexes control the commitment to cell cycle entry from quiescence and the G1 phase. These kinase complexes can phosphorylate RB, releasing E2F to modulate the expression of E2F target genes that are required for S phase entry. We investigated combination of low doses of JQ1 and Palbociclib and observed a profound effect on E2F promoter driven transcriptional activity, and was highly synergistic with JQ1 in a large panel of MM cell lines and primary MM cells from newly diagnosed and relapsed patients. Cell cycle analysis revealed complete G1 arrest after treatment. Importantly, the combination regimen was not effective in healthy donor PBMCs activated with PHA, suggesting a favorable therapeutic index. Transcriptomic changes to assess the impact on promoter and SE-driven processes are ongoing and will be presented.In conclusion, these data implicates the existence of a sequestered cellular functional control that may be perturbed in cancer to maintain the tumor cell state. Simultaneous targeting of non-overlapping promoter and enhancer vulnerabilities impairs the myeloma proliferative program, with potential for development of a promising therapeutic strategy in MM and other malignancies. DisclosuresYoung:Omega Therapeutics: Consultancy, Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Syros Pharmaceuticals: Consultancy, Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Camp4 Therapeutics: Consultancy, Equity Ownership, Membership on an entity's Board of Directors or advisory committees. Anderson:Bristol Myers Squibb: Consultancy; C4 Therapeutics: Equity Ownership, Other: Scientific founder; Celgene: Consultancy; Millennium Takeda: Consultancy; Gilead: Membership on an entity's Board of Directors or advisory committees; OncoPep: Equity Ownership, Other: Scientific founder. Munshi:OncoPep: Other: Board of director.