Lead Scientist Dana-Farber Cancer Institute boston, Massachusetts, United States
Introduction: Multiple myeloma (MM) is characterized by a non-oncogene addiction to the plasma cell lineage factor IRF4, making it a high-value therapeutic target. Using a transcription regulatory network (TRN)-based approach, we here defined the key regulatory activity of IRF4 and its druggable co-factors. This analysis identified lysine acetyl transferase (KAT) function of p300 as the nearest selective neighbor for therapeutic intervention whose deletion mirrors IRF4 deletion. Protein interactome analysis in MM cells revealed direct interaction between IRF4, p300 and key MM transcription factors; and a significant overlap between IRF4 interactome and p300 acetylation substrates. Furthermore, IRF4 and p300 colocalized significantly across the genome at important MM regulatory elements. These data confirmed the existence of an IRF4-p300 convergent functional network in MM cells, that can be perturbed for therapeutic purposes.
Methods: Since TRN analysis suggested a more potent and potentially selective effect of p300/CBP KAT inhibition than historically observed with p300/CBP bromodomain inhibitors, we developed KB528, a highly selective KAT domain inhibitor of p300 and its close homolog CBP. We investigated its activity in vitro in MM cell lines and ex vivo in bone marrow mononuclear cells (BMMNCs) obtained from bone marrow aspirates of MM patients using luminescence- and flow cytometry-based assays; and in vivo in xenograft MM mouse models. Transcriptomic and epigenomic changes after treatment were evaluated by RNA-seq and ChIP-seq analysis respectively.
Results: Treatment of MM cells with KB528 downregulated IRF4 mRNA and protein levels; and caused a reduction in chromatin acetylation at highly specific sites in the genome co-localized with IRF4, in contrast to broader agents like BET bromodomain inhibitors, as well as a disruption of the IRF4 transcriptomic signature. Furthermore, KB528 had a strong antiproliferative effect in MM cell lines, regardless of their cytogenetics or drug sensitivity profile, by downregulating the IRF4 TRN and inducing apoptosis more effectively than p300/CBP bromodomain inhibitors. Ex vivo treatment of BMMNCs of MM patients with stable (n=6) or relapsed-refractory (n=6) disease confirmed induction of apoptotic cell death on CD138+ primary MM cells, while sparing CD138-negative cells, supporting a therapeutic index. Since most successful regimens in MM therapy are combinatorial, we evaluated the activity of KB528 in combination with existing SOC agents and observed synergistic activity in combination with IMiDs and dexamethasone. Finally, KB528 demonstrated activity in vivo, with robust tumor growth inhibition at tolerated doses, together with reduced p300-dependent substrate acetylation and loss of IRF4.
Conclusions: IRF4 dependency is a hallmark of MM that has been challenging to target with existing therapies and this study lays the groundwork for developing p300 KAT inhibitors for MM therapy.