Targeting Lysine Methyltransferases EZH2 and EZH1 for Treating MLL-rearranged Leukemias

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Mixed Lineage Leukemia (MLL) rearrangements are responsible for approximately 70% of infant acute myeloid, lymphoid or mixed-lineage leukemia and about 7-10% of adult cases. Leukemias bearing MLL rearrangement display a particularly poor prognosis with a significantly lower rate of survival. To date, no targeted agents have been approved and clinicians currently rely on relatively non-specific cytotoxic agents that are largely ineffective, thus, highlighting an unmet medical need. EZH2 (enhancer of zeste homolog 2) and EZH1 are the two closely related enzymes that catalyze tri-methylation of histone H3 lysine 27 (H3K27me3), a transcriptionally repressive post-translational modification. It has been shown that EZH2 and EZH1 compensate for one another and both enzymes are critical for sustaining MLL-rearranged leukemias. Therefore, we hypothesized that pharmacological targeting of both EZH2 and EZH1 would provide a novel, targeted therapeutic approach for treating MLL-rearranged leukemias. We previously discovered UNC1999, the only small-molecule inhibitor with high potency (IC50 < 50 nM) for both EZH2 and EZH1. We have shown that UNC1999 suppressed the proliferation of MLL-rearranged leukemia cells in both cellular and mouse cancer models. However, in vivo efficacy of UNC1999 is modest and its tumor-killing action is slow. To address this major weakness, we have applied the PROTAC (proteolysis targeting chimeras) technology to generating bivalent inhibitors of EZH2 and EZH1 by linking UNC1999 to an E3 ubiquitin ligase-binding moiety. We have developed prototype bivalent inhibitors, which inhibited the EZH2/1 enzymatic activity, significantly reduced EZH2 protein levels while UNC1999 did not, were much more effective at inhibiting the proliferation of multiple tumor cell lines than UNC1999, did not exhibit toxicity in non-tumor cells, and were orally bioavailable. Based on these promising preliminary results, we propose to: (1) optimize bivalent inhibitors of EZH2 and EZH1 so that they have potency, selectivity, pharmacokinetic and other drug-like properties consistent with a drug candidate, and (2) evaluate bivalent inhibitors of EZH2 and EZH1 in MLL-rearranged leukemia cellular and mouse models. Our goal is to generate a drug candidate for the treatment of MLL-rearranged leukemias.