Abstract 6286: Targeting Polycomb Repressive Complex 1 with small-molecule inhibitors as a potential therapeutic approach to combat leukemia (2023)

Abstract Background: Polycomb repressive complex 1 (PRC1) is an epigenetic regulatory complex that monoubiquitinates lysine 119 on histone H2A, resulting in the repression of target genes. The canonical core of PRC1 responsible for its E3 ligase activity is a heterodimeric unit comprised of RING1B and BMI1. Emerging data implicates RING1B in various cancers and highlights the importance of PRC1 function in the maintenance and proliferation of leukemic stem cells. Studies demonstrate that loss of PRC1 activity through RING1B knockdown impairs leukemic stem cell proliferation, causes leukemic stem cell differentiation, and enhances survival in MLL-AF9 mouse models. Therefore, development of potent small molecules targeting the activity of PRC1 might offer a new approach for generating new therapeutics for leukemia. Results: Here, we report the discovery and development of the first-in-class small-molecule inhibitors directly binding to RING1B in PRC1. We performed an NMR-based fragment screen against RING1B-BMI1, identifying fragment hit, RB-1, with low-millimolar binding affinity. RB-1 was extensively optimized into a class of inhibitors with considerably enhanced binding affinity to the low-nanomolar range. These inhibitors have been shown to diminish the H2A ubiquitination activity of PRC1 E3 ligase cores. Structural studies revealed that our PRC1 inhibitors bind directly to the RING domain of RING1B and ultimately block the protein-protein interaction between RING1B and the nucleosome. In leukemic cell lines and primary AML samples, PRC1 inhibitor treatment elicits an anti-proliferative effect on cell growth and displays on-target action seen by the reduction of the H2A ubiquitination mark. Additionally, small-molecule inhibition of PRC1 results in the induction of cellular differentiation of leukemic cells, as well as apoptosis. Gene expression analysis in leukemic cells treated with our PRC1 inhibitors reveals upregulation of multiple genes that are repressed by PRC1, which is also supported by the displacement of PRC1 from chromatin assessed by CHIP-seq studies. Furthermore, our PRC1 inhibitors are active in vivo in mouse models of leukemia, demonstrating reduction of leukemia progression upon treatment. Conclusions: We developed first-in class inhibitors directly targeting the E3 ligase core of PRC1. Treatment with our PRC1 inhibitors in leukemic cells elicit an anti-proliferative effect, blocks ubiquitination of H2A, and results in the de-repression of many genes repressed by PRC1. Additionally, our PRC1 inhibitors elicit an anti-proliferative effect on leukemia progression in vivo, demonstrating the traction for developing anti-leukemia therapeutics that inhibit PRC1. Citation Format: Miranda L. Simes, Yiwu Yao, Se Ra Park, Trupta Purohit, Hongzhi Miao, Xiaotian Zhang, Weijiang Ying, Felicia Gray, Shirish Shukla, Qingjie Zhao, Caroline Nikolaidis, Alyssa Winkler, Jolanta Grembecka, Tomasz Cierpicki. Targeting Polycomb Repressive Complex 1 with small-molecule inhibitors as a potential therapeutic approach to combat leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6286.