Abstract 449: Small molecular inhibitors of PRC1 demonstrate potent activity in acute leukemia models
Authors: SeRa Park, Yiwu Yao, Sydney Musser, Miranda Simes, Haiqing He, Hongzhi Miao, Alyssa Winkler, Trupta Purohit, Xiaotian Zhang, Jolanta Grembecka, Tomasz Cierpicki
Journal: Cancer Research (2025)

Abstract Background: Polycomb repressive complex 1 (PRC1) is an epigenetic regulatory complex that monoubiquitinates H2A on lysine 119 and is involved in gene regulation and development, resulting in the repression of target genes important for cellular identity, stemness and differentiation. Activity of PRC1 is maintained by E3 ligases RING1A or RING1B. Recent studies implicate RING1B in various cancers and highlights the importance of PRC1 function in the maintenance and proliferation of leukemic stem cells. This demonstrated that PRC1 has role in acute leukemias, supporting the need to develop PRC1 inhibitors. Therefore, development of potent small molecule inhibitors targeting PRC1 can provide novel therapeutic new approach for leukemia treatment. Results: We have developed potent PRC1 inhibitors represented by our lead compound RB-231. We found that treatment with RB-231 induces potent anti-proliferative effects on cell growth at mid-nanomolar concentrations in multiple acute leukemia cell lines. Analysis of H2A ubiquitination using Western Blots have shown that activity of RB-231 is correlated with global reduction of H2Aub levels. Profiling of gene expression in various AML and ALL cell lines demonstrates de-repression of many genes. Through mechanistic studies using Cut&amp;Run, we validated decrease in H2Aub signal at multiple gene loci, including CDKN1A. We performed Flow cytometry to verify apoptosis and cell differentiation in AML cells. These studies revealed that RB-231 treatment induces cellular differentiation as well as apoptosis in leukemia cells. We also tested the activity of RB-231 in primary AML patient samples. Treatment with RB-231 decreased colony formation in patient samples and induced morphological changes, suggesting differentiation of these cells from leukemic blasts to more mature hematopoietic cells. We also evaluated the activity of RB-231 in normal human CD34+ bone marrow cells to confirm no toxicity. We assessed efficacy of RB-231 in vivo using MV4;11, Jurkat and Reh xenografts. Treatment with PRC1 inhibitor resulted in pronounced reduction of leukemia burden and increase in survival. In accompanying pharmacodynamics PD experiments, RB-231 showed reduced leukemia hCD45+ blast level in blood, bone marrow and spleen compared to the vehicle control, and increased CDKN1A expression. Conclusion: We developed a first- in class inhibitor directly targeting the E3 ligase activity of PRC1. Treatment with our PRC1 inhibitors in leukemic cells elicits an anti-proliferative effect, blocks ubiquitination of H2A, and results in the de-repression of many genes repressed by PRC1. Our data suggests that multiple subtypes of acute leukemia are sensitive to inhibition of PRC1. Furthermore, this study demonstrated that RB-231 is potent PRC1 inhibitor suitable for evaluation in vitro and in vivo models of acute leukemia. Acknowledgement: This research was funded by R01CA282082 grant from NIH. Citation Format: SeRa Park, Yiwu Yao, Sydney Musser, Miranda Simes, Haiqing He, Hongzhi Miao, Alyssa Winkler, Trupta Purohit, Xiaotian Zhang, Jolanta Grembecka, Tomasz Cierpicki. Small molecular inhibitors of PRC1 demonstrate potent activity in acute leukemia models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 449.