Abstract 4670: Identification of a possible therapeutic candidate for multiple myeloma based on dual inhibition of EZH1/EZH2
Authors: Nakagawa Makoto , Fujita Shuhei , Honma Daisuke , Araki Kazushi , Kitabayashi Issay 
Published: July 2017

Multiple myeloma (MM) is largely incurable as the disease eventually relapses despite the recent development of novel therapies. Previous reports show that side population (SP) cells comprise myeloma stem cells. Therefore, targeting SP cells may be a promising strategy for preventing and treating MM relapse. Polycomb repressive complexes 1 (PRC1) and 2 (PRC2) are important epigenetic regulators that maintain the “stemness” of ES cells and other hematopoietic stem cells. Enhancer of zeste homolog 1 and 2 (EZH1/2) are catalytic components of PRC2, which trimethylate histone H3 at lysine 27 to repress transcription of target genes. Mutation and overexpression of EZH2 are associated with many cancers, including MM. Here, we found that SP cells expressed significantly higher levels of EZH1/2 than non-SP cells. These results suggest that overexpression of EZH1/2 is important for maintaining the stemness of MM cells and that EZH1/2 could be a potential therapeutic target. We developed a novel EZH1/2 dual inhibitor, OR-S1, and used it to investigate the effect of pharmacologic inhibition of EZH1/2 on MM. OR-S1 suppressed the proliferation of almost all MM cell lines tested, with an IC50 significantly lower than that of the specific EZH2 inhibitor, GSK126. Furthermore, flow cytometry analysis revealed that OR-S1 significantly depleted the SP cell population. RNA-seq analysis revealed that the transcriptional profiles of MM cell lines treated with OR-S1 were characterized by up-regulation of genes related to the Wnt pathway. qRT-PCR confirmed that expression of Wnt, Frizzled, and Protein kinase C family members increased markedly after exposure to OR-S1. Previous studies show that increased activation of the canonical Wnt signaling pathway down-regulates HSC self-renewal and differentiation. Therefore, we generated β-catenin-overexpressing MM cells to examine the effects of increased Wnt signaling on MM cells. Surprisingly, proliferation of these cells was significantly lower than that of control cells. These results suggested that PRC2 directly targets Wnt signaling, and that over-activation induced by EZH1/2 dual inhibition was responsible for the reduced proliferation of MM cells. Oral administration of OR-S1 to mice bearing MM xenografts led to significant impairment of subcutaneous tumors. Interestingly, long-term administration of the drug at lower doses to mice bearing orthotopic xenografts resulted in complete eradication of minimal residual disease from the bone marrow and complete cure of MM without any serious side effects. Furthermore, OR-S1 treatment of an orthotopic PDX model derived from a relapsed and heavily pretreated MM patient led to a reduction in the levels of human immunoglobulins in the serum. Taken together, these results strongly suggest that dual inhibition of EZH1/2 is a promising therapeutic approach to eradicating myeloma stem cells and could lead to important advances in the treatment of MM.

Citation Format: Makoto Nakagawa, Shuhei Fujita, Daisuke Honma, Kazushi Araki, Issay Kitabayashi. Identification of a possible therapeutic candidate for multiple myeloma based on dual inhibition of EZH1/EZH2 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4670. doi:10.1158/1538-7445.AM2017-4670