Identification of a First-in-Class SETD2 Inhibitor That Shows Potent and Selective Anti-Proliferative Activity in t(4;14) Multiple Myeloma: T(4;14) Multiple Myeloma Cells Are Dependent on Both H3K36 Di and Tri-Methylation

Abstract

t(4;14) chromosome translocations are found in 15% of newly diagnosed multiple myeloma (MM) cases and are associated with high risk. MM cells with t(4;14) over-express the histone methyltransferase (HMT), WHSC1/MMSET/NSD2, which leads to deregulation of gene expression due to increased di-methylation of Histone H3 at lysine 36 (H3K36me2). This activity has been shown to be essential for the survival of t(4;14) MM cells. In addition to WHSC1, another HMT, SETD2, has been shown to methylate H3K36. SETD2 is the only known enzyme capable of tri-methylation of H3K36 and has been reported to play a role in transcriptional elongation and alternative splicing. CRISPR pooled screening has shown that SETD2 activity is required for viability of a variety of cancer cell lines. This led Epizyme to develop small molecule inhibitors of SETD2 enzyme activity in order to understand the role of SETD2 in tumorigenesis. Through our drug discovery efforts, we identified EPZ-040414, a potent and selective inhibitor of SETD2 with low nM cell biochemical activity and broad selectivity against a panel of other HMTs. The proposed role of SETD2 in H3K36 methylation led us to test a panel of MM cells, including 6 t(4;14) cell lines with EPZ-040414. Inhibition of SETD2 resulted in reduced global tri-methylation of H3K36 in t(4;14) bearing MM cell lines. In contrast, there was no effect on global di-methyl H3K36 levels, indicating that WHSC1 activity is not affected by SETD2 inhibition. Moreover, 5/6 t(4;14) MM cell lines showed a cytotoxic response to treatment with EPZ-040414 with IC50s ranging between 60 and 200 nM, while all non-t(4;14) MM cell lines showed limited responses between 1 and 8 μM. Moreover, screening of a 280 cancer cell line panel with a SETD2 inhibitor showed minimal anti-proliferative activity in most cell lines tested. These findings show that t(4;14) MM cell lines require SETD2 activity for survival, suggesting that SETD2 inhibitors are strong candidates for the treatment of this high risk subgroup of MM. Efforts to further understand the interaction between SETD2 and WHSC1 in the molecular pathogenesis of t(4;14) myeloma will be presented. The current chemical series represented by EPZ-040414 is potent, selective, orally available, and currently under further evaluation for its therapeutic potential. Figure. Figure. Disclosures Thomenius: Epizyme Inc.: Employment, Equity Ownership. Totman:Epizyme Inc.: Employment, Equity Ownership. Cosmopoulos:Epizyme Inc.: Employment, Equity Ownership. Brach:Epizyme Inc.: Employment, Equity Ownership. Ci:Epizyme Inc.: Employment, Equity Ownership. Farrow:Epizyme Inc.: Employment, Equity Ownership. Smith:Epizyme Inc.: Employment, Equity Ownership. Chesworth:Epizyme Inc.: Employment, Equity Ownership. Duncan:Epizyme Inc.: Employment, Equity Ownership. Tang:Epizyme Inc.: Employment, Equity Ownership. Riera:Epizyme Inc.: Employment, Equity Ownership. Lampe:Epizyme Inc.: Employment, Equity Ownership.

Discovery of a Non-Nucleoside SETD2 Methyltransferase Inhibitor against Acute Myeloid Leukemia

“…However, as sinefungin is not cell-permeable, it cannot be used for cellular studies. It is worth noting that SETD2 inhibition also generated interest in the pharma sector, since Epizyme (now part of Genentech) reported a potent SETD2 inhibitor with a yet-undisclosed structure [ 22 ].…”

Drug Discovery Targeting Nuclear Receptor Binding SET Domain Protein 2 (NSD2)

“…275 Researchers from Epizyme Inc. disclosed a patent that SETD2 inhibitors with a scaffold of substituted indoles or indolines were used to treat or slow the progression of cancers with NSD2-overexpression, including t(4;14) MM, suggesting that SETD2 inhibitors used as a single agent or in combination with an NSD2 inhibitor might provide a potential therapy for cancers. 276 Some compounds described in this patent significantly suppress the growth of KMS-34 cells in a dosedependent manner. Moreover, these compounds can significantly reduce the level of H3K36me3 while showing no effect on H3K36me2, which was directly associated with its obvious antiproliferation activity.…”

“…NSD2 inhibitors, including polypeptides, polynucleotides comprised of antisense molecule, siRNA and shRNA, and small molecules DZNep, MCTP-39, 10 (LEM06), Ku55933, and AZD0516, were disclosed to increase the sensitivity of diverse cancer cells (e.g., PCa, breast cancer, colorectal cancer, pancreatic cancer, and gastric cancer) to chemotherapeutic agents such as doxorubicin, etoposide, and PI3K inhibitors BKM120BYL719 and RP6530, indicating that combination use of NSD2 and PI3K inhibitors simultaneously shows synergetic antitumor effect . Researchers from Epizyme Inc. disclosed a patent that SETD2 inhibitors with a scaffold of substituted indoles or indolines were used to treat or slow the progression of cancers with NSD2-overexpression, including t(4;14) MM, suggesting that SETD2 inhibitors used as a single agent or in combination with an NSD2 inhibitor might provide a potential therapy for cancers . Some compounds described in this patent significantly suppress the growth of KMS-34 cells in a dose-dependent manner.…”

“…In recent years, developing combination therapy has been gaining increasing attention, especially in the field of cancers, due to its potential superiority to monotherapies, including synergetic therapeutic effects and decreased drug resistance. − Moreover, combination therapy has been universally implemented for epigenetic targets and demonstrated additive and synergetic therapeutic effects. , Substantial efforts have been made in developing combination therapy for NSD2-associated diseases to improve the therapeutic potential. ,,,− …”