Abstract B126: A small molecule inhibitor of the RNA m6A writer METTL3 inhibits the development of acute myeloid leukemia (AML) in vivo
Abstract B126: A small molecule inhibitor of the RNA m6A writer METTL3 inhibits the development of acute myeloid leukemia (AML) in vivo
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DOI：
10.1158/1535-7163.targ-19-b126
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发表时间：
2019-12
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通讯作者：
M. Albertella;W. Blackaby;R. Fosbeary;A. Hendrick;Dan Leggate;Y. Ofir-Rosenfeld;A. Sapetschnig;K. Tzelepis;E. Yankova;T. Kouzarides;Oliver Rausch-
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作者：
M. Albertella;W. Blackaby;R. Fosbeary;A. Hendrick;Dan Leggate;Y. Ofir-Rosenfeld;A. Sapetschnig;K. Tzelepis;E. Yankova;T. Kouzarides;Oliver Rausch-
METTL3是一种RNA甲基转移酶，负责n -6-甲基腺苷（m6A）在SP1等mRNA靶标上的沉积，以调节其稳定性和表达。METTL3被认为是AML细胞生长的重要基因，并被认为是癌症治疗的新靶点（Barbieri 2017）。我们提出了METTL3的新型小分子抑制剂的体外和体内表征，利用药理学审计线索概括了METTL3作为新型癌症靶点的遗传验证。利用结构导向的药物化学平台对2个不同化学系列的小分子抑制剂进行了鉴定和优化。化合物1和2来自不同的系列，均表现出对METTL3酶的生物化学抑制作用，具有个位数纳摩尔效价。SPR证实与METTL3的直接结合具有相当的效力。化合物3是一种无活性的类似物，在酶分析（IC50 - 50微米）中被证实无活性。化合物1-2对METTL3具有选择性，对其他RNA、DNA或蛋白质甲基转移酶没有抑制作用（10微米IC50）。化合物1和2以亚微摩尔的效力抑制SP1和Brd4蛋白的表达，而无活性的类似物化合物3则没有作用，从而证实了细胞靶标的作用。化合物1和2对MOLM13细胞的增殖有抑制作用，这与SP1抑制作用有关，而化合物3则没有作用，说明它们的活性依赖于mettl3。化合物1具有良好的口服生物利用度，在小鼠中具有良好的剂量比例暴露，半衰期为3.5小时，耐受性良好，无体重减轻或临床症状。在MLL-AF9驱动的原发性小鼠AML模型中，化合物1的抗肿瘤作用被评估。与对照组相比，每日口服化合物1 30mg /kg可显著减少AML扩张和脾脏重量，表明其在体内具有明显的抗肿瘤作用。通过mettl3依赖性m6A靶标的减少，骨髓和脾脏的靶标结合得到证实。我们描述了METTL3 RNA甲基转移酶的有效和选择性抑制剂的综合特性，并通过生化、细胞和体内系统证明了它们的活性和效用。我们已经证明，体内小分子抑制METTL3在急性髓性白血病的生理相关模型中具有明显的抗肿瘤作用。据我们所知，这是第一次证明RNA甲基转移酶抑制剂的体内活性，并证明RNA修饰酶是开发新型癌症治疗方法的新靶点。引文格式：Mark Albertella， Wesley Blackaby, Richard Fosbeary, Alan Hendrick, Dan Leggate, Yaara ofirl - rosenfeld, Alexandra Sapetschnig, Konstantinos Tzelepis, Eliza Yankova, Tony Kouzarides, Oliver Rausch。一种小分子的RNA m6A书写者抑制剂METTL3在体内抑制急性髓性白血病（AML）的发展[摘要]。摘自：AACR-NCI-EORTC分子靶点和癌症治疗国际会议论文集；2019年10月26日至30日；波士顿,MA。费城（PA）： AACR；巨蟹座2019；18（12增刊）：摘要nr B126。doi: 10.1158 / 1535 - 7163. -目标- 19 - b126
METTL3 is an RNA methyltransferase which is responsible for the deposition of N-6-methyladenosine (m6A) on mRNA targets such as SP1, to modulate their stability and expression. METTL3 was identified as an essential gene for the growth of AML cells and proposed as a novel target for cancer therapy (Barbieri 2017). We present the in vitro and in vivo characterization of novel small molecule inhibitors of METTL3, which recapitulate the genetic validation of METTL3 as a novel cancer target using a pharmacological audit trail. Small molecule inhibitors from 2 distinct chemical series were identified and optimised using a structure-guided medicinal chemistry platform. Compounds 1 and 2 are from different series, and both showed biochemical inhibition of METTL3 enzyme with single digit nanomolar potency. Direct binding to METTL3 was confirmed by SPR with comparable potency. Compound 3 is an inactive analog which was confirmed inactive in the enzyme assay (IC50 >50microM). Compounds 1-2 are selective for METTL3 and did not inhibit a panel of other RNA, DNA or protein methyltransferases tested (>10microM IC50). Cellular target engagement was confirmed by the demonstration that compounds 1 and 2 inhibited SP1 and Brd4 protein expression with submicromolar potency, whereas the inactive analog compound 3 had no effect. Compounds 1 and 2 treatment of MOLM13 cells inhibited their proliferation which correlated with SP1 inhibition, and compound 3 had no effect, demonstrating that their activity was METTL3-dependent. Compound 1 has excellent oral bioavailability with good dose-proportional exposure in mice and a half-life of 3.5 hrs, and was well-tolerated with no body weight loss or clinical signs. Compound 1 was evaluated for anti-tumor effects in an MLL-AF9 driven primary murine AML model. 30 mg/kg daily oral dosing of compound 1 gave a significant reduction in AML expansion and a reduction in spleen weight compared to vehicle control, indicating a pronounced anti-tumor effect in vivo. Target engagement was confirmed in bone marrow and spleen as measured by reduction of METTL3-dependent m6A targets. We have described the comprehensive characterization of potent and selective inhibitors of the METTL3 RNA methyltransferase, and demonstrated their activity and utility using biochemical, cellular and in vivo systems. We have demonstrated that inhibition of METTL3 by small molecules in vivo leads to a pronounced anti-tumor effect in a physiologically relevant model of acute myeloid leukemia. To our knowledge, this is the first demonstration of in vivo activity of inhibitors of an RNA methyltransferase and providing proof of concept that RNA modifying enzymes are a new target class for the development of novel cancer therapeutics. Citation Format: Mark Albertella, Wesley Blackaby, Richard Fosbeary, Alan Hendrick, Dan Leggate, Yaara Ofir-Rosenfeld, Alexandra Sapetschnig, Konstantinos Tzelepis, Eliza Yankova, Tony Kouzarides, Oliver Rausch. A small molecule inhibitor of the RNA m6A writer METTL3 inhibits the development of acute myeloid leukemia (AML) in vivo [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr B126. doi:10.1158/1535-7163.TARG-19-B126