Targeting Menin Protein Stability In AML

项目摘要 多发性内分泌瘤1型（MEN 1/MENIN）是一种转录调节因子，已知与 赖氨酸甲基转移酶2A（KMT 2A/MLL 1）驱动包括MLL 1在内的白血病侵袭性亚群 急性髓细胞白血病（AML）是指急性髓细胞白血病（AML）的一种，包括NPM 1突变和NUP 98重排的急性髓细胞白血病（MLL-r），以及其他急性髓细胞白血病。 癌症包括胃肠道间质瘤。在临床试验中， 破坏MLL 1/MENIN相互作用以治疗引起MENIN的AML的小分子MENIN抑制剂（MI） 成为重要的药物靶点因此，了解MENIN功能的努力对于 这种蛋白质的持续成功。我，作为一个研究员在博士的指导下工作。 斯科特阿姆斯特朗和埃里克菲舍尔，最近的特点是令人兴奋的方面的机制驱动 MI治疗的疗效。我们已经观察到蛋白质稳定性改变为MENIN和其他蛋白质。 蛋白质是MI敏感性的基本特征。我们对这一机制的初步研究 发现特异性E3连接酶的缺失可通过MENIN蛋白介导MI敏感性降低 稳定化这些发现启发了一种新的方法，即用MENIN蛋白稳定性靶向MENIN蛋白， 一个简单的想法，更多的MENIN蛋白降低药物敏感性，而更少的MENIN蛋白抑制MLL-r 转录程序。本提案中概述的实验旨在表征MENIN蛋白 通过研究这种特异性E3连接酶如何调节有助于MI敏感性的MENIN， 研究新建立的快速MENIN降解系统，作为改善这两种方法的原理证明 MENIN靶向治疗以及研究MENIN功能的方法。我们最近成立了一个 研究MENIN蛋白功能的一组新工具，包括（i）具有编辑的MEN 1的MLL-r白血病细胞系 插入不同蛋白质标签的基因座，使得能够实现高通量MENIN蛋白质稳定性报告系统， （ii）靶向MENIN蛋白的特异性CRISPR文库 功能和（iii）早期候选小分子MENIN蛋白水解靶向嵌合体（PROTAC）。这 建议还旨在提供更多的培训经验，在先进的转录方法， 分析，生物化学结合试验，小分子PROTAC设计和质谱为基础的合作， 免疫沉淀以及指导手稿/赠款写作和领导技能。都传递了 两名资源充足的资深导师的指导，一个优秀的科学咨询委员会 由白血病和染色质生物学领域的专家组成， 在DCFI的环境中，这项提案的资金将使我能够过渡到独立调查员。

PROJECT SUMMARY Multiple Endocrine Neoplasia Type 1 (MEN1/MENIN) is a transcriptional regulator known to interact with Lysine Methyltransferase 2A (KMT2A/MLL1) driving aggressive subsets of leukemia including MLL1 rearranged (MLL-r), NPM1 mutated and NUP98 rearranged acute myeloid leukemia (AML) as well as other cancers including gastrointestinal stromal tumors. Early success has been reported in clinical trials using small molecule MENIN inhibitors (MI) that disrupt the MLL1/MENIN interaction to treat AML causing MENIN to emerge as an important drug target. As a result, efforts to understand MENIN function are paramount for the continued success of targeting this protein. I, as a research fellow working under the mentorship of Drs. Scott Armstrong and Eric Fischer, have recently characterized an exciting aspect of the mechanism driving the therapeutic effect of MI treatments. We have observed that protein stability changes to MENIN and other proteins is an essential feature of MI sensitivity. Our preliminary examination of this mechanism has uncovered that the loss of a specific E3 ligase can mediate decreased MI sensitivities via MENIN protein stabilization. These findings have inspired a new approach of targeting MENIN protein stability with the simple idea that more MENIN protein decreases drug sensitivity while less MENIN protein inhibits MLL-r transcriptional programs. The experiments outline in this proposal aim to characterize MENIN protein stability in AML by studying how this specific E3 ligase regulates MENIN contributing to MI sensitivity and investigate newly established rapid MENIN degradation systems as a proof-of-principle for improving both MENIN targeted therapies as well as the means to study MENIN function. We have recently established a novel set of tools to study MENIN protein function including (i) MLL-r leukemia cell lines with edited MEN1 loci inserting different protein tags enabling high throughput MENIN protein stability reporter systems and dTAG based rapid MENIN degradation systems, (ii) specific CRISPR libraries targeting MENIN protein function and (iii) early candidate small molecule MENIN proteolysis targeting chimeras (PROTAC). This proposal is also designed to provide additional training experiences in advanced methods of transcriptional analysis, biochemical binding assays, small molecule PROTAC design and mass spectrometry-based co- immunoprecipitation as well as mentoring in manuscript/grant writing and leadership skills. Anchored by the guidance of two well-resourced and established mentors, an excellent scientific advisory committee composed of experts in the fields of leukemia and chromatin biology and a highly dynamic research environment at DCFI, funding for this proposal will enable me to transition to an independent investigator.