Targeting Menin Protein Stability In AML

项目总结 多发性内分泌肿瘤1型(MEN1/Menin)是一种已知与之相互作用的转录调节因子 赖氨酸甲基转移酶2A(KMT2A/MLL1)驱动包括MLL1在内的白血病侵袭性亚群 重排(MLL-r)、NPM1突变和NUP98重排急性髓系白血病(AML)等 包括胃肠道间质瘤在内的癌症。据报道，早期的成功在临床试验中使用 阻断MLL1/Menin相互作用的小分子Menin抑制剂(MI)治疗AML引起的Menin 成为重要的毒品目标。因此，努力了解Menin的功能对 靶向这种蛋白质的持续成功。我，作为一名研究员，在麦肯锡博士的指导下工作。 斯科特·阿姆斯特朗和埃里克·费舍尔最近描述了驱动机制令人兴奋的一面 心肌梗死治疗的疗效。我们观察到，蛋白质的稳定性改变为梅宁和其他 蛋白质是心肌梗死敏感性的基本特征。我们对这一机制的初步研究已经 发现特定的E3连接酶的缺失可以通过Menin蛋白介导MI敏感性的降低 稳定状态。这些发现启发了一种新的方法，通过使用 简单地说，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。这项提案中的实验大纲旨在表征Menin蛋白 通过研究这种特定的E3连接酶如何调节Menin对心肌梗死的敏感性和稳定性 研究新建立的薄荷素快速降解系统，作为改善这两种情况的原则证明 Menin靶向治疗以及研究Menin功能的手段。我们最近建立了一个 研究Menin蛋白功能的一套新工具，包括(I)编辑了MEN1的MLL-r白血病细胞系 插入不同蛋白质标签的基因座使能高通量Menin蛋白质稳定性报告系统和 基于Dtag的Menin快速降解系统，(Ii)针对Menin蛋白的特异性CRISPR文库 功能和(Iii)针对嵌合体的早期候选小分子Menin蛋白分解(PROTAC)。这 提案还旨在提供高级转录方法方面的额外培训经验 分析，生化结合分析，小分子PROTAC设计和基于质谱学的共 免疫沉淀以及指导手稿/赠款的撰写和领导技能。锚定在 两位资源雄厚、经验丰富的导师的指导，一个出色的科学咨询委员会 由白血病和染色质生物学领域的专家组成，并进行了一项高度活跃的研究 在DCFI的环境，这项提议的资金将使我能够过渡到一名独立的调查员。

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.