Targeting MBNL1-mediated alternative splicing in MLL-fusion leukemia

提案摘要/摘要： 尽管在过去的几十年里，儿童急性白血病的预后有了显著改善，但婴儿 白血病是一种特别侵袭性的疾病，即使用化疗也难以治愈 强化和骨髓移植。大多数白血病携带MLL基因重排， 这一现象也见于一些新生急性髓性白血病（AML）以及治疗相关的AML， 研究表明，这些白血病的基础是一个共同的基因表达程序， 无论血统如何。通过初步研究，我们发现MLL融合白血病表达高， MBNL 1是一种调节可变RNA剪接的RNA结合蛋白。我们还发现了一个 MLL融合性白血病特有的选择性剪接（AS）特征，并显示MBNL 1缺失导致逆转 并损害MLL融合白血病细胞生长。因此，我们假设MBNL 1是一个关键的 MLL融合特异性AS程序调节子，以及通过MBNL 1抑制导致的该程序的破坏 到白血病细胞死亡。为了验证这些假设，我们的具体目标是：1）确定 潜在的MLL融合白血病对MBNL 1的依赖性，和2）优化小分子的特征 MBNL 1抑制剂作为MLL融合白血病的治疗。为了实现第一个目标，我们将描述变化 在关键的细胞生长和死亡途径，我们假设是负责的影响，看到遗传 击倒。我们还将使用CLIP-seq鉴定MLL融合白血病中新的MBNL 1-mRNA相互作用 （用RNAseq交联免疫沉淀）。对于第二个目标，我们已经证明了概念 MBNL 1的小分子抑制剂可以诱导MLL融合白血病细胞死亡。这种化合物需要 我们将通过应用药物化学原理来优化药效。而且我们 利用其结构特征和已发表的晶体结构， 专有化合物库。这一建议将促进我们对AS在 MLL融合白血病的发病机制，同时也直接导致这种疾病的一流疗法。 申请人目前是辛辛那提儿童医院肿瘤科的讲师 医学中心，将执行这一研究计划，同时从事结构化教学， 如申请中所述，从导师小组接受密切的个人指导。这些科学家 在白血病的分子发病机制方面拥有重要的专业知识。实验，指导， 本职业发展计划中描述的结构化课堂作业将使申请人成功地 过渡到一个独立的研究人员和医生，科学家，在RNA结合的作用的专业知识 蛋白质在白血病发病机制中的作用

Proposal Summary/Abstract: Though outcomes for pediatric acute leukemia have improved dramatically over the last few decades, infant leukemia is a particularly aggressive disease which remains difficult to cure even with chemotherapy intensification and bone marrow transplant. Most of these leukemias carry a rearrangement of the MLL gene, a phenomenon also seen in some de novo acute myeloid leukemias (AML) as well as therapy-related AML, and studies have demonstrated that a common gene expression program underlies these leukemias regardless of lineage. Through preliminary studies, we have found that MLL-fusion leukemias express high levels of MBNL1, an RNA binding protein that regulates alternative RNA splicing. We have also identified an alternative splicing (AS) signature unique to MLL-fusion leukemia, and show that MBNL1 loss causes reversion of this signature and impairs MLL-fusion leukemia cell growth. We thus hypothesize that MBNL1 is a critical regulator of an MLL-fusion specific AS program, and that disruption of this program via MBNL1 inhibition leads to leukemia cell death. To test these hypotheses, our specific aims are 1) to determine the mechanism underlying MLL-fusion leukemia dependence on MBNL1, and 2) optimize characteristics of a small-molecule MBNL1 inhibitor as a treatment for MLL-fusion leukemia. To achieve the first aim, we will characterize changes in key cell growth and death pathways which we hypothesize are responsible for the effects seen with genetic knockdown. We will also identify novel MBNL1-mRNA interactions in MLL-fusion leukemia using CLIP-seq (cross-linking immunoprecipitation with RNAseq). For the second aim, we have shown as a proof of concept that a small molecule inhibitor of MBNL1 can induce MLL-fusion leukemia cell death. This compound requires optimizations for potency, which we will achieve by applying medicinal chemistry principles. Furthermore, we have used its structural characteristics and published crystal structures to initiate an in silico screen of a proprietary compound library. This proposal will advance our understanding of the role of AS in the pathogenesis of MLL-fusion leukemia, while also directly leading to a first-in-class therapy for this disease. The applicant, who is currently an instructor in the Division of Oncology at Cincinnati Children’s Hospital Medical Center, will execute this research plan while simultaneously engaging in structured didactics and receiving close individual guidance from a panel of mentors as described in the application. These scientists possess significant expertise in the molecular pathogenesis of leukemia. The experiments, mentoring, and structured classwork described in this career development plan will position the applicant to successfully transition into an independent researcher and physician-scientist, with expertise in the role of RNA binding proteins in leukemia pathogenesis.