The role of Pleckstrin-2 as a functional node in myeloid proliferation

项目摘要 骨髓增生性肿瘤（MPN）是一组骨髓疾病， 产生血栓形成和发展为急性髓性白血病的风险增加。v617F JAK2的驱动突变是MPN的主要原因之一。这种突变的发现导致了 开发JAK抑制剂以治疗MPN。然而，JAK抑制剂不是治愈性的。此外，MPN 用JAK抑制剂治疗的患者通常由于JAK抑制剂的毒性而产生耐药性和严重的副作用。 JAK2在正常造血中的重要作用。我们一直在研究治疗MPN的新方法， 特别是关注JAK2通路的下游效应子。我们最近发表的研究 通过电流R01揭示了Pleckstrin-2（Plek2）的缺失，其是JAK2-STAT 5通路的新靶点， 改善JAK2V617F诱导的骨髓增殖表型，更重要的是逆转血管生成， 图10显示了JAK2V617F MPN小鼠模型的闭塞和致死率。鉴于Plek2在MPN中的重要性， 发病机制，我们已经确定了Plek2小分子抑制剂的先导化合物，使用基于硅的高， 通量筛选和基于细胞的测定。我们新的未发表的初步数据进一步揭示了Plek 2 Plek2与几种PI3K效应物结合，Plek2的缺失降低Akt活化。初步的体内证据也 表明Plek2对于PI3K-Akt通路是关键的，因为Plek2敲除改善了PI3K-Akt通路。 骨髓增殖和显着延长Pten造血特异性敲除小鼠的生存。这些 研究结果使我们假设Plek2作为一个中心枢纽连接JAK2-STAT和PI3K- Akt通路并促进骨髓增生。为了验证这一假设，并研究其机制， Plek2的功能，我们将使用1）体外生物化学和分子研究，以确定Plek2如何增强 PI3K-Akt信号通路; 2）Pten造血特异性敲除小鼠模型，以揭示Plek2的功能 体内，和3）临床前MPN模型和MPN患者样品，以确定Plek2抑制剂的功效。 该项目的成功完成将导致对Plek 2在糖尿病发病机制中的作用的新见解。 MPN，并为使用Plek2抑制剂作为单药或与 治疗MPN的其他化合物。

PROJECT SUMMARY Myeloproliferative neoplasms (MPNs) are a group of bone marrow diseases that show excessive myeloid cell production with an increased risk of developing thrombosis and evolving to acute myeloid leukemia. V617F driver mutation of JAK2 is one of the leading causes of MPNs. The discovery of this mutation led to the development of JAK inhibitors to treat MPNs. However, JAK inhibitors are not curative. In addition, MPN patients treated with JAK inhibitor often develop drug resistance and severe side effects due to the indispensable roles of JAK2 in normal hematopoiesis. We have been studying new approaches to treat MPNs, especially focusing on the downstream effectors of the JAK2 pathway. Our recently published studies funded by the current R01 revealed that loss of Pleckstrin-2 (Plek2), a novel target of the JAK2-STAT5 pathway, ameliorated JAK2V617F-induced myeloproliferative phenotypes, and more importantly reverted vascular occlusions and lethality of the JAK2V617F MPN mouse model. Given the significance of Plek2 in MPN pathogenesis, we have identified lead compounds of Plek2 small molecule inhibitors using in silico-based high- throughput screenings and cell-based assays. Our novel unpublished preliminary data further reveal that Plek2 binds to several PI3K effectors and loss of Plek2 reduces Akt activation. Preliminary in vivo evidence also demonstrates that Plek2 is critical for the PI3K-Akt pathway in that Plek2 knockout ameliorated myeloproliferation and significantly extended the survival of Pten hematopoietic specific knockout mice. These findings lead us to hypothesize that Plek2 functions as a central hub to connect the JAK2-STAT and the PI3K- Akt pathways and promote myeloproliferation. To test this hypothesis and investigate the mechanism of function of Plek2, we will use 1) in vitro biochemical and molecular studies to determine how Plek2 enhances the PI3K-Akt signaling, 2) Pten hematopoietic specific knockout mouse model to reveal the functions of Plek2 in vivo, and 3) pre-clinical MPN models and MPN patient samples to establish the efficacy of Plek2 inhibitors. Successful completion of this project will lead to novel insights into the role of Plek2 in the pathogenesis of MPNs and lay the foundation for clinical trials using Plek2 inhibitors as single agents or in combination with other compounds to treat MPNs.