FADD在阿西替尼诱导巨核细胞白血病细胞系多倍体化和凋亡过程中作用机制的研究

Polyploidization is crtical for development of megakaryocytes. Failure of polyploidization leads to acute megakaryocytic leukemia (AMKL). Although many Down syndrome AMKL (DS-AMKL) patients respond to current therapies, adults with non-DS-AMKL have a very poor prognosis, with the vast majority relapsing within 1 year of the primary treatment. Recently, induction of polyploidization in AMKL cells is emerging as a novel strategy in development of therapeutic regimen of AMKL. Given that the apoptosis pathways exist in megakaryocytes，it is reasonable that induction of both polyploidization and apoptosis will be more helpful in therapy of AMKL than induction of polyploidization or apoptosis alone. Our preliminary investigation revealed that axitinib can potentially induce both polyploidization and apoptosis in AMKL cell lines. These effects of axitinib on AMKL cell lines may be realted to FADD (FAS-associated death domain protein), S6K1 (ribosomal S6 kinase 1) and AURKA (Aurora-A), and furthermore FADD and its phosphorylation may be involved in regulation of the balance between polyploidization and apoptosis in AMKL cell lines induced by axitinib. Therefore, in this investigating project, we will further explore the roles of FADD, the components involving the assembly of death- and life- inducing signaling complex, and the kinases and peptidyl-prolyl cis-trans isomerase regulating the phosphorylation of FADD in polyploidization and apoptosis of AMKL cell lines treated with axitinib. We will also explore the mutual interaction among FADD, S6K1 and AURKA. The findings from this project will provide experimental data for clinical translation of axitinib, a rationale to support clinical trials of aixitinb, and new target for further drug development.

多倍体化对巨核细胞发育至关重要，其失败将导致急性巨核细胞白血病（AMKL）。虽然许多唐氏综合症AMKL(DS-AMKL)患者对现有治疗有效，但成人非DS-AMKL患者预后极差，绝大部分患者首次治疗1年后复发。最近，诱导AMKL细胞多倍体化已成为研究治疗AMKL的新策略。鉴于巨核细胞内存在凋亡途径，如能同时诱导多倍体化和凋亡，可能更有利于AMKL的治疗。我们初步的研究发现阿西替尼有这方面潜质，其作用可能与FADD，S6K1和AURKA有关，而且FADD和其磷酸化修饰可能调节凋亡和多倍体化间的平衡。因此，本项目将深入研究FADD，与其组成介导死亡和存活信号复合物的其他组分和参与其磷酸化修饰激酶和异构酶，在阿西替尼诱导多倍体化和凋亡过程中的作用，以及FADD，S6K1和AURKA相互作用的关系。研究成果，将为基于诱导多倍体化和凋亡AMKL治疗策略，阿西替尼的临床转化，提供实验数据和理论依据。

本研究发现，（1）低分化CMK11-5细胞系、中度分化HEL细胞系和高度分化Dami细胞系可以作为巨核细胞不同发育阶段的研究模型；（2）在巨核细胞的发育早期诱导分化和成熟，并抑制凋亡，有利于诱导多倍体化；(3) 在巨核细胞的发育早和中期，成熟与多倍体化偶联，而在发育的晚期二者解偶联；（4）Ly2874455是一个与目前已经完成I期临床试验，有望用于巨核细胞白血病和原发性骨髓纤维化治疗药物（Alisertib）体外生物学活性基本相同，但作用机制不同的小分子化合物；（5）Ly2874455的作用靶点可能是PIM-1。鉴于目前尚不能获得大量原代和体外培养的同步化的巨核细胞，三个细胞模型对于研究巨核细胞发育过程中分化、多倍体化、成熟和凋亡间的关系以及相关分子和信号转导机制，将有重要的推动作用；该化合物有望申报治疗巨核细胞相关疾病的专利。本项研究不但有重要的理论意义，也有潜在的临床转化价值。