GAS5基因多态性及其表达对急性髓系白血病阿糖胞苷联合蒽环类化疗预后的影响及机制探讨

cytosine arabinoside (Ara-c) combined with anthracycline-based chemotherapy is the standard chemotherapy treatment for acute-myeloid leukemia (AML). However, both chemosensitivity and prognosis AML patients show remarkable individual differences. Genetic factors are acknowledged to be one of the main causes for this individual differences. Our previous study have observed that the long non-conding RNA GAS5 promoter single nucleotide polymorphism rs55829688 was significantly associated with AML prognosis as indicated by overall survival (OS). Moreover, this polymorphism could alter the promoter activity and the expression of GAS5 through intervening the action of transcript factor TP63. Study has shown that GAS5 can interfere with the actions glucocorticoids (GCs) through binding activated GC receptors, and in consideration of the important role of GCs in stimulating bone marrow hematopoiesis, while myelosupression is the most important cause of death in AML patients during chemotherapy, this program is designed to make clear the influence of GAS5 rs55829688 polymorphism on AML prognosis is related to inhibition in recovery of myelosupression after chemotherapy by both clinical and cellular studies. By isolating and culture of hemopoietic stem cell (HSCs) from human umbilical cord blood and using Ara-c induced HSCs damage, through interfering with GAS5 expression by overexpression and siRNA, to explore the role of GAS5 in hematopoiesis under Ara-c exposure and the possible mechanisms.

阿糖胞苷联合蒽环类药物治疗是急性髓系白血病（AML）标准化疗方案，但患者的疗效和和预后不一，遗传因素是AML药物反应及预后差异的主要原因之一。本项目前期研究发现lncRNA GAS5 rs55829688多态性与AML患者生存期相关，其功能可能与影响TP63对GAS5启动子的调节从而升高其表达有关。由于GAS5可抑制糖皮质激素受体的功能，糖皮质激素具有促进造血的作用，而骨髓抑制是AML的主要死因，本研究拟通过临床研究结合细胞实验，查明GAS5 rs55829688多态性影响AML的预后是否与其影响联合化疗后骨髓抑制的恢复有关；并通过分离培养人脐带血造血干细胞（HSCs），在阿糖胞苷诱导损伤的HSCs细胞模型，通过干预GAS5表达和质谱分析，查明GAS5在HSCs造血中的作用及其分子机制。本项目旨在探讨GAS5影响AML预后的机制，为促进AML的个体化治疗和骨髓抑制的恢复提供新的思路。

本项目从药物基因组学角度出发，利用基础分子生物学和临床相结合的手段，查明了GAS5 rs55829688多态性与AML患者的预后的关系并进行初步机制探索，并从多角度发现与AML阿糖胞苷联合蒽环类药物化疗疗效、不良反应及预后相关的药物基因组生物标志物，初步建立了AML化疗敏感性及预后预测模型。项目发现rs55829688 CC型患者的血小板恢复时间更长，而GAS5过表达后造血干细胞（HSPC）增殖减慢、凋亡增加、造血集落的形成减少，该过程可能与GATA1、SPI1等造血相关转录因子的表达水平有关。项目还从药物代谢和AML发病机制相关基因出发进行系统的药物基因组学研究，发现DCK rs4643786、NME2 rs3744660、RRM1 rs183484、UGT1A1*28和*6、E2F1 rs3213150等多个多态性位点与AML诱导化疗的反应性和预后相关。项目纳入与Ara-C代谢相关的8个SNP，并分别通过Logistic回归分析、Fisher判别分析和决策树构建AML药物反应性模型并发现模型的效果较好，该发现为AML的个体化治疗和预后预测奠定了理论基础。项目还基于甲基化芯片分析发现TBC1D16 cg16030878甲基化水平与TBC1D16 mRNA表达水平呈正相关，且TBC1D16 低表达患者的OS显著延长；TBC1D16敲减导致细胞出现轻微的G1/S期细胞周期阻滞、生长速度减慢，期机制可能与p-ERK表达下调有关。项目还应用公共数据库，基于Ara-C敏感性相关基因mRNA表达水平构建了基于42个与阿糖胞苷反应性和AML预后基因的预测模型，为从全基因组水平探讨AML化疗抵抗和预后个体差异机制、构建个体化用药与预后预测模型提供了范例。