目前普遍认为常规多药耐药（MDR）白血病/肿瘤细胞对砷剂敏感。我们采用阿霉素诱导获得一株具有高度三氧化二砷耐药性的白血病MDR细胞（HL-60/RS细胞）。本项目以砷剂敏感的经典MDR细胞K562/ADM细胞为参照，通过比较研究HL-60/RS细胞砷剂耐药特点、基因表达谱、细胞增殖-分化-死亡调控及药物代谢调控信号通路、砷转运蛋白、PML-RARA融合基因或PML基因突变，初步探讨常规MDR细胞对砷剂敏感抑或耐药的分子基础。比较研究HL-60/RS和K562/ADM细胞群体中白血病干细胞（LSCs）砷剂敏感性及其分子机制，明确LSCs在常规MDR细胞砷剂耐受抑或敏感中的作用，以及砷剂耐受MDR细胞是否存在三氧化二砷激活休眠LSC的缺陷。探索常规MDR细胞与砷剂交叉耐受的分子靶点和调控基础，为靶向预防和干预白血病/肿瘤治疗中含砷药物的耐药问题奠定基础。

The current ex vivo studies and clinic practice confirm that conventional multi-drug resistant(MDR) leukemia/tumor cells are sensitive to arsenicals, but we unexpectedly acquired a typical MDR cell line with high-resistance to arsenic trioxide by long-term and continuous stepwise adriamycin-selection of human leukemia HL-60 cells, named as HL-60/RS cells. Based on the uncovering of existence of unique arsenic-resistance in MDR leukemia cell population by our previous studies, the arsenic-sensitive classical MDR cells, K562/ADM cells, are employed as reference cells, this study aimed to comparatively investigate characteristics of arsenic-resistance, gene expression profiles, cellular proliferation-differentiation-death and drug metabolism related signal transduction pathways, arsenic transfers and mutations in PML-RARA fusion gene or PML gene, and to explore the preliminary molecular basis of sensitivity of conventional MDR cells to arsenic trioxide. The comparison of the arsenic sensitivity and molecular mechanism of leukemia stem cells(LSCs) in HL-60/RS and K562/ADM cells are done to reveal the action of LSCs in arsenic-sensitive or –resistance of conventional MDR cells, and whether exist imperfection in activation of arsenic-resistant LSC dormancy by arsenic trioxide. This study may figure out the molecular targets and regulation of conventional MDR leukemia cells cross-resistant to arsenic, and lay the foundation for guiding prevention and intervention of arsenic resistance in arsenic-based induction therapy of leukemias and tumors.

目前普遍认为常规多药耐药（MDR）白血病/肿瘤细胞对砷剂敏感。本项目采用阿霉素诱导成功建立具有典型多药耐药（MDR）特性和独特高三氧化二砷耐药性的白血病HL-60/RS耐药细胞，以砷剂敏感的经典MDR白血病K562/ADM细胞为参照，深入研究HL-60/RS细胞的生物学特性及白血病MDR细胞对三氧化二砷（ATO）的敏感性，通过比较研究HL-60/RS细胞砷剂耐药特点、基因表达谱、砷转运蛋白、PML-RARA融合基因、细胞增殖-分化-死亡调控及药物代谢调控信号通路，初步探讨常规MDR细胞对砷剂敏感抑或耐药的分子基础。研究结果证实HL-60/RS细胞具有MDR和砷耐受的双重特性，高表达ABC转运体和PML-RARA融合基因，具有更高含量的白血病干细胞（LSC）和自我更新能力。与K562/ADM细胞不同，HL-60/RS细胞对ATO和硫化砷具有很高的耐受性，其砷耐受性的机制不仅与砷转运体MPR1、MPR2和ASNA1等砷转运相关蛋白的表达水平密切相关，而且与ATO对这些转运蛋白表达的作用高度相关。源于白血病MDR细胞K562/ADM细胞的LSCK562/ADM对ATO诱发的细胞增殖抑制和凋亡敏感性高于来源于敏感K562细胞的LSCK562，与其相应群体细胞的砷敏感性一致。功能分类基因PCR芯片技术初步分析证实LSCK562/ADM细胞与LSCK562细胞在NF-？B、P53、PI3K/AKT、Wnt/β-catenin等信号通路存在较大的差异。白血病MDR细胞及其敏感细胞能量代谢特别是有氧糖酵解能力存在较大差异，常氧条件下白血病MDR细胞较敏感细胞具有更强的有氧糖酵解能力，ATO可通过AKT-c-MYC-mTOR途径抑制白血病MDR细胞有氧糖酵解、抑制其能量代谢，增高药物敏感性。应用二代测序（NGS）技术对HL-60/RS细胞和HL-60细胞及K562/ADM细胞和K562细胞两对细胞进行了高通量测序，初步发现两对细胞在细胞增殖、凋亡、自噬、药物转运、能量代谢等调控上存在很大的差异，白血病MDR是否对ATO耐受涉及多条信号通路，并有未知的新基因参与。综合上述研究结果，白血病MDR细胞是否对砷剂交叉耐受与PML-RARA融合基因表达、LSC比例、有氧糖酵解能力、ABC转运体特别是砷转运蛋白的表达，以及砷对砷转运蛋白及糖代谢的作用密切相关，NGS证实调控MDR细胞对砷敏感或耐