Hu Lei, Lv Jin-Feng, Zhuo Wei, Zhang Cong-Min, Zhou Hong-Hao, Fan Lan
Department of Clinical Pharmacology, Xiangya Hospital, Central South University, Changsha, China.
Institute of Clinical Pharmacology, Hunan Key Laboratory of Pharmacogenetics, Central South University, Changsha, China.
J Pharm Pharmacol. 2016 Sep;68(9):1193-202. doi: 10.1111/jphp.12591. Epub 2016 Jul 1.
All-trans-retinoic acid (ATRA), a naturally occurring metabolite of vitamin A, has been shown to have great potential as an antitumorigenic drug to treat acute leukaemia by promoting cancer cell differentiation. Cytochrome P450 oxidoreductase (POR) is the only obligate electron donor for all of the microsomal cytochrome P450 enzymes including CYP26A1 which is highly specific for ATRA metabolism and efficacy in human myeloid leukaemia cells. In this study, we aimed to investigate the effect of POR on ATRA efficacy and CYP26A1 expression in human myeloid leukaemia HL-60 cells.
Stably expressed POR and POR-RNAi HL-60 cell lines were established by transfecting POR overexpression or RNAi (RNA interference) vectors mediated by lentivirus. The protein expression of POR and CYP26A1 was examined by Western blot. The potential roles of POR on ATRA efficacy in HL-60 cells were explored by cell viability assay, cell cycle distribution, cellular differentiation and apoptosis analysis.
All-trans-retinoic acid treatment caused the expression of POR upregulation and CYP26A1 downregulation in dose- and time-dependent manners. POR overexpression decreased CYP26A1 expression in HL-60 cells. When POR gene was interfered, the downregulation of CYP26A1 expression by ATRA was abolished. In addition, POR overexpression in HL-60 cells significantly compromised ATRA-induced cell proliferation inhibition, cell cycle arrest, differentiation and apoptosis, whereas downregulation of POR significantly potentiated ATRA effects.
Our study therefore suggested that POR played an important role in regulating ATRA efficacy and CYP26A1 expression in HL-60 cells.
全反式维甲酸（ATRA）是维生素A的一种天然代谢产物，已显示出作为一种抗肿瘤药物通过促进癌细胞分化来治疗急性白血病的巨大潜力。细胞色素P450氧化还原酶（POR）是所有微粒体细胞色素P450酶的唯一必需电子供体，包括对人髓系白血病细胞中ATRA代谢和疗效具有高度特异性的CYP26A1。在本研究中，我们旨在探讨POR对人髓系白血病HL-60细胞中ATRA疗效和CYP26A1表达的影响。
通过转染慢病毒介导的POR过表达或RNAi（RNA干扰）载体，建立稳定表达POR和POR-RNAi的HL-60细胞系。通过蛋白质印迹法检测POR和CYP26A1的蛋白表达。通过细胞活力测定、细胞周期分布、细胞分化和凋亡分析，探讨POR对HL-60细胞中ATRA疗效的潜在作用。
全反式维甲酸处理以剂量和时间依赖性方式导致POR表达上调和CYP26A1表达下调。POR过表达降低了HL-60细胞中CYP26A1的表达。当POR基因受到干扰时，ATRA对CYP26A1表达的下调作用被消除。此外，HL-60细胞中POR过表达显著削弱了ATRA诱导的细胞增殖抑制、细胞周期阻滞、分化和凋亡，而POR的下调则显著增强了ATRA的作用。
因此，我们的研究表明POR在调节HL-60细胞中ATRA疗效和CYP26A1表达方面发挥着重要作用。
