基于OCT2/MATEs机制的延胡索乙素降低顺铂肾毒性研究

Cisplatin is one of the most widely used and most potent chemotherapy drugs, however severe nephrotoxicity limits its clinical application. Recent research has shed significant new lights on the mechanism of cisplatin nephrotoxicity, especially on renal organic transporters, such as OCT2, MATE1 and MATE2-K, which have been linked to cisplatin-induced nephrotoxicity. Previously, the renal toxicity of cisplatin was suggested to be a result of OCT2-mediated extensive renal accumulation from the circulation, and weak tubular secretion into the urine by multidrug and toxin extrusion protein 1 and 2 (MATE1 and MATE2-K), primarily located at the apical membrane of proximal tubular cells. Currently, it has not yet been found an effective drug in clinic, which has the potential to prevent cisplatin-induced nephrotoxicity in patients, mainly based on the effect of OCT2- or MATEs-mediated renal transport of cisplatin. Our previous work demonstrated that (-)-THP ((-)-tetrahydropalmatine) had significant inhibitory effect on OCT2, and protected against severe cisplatin-induced cytoxicity on MDCK-hOCT2 and MDCK-hOCT2/hMATE1 cells in a concentration-dependent manner. Therefore, we proposed that (-)-THP co-administrated with cisplatin, might have an effect on cisplatin accumulation via OCT2 and/or MATEs, then reduces cisplatin-induced nephrotoxicity. To confirm the hypothesis, the models of the transgenic cells, renal primary cells, renal cortical slices and Oct1/2 (-/-) mouse would be used to investigate the molecular mechanism in the protective effect of (-)-THP on the cisplatin-induced nephrotoxicity based on the renal transport of cisplatin mediated by OCT2, MATE1 and MATE2-K. If the results as our expect，the concomitant administration of (-)-THP was revealed to be a useful strategy preventing cisplatin-induced nephrotoxicity，meanwhile (-)-THP could ameliorate cancer pain in patients. The results offer the scientific basis for the future clinical (-)-THP as a specific modifier of cisplatin-induced nephrotoxicity.

顺铂是目前临床上最有效的广谱抗肿瘤药物之一，但严重的肾毒性限制了其在临床上的使用。肾小管上皮细胞基底膜侧高表达的OCT2将顺铂摄取入细胞，使其毒性增强；而顶膜侧MATE1和MATE2-K介导顺铂泵出细胞，降低其毒性。目前，尚未发现安全有效的临床药物，通过影响OCT2/MATEs介导顺铂肾脏转运，达到抑制或降低顺铂肾毒性的效果。申请者研究发现，左旋延胡索乙素 ((-)-THP) 对hOCT2具有显著抑制作用，能抑制顺铂在MDCK-hOCT2与MDCK-hOCT2/hMATE1细胞上的毒性。提出(-)-THP可能通过对OCT2/ MATEs介导顺铂跨膜转运的影响，减少顺铂肾脏蓄积，从而达到减弱或抑制顺铂肾毒性的预期结果。为此，本项目拟运用细胞、组织、基因敲除小鼠等模型，阐明基于OCT2/MATEs 介导的(-)-THP降低顺铂肾毒性机制，为临床(-)-THP用于降低顺铂肾毒性提供科学依据。

顺铂是目前临床上最有效的广谱抗肿瘤药物之一，广泛用于卵巢癌、前列腺癌、肺癌及乳腺癌等多种实体肿瘤治疗，但其严重的肾毒性限制了其临床应用。L001是一种生物碱，具有缓解头痛、胸痛等多种疼痛的作用，且具有非成瘾性、毒副作用小的优点。那么，L001是否会通过影响肾脏基底膜侧特异性高表达的有机阳离子转运体2（organic cation transporter, OCT2）及顶膜侧多药及毒素外排蛋白(multidrug and toxin extrusion proteins, MATEs)而拮抗顺铂引起的肾脏细胞毒性。研究发现，在MDCK-hOCT2 /hMATE1 单层细胞上，L001通过选择性强抑制肾脏细胞膜转运体OCT2对顺铂的摄取，对MATE1外排顺铂的较弱抑制，使得顺铂的细胞积聚降低，并由此减弱顺铂的细胞毒性。L001的细胞保护作用在原代培养的小鼠肾细胞中得到了进一步的验证。相似的，以健康小鼠为动物模型，顺铂合用 5-40 mg/kg L001能显著降低顺铂的肾脏积聚为单独使用顺铂的75.0%-52.3%，同时，显著缓解顺铂的肾脏毒性。另外，L001在H460 和HCT116荷瘤裸鼠体内在保护肾脏免受损伤的同时，并不会因此而降低顺铂抗肿瘤的效应。在H460荷瘤裸鼠体内，10, 20 和 40 mg/kg L001分别与10 mg/kg顺铂合用，肿瘤抑制率分别达到71.4%, 70.4% 和69.4%，高于单独使用10 mg/kg顺铂(60.6%)。而在HCT116荷瘤裸鼠体内， 20 mg/kg L001与10 mg/kg顺铂合用，肿瘤抑制率达到34.5%，也显著高于单独使用10 mg/kg顺铂（25.4%）。L001通过减少肾脏细胞对顺铂的摄取从而减缓顺铂的细胞毒性在原代培养的人肾脏细胞也得到了较好的验证。因此，L001通过抑制顺铂进入肾脏细胞减弱顺铂的肾脏毒性，但并未影响顺铂的抗肿瘤效应，为临床铂类药物肾毒性的防护提供新途径。