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Abstract：
Anticancer therapeutics can provoke severe side effects that impair the patient\'s quality of life. A frequent dose-limiting side effect of platinum-based anticancer therapy is neurotoxicity. Its pathophysiology is poorly understood, and effective preventive or therapeutic measures are missing. Therefore, elucidation of the molecular mechanism of platinating drug-induced neurotoxicity and the development of preventive strategies is urgently needed. To this end, we aim to use C. elegans as a 3R-compliant in vivo model. The 3R principles were conceived for animal welfare in science concerning animal experiments, which should be replaced, reduced or refined. We can analytically demonstrate dose-dependent uptake of cisplatin (CisPt) in C. elegans, as well as genotoxic and cytotoxic effects based on DNA adduct formation (i.e., 1,2-GpG intrastrand crosslinks), induction of apoptosis, and developmental toxicity. Measuring the impairment of pharyngeal pumping as a marker of neurotoxicity, we found that especially CisPt reduces the pumping frequency at concentrations where basal and touch-provoked movement were not yet affected. CisPt causes glutathione (GSH) depletion and RNAi-mediated knockdown of the glutamate-cysteine ligase GCS-1 aggravates the CisPt-induced inhibition of pharyngeal pumping. Moreover, N-acetylcysteine (NAC) mitigated CisPt-triggered toxicity, indicating that GSH depletion contributes to the CisPt-induced pharyngeal damage. In addition to NAC, amifostine (WR1065) also protected the pharynx of C. elegans from the toxic effects of CisPt. Measuring pharyngeal activity by the electrophysiological recording of neurotransmission in the pharynx, we confirmed that CisPt is neurotoxic in C. elegans and that NAC is neuroprotective in the nematode. The data support the hypothesis that monitoring the pharyngeal activity of C. elegans is a useful surrogate marker of CisPt-induced neurotoxicity. In addition, a low GSH pool reduces the resistance of neurons to CisPt treatment, and both NAC and WR1065 are capable of attenuating platinum-induced neurotoxicity during post-incubation in C. elegans. Overall, we propose C. elegans as a 3R-compliant in vivo model to study the molecular mechanisms of platinum-induced neurotoxicity and to explore novel neuroprotective therapeutic strategies to alleviate respective side effects of platinum-based cancer therapy.
摘要：
抗癌疗法可以引起严重的副作用，损害患者的生活质量。基于铂的抗癌疗法的常见剂量限制性副作用是神经毒性。它的病理生理学知之甚少，缺乏有效的预防或治疗措施。因此,迫切需要阐明铂类药物诱导的神经毒性的分子机制并制定预防策略.为此,我们的目标是使用秀丽隐杆线虫作为3R兼容的体内模型。3R原则是为动物实验科学中的动物福利而构想的，应该更换，减少或精炼。我们可以通过分析证明秀丽隐杆线虫中顺铂（CisPt）的剂量依赖性摄取，以及基于DNA加合物形成的基因毒性和细胞毒性作用(即，1,2-GpG链内交联)，诱导凋亡，和发育毒性。测量咽泵的损伤作为神经毒性的标志，我们发现,在尚未影响基础运动和触感运动的浓度下,尤其是CisPt会降低泵浦频率.CisPt导致谷胱甘肽（GSH）耗竭，RNAi介导的谷氨酸-半胱氨酸连接酶GCS-1的敲低加重了CisPt诱导的咽泵的抑制。此外，N-乙酰半胱氨酸（NAC）减轻CisPt触发的毒性，表明GSH耗竭有助于CisPt引起的咽部损伤。除了NAC,氨磷汀(WR1065)还保护秀丽隐杆线虫的咽部免受CisPt的毒性作用。通过咽部神经传递的电生理记录来测量咽部活动，我们证实CisPt对秀丽隐杆线虫具有神经毒性,而NAC对线虫具有神经保护作用。数据支持以下假设：监测秀丽隐杆线虫的咽部活性是CisPt诱导的神经毒性的有用替代标记。此外,低GSH池降低了神经元对CisPt治疗的抵抗力，NAC和WR1065都能够减弱在秀丽隐杆线虫中孵育后的铂诱导的神经毒性。总的来说,我们建议将秀丽隐杆线虫作为一种符合3R标准的体内模型来研究铂诱导的神经毒性的分子机制,并探索新的神经保护性治疗策略,以减轻铂类癌症治疗的副作用.
