Membrane Perturbations and Assay Interferences by Ivermectin Explain Its In Vitro SARS-CoV‑2 Antiviral Activities and Lack of Translatability

The antiparasitic drug ivermectin was proposed as a repurposed drug for the treatment of SARS-CoV-2 infection based on in vitro studies, but proved ineffective in high-quality clinical trials. When exploring possible reasons for this disconnect, we found that ivermectin interferes with AlphaScreen assays by quenching singlet oxygen transmission, calling into question the original justifications for pursuing ivermectin as an antiviral agent. Furthermore, at the low micromolar concentrations where ivermectin reduced SARS-CoV-2 viral burden in vitro, ivermectin decreased cell viability, modified membrane bilayer properties, and nonspecifically dysregulated membrane protein functions. In this Perspective, we provide molecular-level rationale for why ivermectin, an effective and safe antiparasitic drug at low nanomolar concentrations, becomes cytotoxic at low micromolar concentrations and, in turn, why ivermectin has not translated into an effective antiviral agent. We highlight lessons learned from the failed ivermectin repurposing effort and provide a workflow for identifying membrane-perturbing bioactivity early in drug development.基于体外实验结果，抗寄生虫药物伊维菌素（ivermectin）曾被提出作为治疗严重急性呼吸综合征冠状病毒2型（SARS-CoV-2）感染的老药新用候选药物，但在高质量临床试验中被证实无效。在探究这种结果不一致的潜在原因时，我们发现伊维菌素会通过淬灭单线态氧传递来干扰AlphaScreen检测，这使得此前将伊维菌素作为抗病毒药物开发的依据受到质疑。此外，在体外实验中伊维菌素可降低SARS-CoV-2病毒载量所需的低微摩尔浓度下，伊维菌素会降低细胞活力、改变细胞膜双层结构特性，并非特异性地导致膜蛋白功能异常。在本展望文章中，我们从分子层面阐释了为何在低纳摩尔浓度下兼具高效与安全性的抗寄生虫药物伊维菌素，在低微摩尔浓度下会产生细胞毒性，并进而说明为何伊维菌素未能成为有效的抗病毒药物。我们总结了伊维菌素老药新用项目失败的经验教训，并提供了一套可在药物开发早期识别膜扰动生物活性的工作流程。