Center of Research Excellence on Therapeutic Proteins and Antibody Engineering, Department of Parasitology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, 10700, Thailand.
Department of Research and Development, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, 10700, Thailand.
Emerg Microbes Infect. 2018 Mar 21;7(1):41. doi: 10.1038/s41426-018-0031-3.
Small molecular inhibitors and passive immunization against Ebola virus disease (EVD) have been tested in animal models, including rodents and non-human primates, as well as in clinical trials. Nevertheless, there is currently no Food and Drug Administration (FDA)-approved therapy, and alternative strategies must be pursued. The aim of this study was to produce cell-penetrable human single-chain antibodies (transbodies) that are able to interfere with the activities of interferon inhibitory domain (IID) of the VP35 protein, a multifunctional virulence factor of Ebola virus (EBOV). We speculated that effective VP35-IID-specific transbodies could inspire further studies to identify an alternative to conventional antibody therapies. Phage display technology was used to generate Escherichia coli-derived human single-chain antibodies (HuscFvs) that bind to IID. HuscFvs were linked to nona-arginine (R9) to make them cell penetrable. Transbodies of transformed E. coli clones 13 and 3, which were predicted to interact with first basic patch residues (R9-HuscFv13), central basic patch, and end-cap residues (R9-HuscFv3), effectively inhibited EBOV minigenome activity. Transbodies of E. coli clones 3 and 8 antagonized VP35-mediated interferon suppression in VP35-transduced cells. We postulate that these transbodies formed an interface contact with the IID central basic patch, end-cap, and/or residues that are important for IID multimeric formation for dsRNA binding. These transbodies should be evaluated further in vitro using authentic EBOV and in vivo in animal models of EVD before their therapeutic/prophylactic effectiveness is clinically evaluated.
小分子抑制剂和针对埃博拉病毒病（EVD）的被动免疫已在动物模型中进行了测试，包括啮齿动物和非人类灵长类动物，以及临床试验。然而，目前尚无获得食品和药物管理局（FDA）批准的治疗方法，必须寻求替代策略。本研究旨在产生能够干扰埃博拉病毒（EBOV）多功能毒力因子 VP35 蛋白干扰素抑制结构域（IID）活性的细胞穿透性人单链抗体（transbodies）。我们推测，有效的 VP35-IID 特异性 transbodies 可以激发进一步的研究，以确定替代传统抗体疗法的方法。噬菌体展示技术用于生成与 IID 结合的大肠杆菌衍生的人单链抗体（HuscFvs）。将 HuscFvs 与非九聚精氨酸（R9）连接以使它们具有细胞穿透性。预测与第一碱性补丁残基（R9-HuscFv13）、中央碱性补丁和端帽残基（R9-HuscFv3）相互作用的大肠杆菌克隆 13 和 3 的 transbodies 有效地抑制了 EBOV 小基因活性。大肠杆菌克隆 3 和 8 的 transbodies 拮抗了 VP35 转导细胞中 VP35 介导的干扰素抑制。我们假设这些 transbodies 与 IID 中央碱性补丁、端帽和/或对于 IID 多聚体形成和 dsRNA 结合很重要的残基形成界面接触。这些 transbodies 应在使用真实 EBOV 的体外进一步评估，并在 EVD 的动物模型中进行体内评估，然后再在临床上评估其治疗/预防效果。
