The Institute of Virology and Immunology IVI, Mittelhäusern, Switzerland.
Department of Infectious Diseases and Pathobiology, Vetsuisse Faculty, University of Bern, Bern, Switzerland.
Front Immunol. 2021 Jan 28;11:622385. doi: 10.3389/fimmu.2020.622385. eCollection 2020.
Self-amplifying replicon RNA (RepRNA) promotes expansion of mRNA templates encoding genes of interest through their replicative nature, thus providing increased antigen payloads. RepRNA derived from the non-cytopathogenic classical swine fever virus (CSFV) targets monocytes and dendritic cells (DCs), potentially promoting prolonged antigen expression in the DCs, contrasting with cytopathogenic RepRNA. We engineered pestivirus RepRNA constructs encoding influenza virus H5N1 (A/chicken/Yamaguchi/7/2004) nucleoprotein (Rep-NP) or hemagglutinin (Rep-HA). The inherent RNase-sensitivity of RepRNA had to be circumvented to ensure efficient delivery to DCs for intracellular release and RepRNA translation; we have reported how only particular synthetic delivery vehicle formulations are appropriate. The question remained concerning RepRNA packaged in virus replicon particles (VRPs); we have now compared an efficient polyethylenimine (PEI)-based formulation (polyplex) with VRP-delivery as well as naked RepRNA co-administered with the potent bis-(3',5')-cyclic dimeric adenosine monophosphate (c-di-AMP) adjuvant. All formulations contained a Rep-HA/Rep-NP mix, to assess the breadth of both humoral and cell-mediated defences against the influenza virus antigens. Assessment employed pigs for their close immunological relationship to humans, and as natural hosts for influenza virus. Animals receiving the VRPs, as well as PEI-delivered RepRNA, displayed strong humoral and cellular responses against both HA and NP, but with VRPs proving to be more efficacious. In contrast, naked RepRNA plus c-di-AMP could induce only low-level immune responses, in one out of five pigs. In conclusion, RepRNA encoding different influenza virus antigens are efficacious for inducing both humoral and cellular immune defences in pigs. Comparisons showed that packaging within VRP remains the most efficacious for delivery leading to induction of immune defences; however, this technology necessitates employment of expensive complementing cell cultures, and VRPs do not target human cells. Therefore, choosing the appropriate synthetic delivery vehicle still offers potential for rapid vaccine design, particularly in the context of the current coronavirus pandemic.
自我扩增复制子 RNA (RepRNA) 通过其复制特性促进编码感兴趣基因的 mRNA 模板的扩增，从而提供增加的抗原负荷。源自非致细胞病变性经典猪瘟病毒 (CSFV) 的 RepRNA 靶向单核细胞和树突状细胞 (DC)，可能促进 DC 中抗原的延长表达，与致细胞病变性 RepRNA 形成对比。我们设计了瘟病毒 RepRNA 构建体，编码流感病毒 H5N1（A/鸡/山口/2004）核蛋白 (Rep-NP) 或血凝素 (Rep-HA)。必须规避 RepRNA 的固有核糖核酸酶敏感性，以确保其有效地递送至 DC 以进行细胞内释放和 RepRNA 翻译；我们已经报告了只有特定的合成递送载体配方是合适的。RepRNA 包装在病毒复制子颗粒 (VRP) 中的问题仍然存在；我们现在比较了一种有效的聚乙二醇化 (PEI) 基配方 (多聚物) 与 VRP 递送以及与强效双-(3'，5')-环二核苷酸单磷酸 (c-di-AMP) 佐剂共同给药的裸露 RepRNA。所有配方均含有 Rep-HA/Rep-NP 混合物，以评估针对流感病毒抗原的体液和细胞介导防御的广度。评估使用猪作为其与人的密切免疫关系，以及作为流感病毒的天然宿主。接受 VRPs 以及 PEI 递送的 RepRNA 的动物表现出针对 HA 和 NP 的强烈体液和细胞反应，但 VRPs 更有效。相比之下，裸露的 RepRNA 加 c-di-AMP 只能在五头猪中的一头中诱导低水平的免疫反应。总之，编码不同流感病毒抗原的 RepRNA 可有效诱导猪的体液和细胞免疫防御。比较表明，在 VRP 内包装仍然是最有效的递送方法，可导致免疫防御的诱导；然而，这项技术需要使用昂贵的互补细胞培养物，并且 VRPs 不靶向人类细胞。因此，选择合适的合成递送载体仍然为快速疫苗设计提供了潜力，特别是在当前冠状病毒大流行的背景下。
