Tran Pham-Tue-Hung, Asghar Naveed, Höglund Urban, Larsson Olivia, Haag Lars, Mirazimi Ali, Johansson Magnus, Melik Wessam
School of Medical Science, Inflammatory Response and Infection Susceptibility Centre (iRiSC), Örebro University, 703 62 Örebro, Sweden.
Adlego Biomedical AB, P.O. Box 42, 751 03 Uppsala, Sweden.
Microorganisms. 2020 Nov 29;8(12):1890. doi: 10.3390/microorganisms8121890.
Kunjin virus (KUNV) is an attenuated strain of the severe neurotropic West Nile virus (WNV). The virus has a single-strand positive-sense RNA genome that encodes a polyprotein. Following gene expression, the polyprotein is cleaved into structural proteins for viral packaging and nonstructural proteins for viral replication and expression. Removal of the structural genes generate subgenomic replicons that maintain replication capacity. Co-expression of these replicons with the viral structural genes produces reporter virus-like particles (RVPs) which infect cells in a single round. In this study, we aimed to develop a system to generate multivalent RVPs based on KUNV to elicit an immune response against different viruses. We selected the Ebola virus (EBOV) glycoprotein (GP) and the matrix protein (VP40) genes, as candidates to be delivered by KUNV RVPs. Initially, we enhanced the production of KUNV RVPs by generating a stable cell line expressing the KUNV packaging system comprising capsid, precursor membrane, and envelope. Transfection of the DNA-based KUNV replicon into this cell line resulted in an enhanced RVP production. The replicon was expressed in the stable cell line to produce the RVPs that allowed the delivery of EBOV GP and VP40 genes into other cells. Finally, we immunized BALB/cN mice with RVPs, resulting in seroconversion for EBOV GP, EBOV VP40, WNV nonstructural protein 1, and WNV E protein. Thus, our study shows that KUNV RVPs may function as a WNV vaccine candidate and RVPs can be used as a gene delivery system in the development of future EBOV vaccines.
库宁病毒（KUNV）是严重嗜神经性西尼罗河病毒（WNV）的减毒株。该病毒具有单链正义RNA基因组，编码一种多聚蛋白。基因表达后，多聚蛋白被切割成用于病毒包装的结构蛋白和用于病毒复制与表达的非结构蛋白。去除结构基因可产生维持复制能力的亚基因组复制子。这些复制子与病毒结构基因共表达可产生报告病毒样颗粒（RVP），其可单次感染细胞。在本研究中，我们旨在开发一种基于KUNV产生多价RVP的系统，以引发针对不同病毒的免疫反应。我们选择了埃博拉病毒（EBOV）糖蛋白（GP）和基质蛋白（VP40）基因，作为由KUNV RVP递送的候选基因。最初，我们通过构建表达包含衣壳、前体膜和包膜的KUNV包装系统的稳定细胞系来提高KUNV RVP的产量。将基于DNA的KUNV复制子转染到该细胞系中可提高RVP产量。复制子在稳定细胞系中表达以产生RVP，从而将EBOV GP和VP40基因递送至其他细胞。最后，我们用RVP免疫BALB/cN小鼠，导致针对EBOV GP、EBOV VP40、WNV非结构蛋白1和WNV E蛋白的血清转化。因此，我们的研究表明，KUNV RVP可能作为WNV疫苗候选物，并且RVP可在未来EBOV疫苗的开发中用作基因递送系统。
