Wang Bao-Zhong, Quan Fu-Shi, Kang Sang-Moo, Bozja Jadranka, Skountzou Ioanna, Compans Richard W
Department of Microbiology and Immunology and Emory Vaccine Center, Emory University School of Medicine, 1510 Clifton Road, Atlanta, Georgia 30322, USA.
J Virol. 2008 Dec;82(23):11813-23. doi: 10.1128/JVI.01076-08. Epub 2008 Sep 10.
We have designed a membrane-anchored form of the Toll-like receptor 5 ligand flagellin, the major proinflammatory determinant of enteropathogenic Salmonella, which was found to be glycosylated and expressed on cell surfaces. A chimeric influenza virus-like particle (cVLP) vaccine candidate containing A/PR8/34 (H(1)N(1)) hemagglutinin (HA), matrix protein (M1), and the modified flagellin as a molecular adjuvant was produced. The immunogenicity, including the serum antibody levels and cellular immune responses, and the protective efficacy against homologous and heterologous live virus challenge of the resulting VLPs were tested after intramuscular administration in a mouse model. The results demonstrated that flagellin-containing VLPs elicited higher specific immunoglobulin G (IgG) responses than standard HA and M1 VLPs, indicating the adjuvant effect of flagellin. Enhanced IgG2a and IgG2b but not IgG1 responses were observed with flagellin-containing VLPs, illuminating the activation of Th1 class immunity. The adjuvant effects of flagellin were also reflected by enhanced specific cellular responses revealed by the secretion of cytokines by freshly isolated splenocyte cultures when stimulated with pools of major histocompatibility complex class I or II peptides. When immunized mice were challenged with homologous live PR8 virus, complete protection was observed for both the standard and cVLP groups. However, when a heterosubtypic A/Philippines (H(3)N(2)) virus was used for challenge, all of the standard VLP group lost at least 25% of body weight, reaching the experimental endpoint. In contrast, for the cVLP group, 67% of mice survived the challenge infection. These results reveal that cVLPs designed by incorporating flagellin as a membrane-anchored adjuvant induce enhanced cross-protective heterosubtypic immune responses. They also indicate that such cVLP vaccines are a promising new approach for protection against pandemic influenza viruses.
我们设计了一种膜锚定形式的Toll样受体5配体鞭毛蛋白，它是肠道致病性沙门氏菌的主要促炎决定因素，已发现其被糖基化并在细胞表面表达。制备了一种嵌合流感病毒样颗粒（cVLP）候选疫苗，其包含A/PR8/34（H(1)N(1)）血凝素（HA）、基质蛋白（M1）以及作为分子佐剂的修饰鞭毛蛋白。在小鼠模型中进行肌肉注射后，测试了所得VLP的免疫原性，包括血清抗体水平和细胞免疫反应，以及对同源和异源活病毒攻击的保护效力。结果表明，含鞭毛蛋白的VLP比标准HA和M1 VLP引发更高的特异性免疫球蛋白G（IgG）反应，表明鞭毛蛋白具有佐剂作用。含鞭毛蛋白的VLP观察到IgG2a和IgG2b反应增强，但IgG1反应未增强，这表明Th1类免疫被激活。当用主要组织相容性复合体I类或II类肽库刺激时，新鲜分离的脾细胞培养物分泌细胞因子所显示的增强的特异性细胞反应也反映了鞭毛蛋白的佐剂作用。当用同源活PR8病毒攻击免疫小鼠时，标准组和cVLP组均观察到完全保护。然而，当用异亚型A/菲律宾（H(3)N(2)）病毒进行攻击时，所有标准VLP组小鼠体重至少减轻25%，达到实验终点。相比之下，对于cVLP组，67%的小鼠在攻击感染中存活。这些结果表明，通过掺入作为膜锚定佐剂的鞭毛蛋白设计的cVLP可诱导增强的交叉保护性异亚型免疫反应。它们还表明，此类cVLP疫苗是预防大流行性流感病毒的一种有前途的新方法。
