New Approaches To Passive And Active Immunoprophylaxis

HVS与比利时Rixensart的SmithKline Beecham合作开发了几种候选甲型肝炎减毒活疫苗。此外，人类免疫研究所还开发了一种前景看好的候选重组戊型肝炎疫苗，目前正在进行临床试验。HVS正在研究DNA疫苗的技术，并建立了一个基于乙肝疫苗的模型系统，HVS在乙肝疫苗方面已经有了丰富的经验。我们测试了一种免疫刺激剂(CpG)作为DNA疫苗和蛋白质疫苗佐剂的有效性。此外，还探讨了DNA疫苗在丙型肝炎病毒控制中的应用。一种基于丙型肝炎病毒E2包膜蛋白的DNA疫苗在小鼠和猕猴中被证明具有高度的免疫原性，在黑猩猩中具有中等的免疫原性，但当黑猩猩受到活的丙型肝炎病毒攻击时，它们并没有得到完全保护。一种类似的方法已经用于基于丙型肝炎病毒包膜蛋白的DNA疫苗的研究。不同结构的E1基因被制备成DNA疫苗(表达载体质粒)和载体疫苗(重组痘苗)。用DNA疫苗(“PRIME”)接种小鼠，然后用重组牛痘病毒(“Boost”)接种。这些小鼠对DNA疫苗和牛痘疫苗都有很好的免疫反应。基于这些结果，研究将扩展到包括黑猩猩在内的非人类灵长类动物，以确定这种免疫反应是否具有保护性。被动免疫预防也是一种重要的公共卫生工具。例如，正常的免疫球蛋白在预防甲型肝炎方面一直很重要。然而，单抗制剂可能更有效，为特定的中和表位量身定做，效力高度一致。我们从实验中先后感染了五种人类肝炎病毒的黑猩猩的骨髓中制备了组合文库。黑猩猩的球蛋白与人类的免疫球蛋白几乎完全相同，这使它们成为免疫预防和免疫治疗药物的诱人选择。到目前为止，我们已经分离出与甲型肝炎病毒、乙肝病毒、丁型肝炎病毒和戊型肝炎病毒发生反应的单抗免疫球蛋白。在其他研究中，我们已经恢复了与丙型肝炎病毒反应的人类单抗。上面描述的许多单抗是中和的，它们的生产正在扩大规模，用于对黑猩猩进行被动免疫预防测试。针对被实验感染登革热病毒1至4和诺沃克病毒的黑猩猩，正在进行类似的骨髓组合文库构建。诺沃克病毒是成人病毒性胃肠炎的最常见原因之一。在某些情况下(登革热)，重组单抗可能被证明是重要的预防和治疗药物。在其他情况下(诺沃克病毒)，单抗将用作诊断试剂。我们计划将这些研究扩展到其他感兴趣的病毒，这些病毒可以在黑猩猩身上进行实验。

The HVS in collaboration with SmithKline Beecham, Rixensart, Belgium, has developed several candidate live attenuated HAV vaccines. In addition, the HVS has developed a candidate recombinant hepatitis E vaccine that is highly promising and that is currently in clinical trials. The HVS is studying the technology of DNA vaccines with a model system based upon hepatitis B virus (HBV) vaccine, a vaccine with which the HVS has had extensive experience. We have tested the efficacy of an immunostimulant (CpG) as an adjuvant for DNA vaccines, as well as for protein vaccines. In addition, the utility of DNA vaccines for the control of hepatitis C virus (HCV) has been explored. A DNA vaccine based on the E2 envelope protein of HCV proved to be highly immunogenic in mice and rhesus monkeys and moderately immunogenic in chimpanzees, but the chimpanzees were not fully protected when they were challenged with live HCV. A similar approach has been utilized in the study of a DNA vaccine based on the E1 envelope protein of HCV. Various constructs of the E1 gene were prepared as DNA vaccines (expression vector plasmids) and as vectored vaccines (recombinant vaccinia). Mice were vaccinated with the DNA vaccine ("prime"), followed by vaccination with the recombinant vaccinia viruses ("boost"). The mice had excellent immune responses to the DNA vaccine as well as to the vaccinia boost. Based on these results, the studies will be extended to non-human primates, including chimpanzees, to determine if such immune responses are protective. Passive immunoprophylaxis has also been an important public health tool. For example, normal immunoglobulin has been important in the prevention of hepatitis A. However, monoclonal preparations could be more potent, tailored to specific neutralization epitopes and highly consistent in potency. We have prepared combinatorial libraries from the bone marrow of chimpanzees that had been experimentally infected in sequence with each of the five human hepatitis viruses. Chimpanzee globulins are virtually identical to human immunoglobulins, making them attractive choices for immunoprophylactic and immunotherapeutic agents. To date, we have isolated monoclonal immunoglobulins that react with HAV, HBV, HDV and HEV. In other studies, we have recovered human monoclonal antibodies that react with HCV. Many of the monoclonal antibodies described above are neutralizing and their production is being scaled up for tests of passive immunoprophylaxis in chimpanzees. Similar construction of combinatorial libraries from bone marrow is being carried out for chimpanzees that have been experimentally infected with dengue viruses 1 through 4 and the Norwalk virus, one of the most common causes of adult viral gastroenteritis. In some cases (dengue fever) recombinant monoclonal antibodies may prove to be important prophylactic and therapeutic agents. In other cases (Norwalk virus), the monoclonal antibodies will be useful as diagnostic reagents. We plan to extend these studies to other viruses of interest that can be experimentally administered to chimpanzees.