DEVELOPMENT OF A DETECTIVE HEPATITIS A VIRUS VACCINE

到目前为止，生产甲型肝炎疫苗的尝试尚未成功，原因是 几个因素，包括组织培养中病毒生长不良， 没有体外衰减的标志物，以及分离出的 病毒蛋白不显示病毒中和表位。作为一种 经典灭活病毒或减毒病毒疫苗的替代品 重组DNA衍生亚单位疫苗的研究方法--缺陷甲型肝炎病毒 这种方法将作为甲型肝炎疫苗开发和测试。一种新的甲型肝炎病毒株 会引起裂解性感染，并在组织培养中生长到高滴度 被利用。在第一阶段，这种快速增长的甲型肝炎病毒变种的基因组将是 并对分离克隆的生物活性进行评估。 通过将其导入易感的人肝癌细胞。在第二阶段， 缺失或替换突变将被引入到 产生缺陷甲型肝炎病毒的基因组。互补性细胞系 将构建表达2C基因的基因并用于生长缺陷 病毒。一旦有缺陷的病毒和补体细胞系 构建，研究以评估突变病毒和 将进行病毒生产水平的测定，以确定 将该病毒用作疫苗的可行性。

Attempts to generate HAV vaccines have been unsuccessful to date due to several factors including poor growth of the virus in tissue culture, absence of in vitro markers of attenuation, and the fact that isolated viral proteins do not display virus neutralizing epitopes. As an alternative to classical killed virus or a attenuated virus vaccine approaches or recombinant DNA derived subunit vaccines, a defective HAV approach will be developed and tested as a HAV vaccine. A new HAV strain that causes lytic infection and grows to high titer in tissue culture will be used. In Phase I, the genome of this fast growing HAV variant will be cloned and the biological activity of the isolated clone will be assessed by transfection into susceptible human hepatoma cells. In Phase II, deletion or substitution mutations will be introduced into the 2C region of the genome to generate a defective HAV. A complementing cell line expressing the 2C gene will be constructed and used to grow the defective virus. Once the defective virus and complementing cell line are constructed, studies to assess the stability of the mutant virus and the production level of the virus will be done in order to determine the feasibility of using the virus as a vaccine.