Gammaretroviruses in Biologics

摘要：使用猪组织或细胞的临床试验正在进行中。由于所有猪的基因组中都携带猪内源性逆转录病毒（PERV），因此这些异种移植手术存在接触和传播PERV的风险。今年，我们分析了来自几种不同物种（恒河猴、狒狒和非洲绿猴）的非人类灵长类动物的原代细胞和细胞系是否允许PERV复制。我们进行了这项分析，以确定非人类灵长类动物是否是评估PERV体内传染性和潜在致病性的合适模型。在所有病例中，我们发现病毒可以进入细胞，整合并表达病毒RNA，但病毒感染没有在NHP培养物中传播。进一步的分析表明，允许复制的阻滞可能发生在几个不同的阶段，包括病毒进入效率低下和病毒组装（A. Ritzhaupt等，J. Virol）。，出版)。分析表明，NHP不太可能作为PERV在体内复制的许可模型，因此，来自此类研究的数据应谨慎解释。作为该项目的一个组成部分，我们开发了一种检测和定量PERV RNA和DNA的灵敏方法，可用于分析细胞培养实验、动物组织和人类临床样本（Argaw等人，J. Virol）。方法，出版中)。我们完成的第二个项目涉及对猪细胞衍生的PERV进行分析，这些细胞被改造成表达一种人类补体调节蛋白CD59 （Takefman, et al . J. Virol. 75:4551）。这项研究的目的是再现这种分子转基因猪身上可能发生的情况。有人提出，从猪中表达的PERV转基因人crp可能对补体介导的病毒溶解和中和活性具有抗性。我们发现人CD59蛋白被整合到来自表达hCD59的猪细胞的病毒粒子中，并且该分子的存在抑制补体介导的裂解，但不能中和PERV的传染性。因此，补体介导的抗PERV感染保护的一个组成部分仍然被保留。

Summary: Clinical trials using porcine tissues or cells are underway. Since all pigs carry in their genome porcine endogenous retroviruses (PERV), these xenotransplantation procedures carry the risk of exposure to and transmission of PERV. This year, we analyzed whether primary cells and cell lines derived from non-human primates of several different species (rhesus macaque, baboon, and African green monkey) were permissive to PERV replication. We performed this analysis to determine whether non-human primates are a suitable model for assessing in vivo infectivity and potential for pathogenicity of PERV. In all cases, we found that the virus could enter the cells, become integrated, and express viral RNA, but the viral infection did not spread in the NHP cultures. Further analyses demonstrated that the block to permissive replication may occur at several different stages, including inefficient viral entry and viral assembly (A. Ritzhaupt, et al, J. Virol., In Press). The analysis demonstrated that NHP do not likely serve as a permissive model for PERV replication in vivo, and therefore, data from such studies should be interpreted with caution. As a component of this project, we developed a sensitive assay for detection and quantitation of PERV RNA and DNA that can be used to analyze samples from cell culture experiments, animal tissues, and human clinical samples (Argaw, et al, J. Virol. Methods, In Press). The second project we completed involved an analysis of PERV derived from pig cells that were engineered to express one of the human complement regulatory proteins, CD59 (Takefman, et al, J. Virol. 75:4551). The aim of this study was to reproduce what would likely happen in pigs transgenic for this molecule. It has been proposed that PERV expressed from pigs that are transgenic for human CRPs may be resistant to the virolysis and neutralizing activity of complement-mediated protection. We found that the human CD59 protein is incorporated into virions derived from pig cells expressing hCD59, and that the presence of this molecule inhibited complement-mediated lysis, but not neutralization of infectivity of PERV. Therefore, one component of complement-mediated protection against PERV infection is still retained.