Chu T H, Dornburg R
Department of Molecular Genetics and Microbiology, Robert Wood Johnson Medical School, University of Medicine and Dentistry of New Jersey, Piscataway 08854, USA.
J Virol. 1997 Jan;71(1):720-5. doi: 10.1128/JVI.71.1.720-725.1997.
Recently, we constructed retroviral vector particles derived from spleen necrosis virus (SNV) that display a single-chain antibody (scA) on the viral surface. By transient transfection protocols, we showed that such particles are competent for infection and cell type specific. Efficient infection was dependent on the presence of wild-type envelope, although wild-type SNV was not infectious on target cells (T.-H. T. Chu and R. Dornburg, J. Virol. 69:2659-2663, 1995; T.-H. T. Chu, I. Martinez, W. C. Sheay, and R. Dornburg, Gene Ther. 1:292-299, 1994). In this study, stable packaging lines were constructed and detailed biological and biochemical studies were performed. Chimeric scA-envelope fusion proteins were expressed as efficiently as wild-type envelope and were stable over a period of at least 6 h. Only a fully functional wild-type envelope could act as a helper for efficient virus penetration. The ratio of wild-type envelope protein to chimeric envelope protein appears to determine the efficiency of infection. Virus titers of targeting vectors obtained from stable packaging lines were as high as 10(4) CFU/ml. A 25-fold concentration of vector virus stocks resulted in a 200-fold increase in virus titers (up to 10(6) CFU/ml). These data indicate that an inhibitor of infection was (at least partially) removed by the concentration protocol. Our data show that this technology has several variables for further improvements and, therefore, has the potential to become a powerful tool for cell-type-specific in vivo human gene therapy.
最近，我们构建了源自脾坏死病毒（SNV）的逆转录病毒载体颗粒，这些颗粒在病毒表面展示单链抗体（scA）。通过瞬时转染方案，我们表明此类颗粒具有感染能力且具有细胞类型特异性。高效感染依赖于野生型包膜的存在，尽管野生型SNV对靶细胞无感染性（T.-H. T. Chu和R. Dornburg，《病毒学杂志》69:2659 - 2663，1995；T.-H. T. Chu、I. Martinez、W. C. Sheay和R. Dornburg，《基因治疗》1:292 - 299，1994）。在本研究中，构建了稳定的包装细胞系并进行了详细的生物学和生物化学研究。嵌合scA - 包膜融合蛋白的表达效率与野生型包膜一样高，并且在至少6小时内保持稳定。只有完全功能的野生型包膜才能作为有效病毒穿透的辅助因子。野生型包膜蛋白与嵌合包膜蛋白的比例似乎决定了感染效率。从稳定包装细胞系获得的靶向载体的病毒滴度高达10⁴CFU/ml。载体病毒原液25倍的浓缩导致病毒滴度增加200倍（高达10⁶CFU/ml）。这些数据表明感染抑制剂（至少部分）通过浓缩方案被去除。我们的数据表明该技术有几个可进一步改进的变量，因此有潜力成为用于体内细胞类型特异性人类基因治疗的强大工具。
