Zhao Y, Zhu L, Lee S, Li L, Chang E, Soong N W, Douer D, Anderson W F
Gene Therapy Laboratories, Norris Cancer Center, University of Southern California School of Medicine, 1441 Eastlake Avenue, Los Angeles, CA 90033, USA.
Proc Natl Acad Sci U S A. 1999 Mar 30;96(7):4005-10. doi: 10.1073/pnas.96.7.4005.
A chimeric retroviral vector (33E67) containing a CD33-specific single-chain antibody was generated in an attempt to target cells displaying the CD33 surface antigen. The chimeric envelope protein was translated, processed, and incorporated into viral particles as efficiently as wild-type envelope protein. The viral particles carrying the 33E67 envelope protein could bind efficiently to the CD33 receptor on target cells and were internalized, but no gene transfer occurred. A unique experimental approach was used to examine the basis for this postbinding block. Our data indicate that the chimeric envelope protein itself cannot participate in the fusion process, the most reasonable explanation being that this chimeric protein cannot undergo the appropriate conformational change that is thought to be triggered by receptor binding, a suggested prerequisite to subsequent fusion and core entry. These results indicate that the block to gene transfer in this system, and probably in most of the current chimeric retroviral vectors to date, is the inability of the chimeric envelope protein to undergo this obligatory conformational change.
为了靶向表达CD33表面抗原的细胞，构建了一种包含CD33特异性单链抗体的嵌合逆转录病毒载体（33E67）。嵌合包膜蛋白的翻译、加工以及整合到病毒颗粒中的效率与野生型包膜蛋白一样高。携带33E67包膜蛋白的病毒颗粒能够有效地结合靶细胞上的CD33受体并被内化，但未发生基因转移。采用了一种独特的实验方法来研究这种结合后阻断的基础。我们的数据表明，嵌合包膜蛋白本身不能参与融合过程，最合理的解释是这种嵌合蛋白不能经历被认为由受体结合触发的适当构象变化，而这是后续融合和核心进入的一个假定前提条件。这些结果表明，该系统中以及可能到目前为止的大多数当前嵌合逆转录病毒载体中基因转移的阻断原因是嵌合包膜蛋白无法经历这种必需的构象变化。
