Department of Biochemistry, University of Medicine and Dentistry of New Jersey–Robert Wood Johnson Medical School, 675 Hoes Lane, Piscataway, NJ 08854, USA
Virol J. 2013 Jun 14;10:194. doi: 10.1186/1743-422X-10-194.
Osteosarcomas are the most common primary bone malignancies found in children and adolescents. An optimized system was developed for efficient retroviral gene delivery into solid 143B osteosarcoma tumors in mice using a retargeted Env. In these studies, the viral Env CP was isolated from an in vitro screen of a library of feline leukemia virus Env randomized in the receptor-binding domain and maintained high titer on human 143B osteosarcoma cell line.
The vector developed to express the random Env libraries encoded the drug selectable marker neo. To adapt this for studies in live animals, the murine based vector was modified to express the luciferase gene. The bicistronic vector developed expressed both the CP Env and luciferase in the presence of either the MPMV CTE or a WPRE element. Virus bearing the CP FeLV Env variant maintained high titers after concentration allowing for direct visualization of delivery of the luciferase gene in subcutaneous 143B osteosarcoma tumors.
This system serves as a proof-of-concept for the use of novel FeLV Env pseudotyped MLV particles for in vivo gene delivery. Gene delivery and expression of lucerifase from viral particles bearing the CP Env was readily detected in live mice after a single round of intratumor injection.
骨肉瘤是儿童和青少年中最常见的原发性骨恶性肿瘤。我们开发了一种优化的系统，使用重定向的 Env 高效地将逆转录病毒基因递送至小鼠的实体 143B 骨肉瘤肿瘤中。在这些研究中，病毒 Env CP 是从猫白血病病毒 Env 文库的体外筛选中分离出来的，该文库在受体结合域中随机化，并在人 143B 骨肉瘤细胞系上保持高滴度。
开发的表达随机 Env 文库的载体编码了药物选择标记 neo。为了将其适应于活体动物研究，我们对基于鼠的载体进行了修饰以表达荧光素酶基因。开发的双顺反子载体在存在 MPMV CTE 或 WPRE 元件的情况下表达 CP Env 和荧光素酶。携带 CP FeLV Env 变体的病毒在浓缩后仍保持高滴度，从而允许直接观察到荧光素酶基因在皮下 143B 骨肉瘤肿瘤中的递送。
该系统证明了使用新型 FeLV Env 假型 MLV 颗粒进行体内基因递送的概念验证。在单次肿瘤内注射后，在活体小鼠中很容易检测到携带 CP Env 的病毒颗粒的基因传递和荧光素酶的表达。
