融合IL2的慢病毒实现对T细胞的高效基因转导

• 作 者: 王依依 杜晓 胡志嵩 刘欣禹 欧阳清 赵晶 杨安钢 阎博
• 机 构: 新乡医学院医学检验学院 河南省新乡市, 453004;第四军医大学生物化学与分子生物学教研室, 肿瘤生物学国家重点实验室 西安市, 710032;第四军医大学生物化学与分子生物学教研室, 肿瘤生物学国
• 出 处: 医学分子生物学杂志
• 发表日期: 2022年1期
• 关键词: T lymphocytes envelope glycoprotein interleukin-2-displaying lentiviruses transd

目的 制备融合IL2的慢病毒以提高对人原代T淋巴细胞的转导效率.方法 将白细胞介素-2(IL2)基因定向克隆至水疱性口炎病毒G蛋白(VSV-G)编码基因5′端,构建新型慢病毒包装辅助质粒pMD2.IL2-G.使用不同剂量pMD2.IL2-G进行慢病毒包装,应用免疫印迹和ELISA比较病毒滴度,流式细胞术检测病毒对T细胞的转导效率.结果 单独使用pMD2.IL2-G显著降低病毒得率.与传统慢病毒相比,pMD2.IL2-G与原始辅助质粒pMD2.G以1:4混合制备的慢病毒,能够显著提高EGFP基因导入T细胞的效率(89.2％vs.40.2％),且病毒得率相当.结论 融合IL2的慢病毒能够对T细胞进行高效的基因转导,为基因修饰T细胞的临床应用提供有力支持.“,”Objective To construct the interleukin 2 (IL2) -displaying lentivirus to improve the transduction efficiency of human primary T lymphocytes. Methods The IL2 gene was fused to the N-terminus of vesicular stomatitis virus G protein to produce the novel lentivirus-packaging plas-mid pMD2.IL2-G. The yields of lentiviral particles pseudotyped with different doses of pMD2.IL2-G were compared using immunoblotting and ELISA. The infected capacity of IL2-displaying lentiviruses to T lymphocytes from healthy donors was tested using flow cytometry. Results The yield of lentivi-ral particles pseudotyped with pMD2.IL2-G was significantly less than that of wild-type lentiviruses packaged by pMD2.G. The mixture of pMD2.IL2-G and pMD2.G at a ratio of 1: 4 could obtain the equivalent yield of lentiviruses that of the wild-type, and significantly improve the transduction effi-ciency of enhanced green fluorescent protein into the T lymphocytes compared to the wild-type lenti-viruses (89.2 ％vs.40.2 ％). Conclusion Our results demonstrate that IL2-displaying lentiviral particles have a greatly enhanced transduction efficiency into human T lymphocytes. This research is expected to provide strong support for the clinical application of genetically modified T cells.