肿瘤细胞治疗研究
英文原题：Anti-Her2 CAR-NK92 Cells and Their Exosomes: Generation, Characterization, and Selective Cytotoxicity Against Her2-Positive Tumor Cells.
嘌呤霉素筛选有效富集了表达 CAR 的细胞，GFP 阳性率达到 99.8%，CAR 表面表达较野生型细胞增加 15 倍。
CAR 工程化 NK 细胞是 Her2 阳性癌症靶向免疫治疗的一种有前景方法。本研究旨在制备抗 Her2 CAR-NK92 细胞，评估其对 Her2 阳性癌细胞的选择性细胞毒性，并分离和表征其释放的外泌体。研究者使用编码抗 Her2 CAR 和辅助转座酶的 piggyBac 转座子载体对 NK92 细胞进行电转，并通过嘌呤霉素筛选富集转导细胞。采用流式细胞术评估 CAR 和 GFP 表达，并对外泌体的蛋白货物及表面蛋白表达进行分离和表征。通过实时细胞分析，评估其对 Her2 阳性 SK-BR3 细胞和 Her2 阴性 MCF-7 细胞的细胞毒性。电转未显著影响 NK92 细胞活率。嘌呤霉素筛选有效富集 CAR 表达细胞，GFP 阳性率达到 99.8%，CAR 表面表达较野生型细胞提高 15 倍。CAR-NK92 细胞表现出强效、依赖效靶比的 Her2 特异性细胞毒性，在 10:1 效应细胞与靶细胞比例时作用最强。CAR-NK92 来源外泌体携带 CAR 分子，并选择性靶向 Her2 阳性细胞。抗 Her2 CAR-NK92 细胞及其外泌体对 Her2 阳性癌细胞均表现出强且选择性的细胞毒性，支持其作为实体瘤创新免疫治疗药物的潜力。
Chimeric antigen receptor (CAR)-engineered NK cells are a promising approach for targeted immunotherapy in Her2-positive cancers. This study aimed to generate anti-Her2 CAR-NK92 cells, to evaluate their selective cytotoxicity against Her2-positive cancer cells, and to isolate and characterize their released exosomes. NK92 cells were electroporated with piggyBac transposon vectors encoding anti-Her2 CAR and the helper transposase. Puromycin selection was performed to enrich the transduced cells. CAR and GFP expression were assessed by flow cytometry, and exosomes were isolated and characterized in terms of protein cargo and surface protein expression. Cytotoxicity was evaluated using real-time cell analysis against Her2-positive SK-BR3 cells and Her2-negative MCF-7 cells. Electroporation did not significantly affect NK92 cell viability. Puromycin selection efficiently enriched for CAR-expressing cells, with GFP positivity reaching 99.8% and a 15-fold increase in CAR surface expression compared to wild-type cells. CAR-NK92 cells demonstrated robust, Her2-specific cytotoxicity in a E:T-dependent manner, with the greatest effect observed at a 10:1 effector-to-target ratio. Exosomes derived from CAR-NK92 cells contained CAR molecules and selectively targeted Her2-positive cells. Anti-Her2 CAR-NK92 cells and their exosomes exhibit potent and selective cytotoxicity against Her2-positive cancer cells, supporting their potential as innovative immunotherapeutic agents for solid tumors.
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