敲除CISH敲入并IL-15显著增强NK细胞抗肿瘤活性的研究
CISH deletion and IL-15 up-regulation of NK cell greatly improve its antitumor activity
刘传玲 1李营营 2李燕荣 2胡梦 2唐立春 2王海燕 2焦顺昌 3卢社莲 2赵卫红3
作者信息
- 1. 解放军总医院第一医学中心肿瘤内科,北京 100853
- 2. 鼎成肽源生物有限公司研发部,北京 102200
- 3. 解放军总医院肿瘤医学部,北京 100853
- 折叠
摘要
背景 白细胞介素(interleukin,IL)-15是调节自然杀伤细胞(natural killer cell,NK细胞)功能的关键细胞因子,而CISH可负向调控IL-15信号通路,从而影响NK细胞功能.靶向CISH基因和IL-15基因对NK细胞进行基因编辑,可增强NK细胞抗肿瘤活性.目的 探讨CISH基因、IL-15基因对NK细胞的细胞毒性的影响.方法 采用AsCas12a Ultra基因编辑工具敲除CISH基因,并敲入IL-15基因,对NK细胞进行基因编辑,并通过基因测序检测CISH基因敲除效率,采用ELISA检测IL-15 的表达,用LDH检测基因编辑后的NK细胞对H358细胞系的杀伤活性,评估CISH基因敲除、IL-15基因敲入对NK细胞抗肿瘤功能的影响.结果 与对照组相比,CISH基因敲除组和CISH基因敲除+IL-15基因敲入组杀伤效率显著增强[E∶T=2∶1组,34.57%±6.32%vs 71.40%±5.81%和 69.23%±4.28%,P＜0.001].结论 敲除CISH基因可上调IL-15信号通路,增强NK细胞抗肿瘤活性.
Abstract
Background IL-15 plays a crucial role in regulating the function of natural killer(NK)cells,while CISH acts as a negative regulator of the IL-15 signaling pathway,thereby affecting NK cell function.Gene editing targeting the CISH and IL-15 genes in NK cells has been shown to enhance their anti-tumor activity.Objective To investigate the effect of CISH gene and IL-15 gene on cytotoxicity of NK cells.Methods The CISH gene was disrupted using the AsCas12a Ultra gene editing tool,while the IL-15 gene was introduced to facilitate genetic modification of NK cells.The knockout efficiency of the CISH gene was assessed through gene sequencing,and the expression of IL-15 was measured via ELISA.Subsequently,the cytotoxicity of NK cells against H358 cell line post-gene editing was evaluated using LDH assay,and the impact of CISH knockout on IL-15-mediated anti-tumor function in NK cells was examined.Results The killing efficiency of the CISH gene knockout group and the CISH KO+IL-15 KI group was significantly enhanced compared to the control group(E:T=2:1 group,34.57%±6.32%vs71.40±5.81%and 69.23%±4.28%,P<0.001).Conclusion The knockout of the CISH gene can enhance the IL-15 signaling pathway and augment the anti-tumor activity of NK cells.
关键词
自然杀伤细胞/CISH/白细胞介素-15/AsCas12a/UltraKey words
NK cell/CISH/IL-15/AsCas12a ultra引用本文复制引用
出版年
2024
国家科技期刊平台
NSTL
万方数据