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

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。

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%vs 71.40±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 antitumor activity of NK cells.