敲除CISH敲入IL-15显著增强NK细胞抗肿瘤活性的研究

刘传玲; 李营营; 李燕荣; 胡梦; 唐立春; 王海燕; 焦顺昌; 卢社莲; 赵卫红

doi:10.12435/j.issn.2095-5227.2023.129

敲除CISH敲入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 NK cell antitumor activity

摘要

摘要:
背景 IL-15是调节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、CISH基因敲除 + IL-15基因敲入组杀伤效率明显增强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 impacting 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 anti-tumor activity of NK cells.

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