[Modification with IL-21 and CCL19 enhances killing efficiency and tumor infiltration of NKP30 CAR-T cells in lung cancer].

Q3 Medicine
Z Zhou, S Liu, J Li, M Chen, H Lin, Y Chen, W Chen, J Lin, H Zhou, Q Zheng
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引用次数: 0

Abstract

Objective: To investigate whether modification with IL-21 and CCL19 enhances killing and tumor-infiltrating efficiency of NKP30 CAR-T cells in lung cancer.

Methods: The modified IL-21-CCL19 NKP30 CAR-T cells expressing IL-21 and CCL19 fusion gene was constructed based on NKP30 CAR-T cells and stimulated with CD3CD28 antibodies and IL-2. The immunophenotype and migration of the cells in the presence of IL-21 were investigated using flow cytometry and migration experiments. Lactate dehydrogenase (LDH) release and sphere formation assays were used to assess the killing and infiltration capabilities of CAR-T cells, and the secretion levels of IFN-γ, IL-21 and CCL19 were determined with enzyme-linked immunospot assay (ELISPOT) and ELISA. A zebrafish model bearing HCG-27 cell xenograft was established by microinjection of the tumor cells into the yolk sac followed 24 h later by injection of the immune cells at the same site, and the fluorescence signals were captured using a fluorescent microscopy.

Results: The NKP30 ligand B7H6, which was almost undetectable in normal tissues and blood cells, was highly expressed (over 90%) in lung cancer cells. Compared with NKP30 CAR-T cells and conventional T cells, IL-21-CCL19 NKP30 CAR-T cells exhibited stronger proliferative and migration capabilities with the formation of central memory T cells. The reduced expressions of CTLA4 and PD1 in the constructed cells resulted in enhanced killing efficiency against lung cancer cells accompanied by significantly increased production of IFN-γ, IL-21 and CCL19. In the zebrafish models, CAR-T cells exhibited stronger cytotoxicity and proliferative abilities than typical T cells, but these differences were not statistically significant between the two CAR-T cells.

Conclusion: Modification of NKP30 CAR-T cells with IL-21 and CCL19 facilitates their access into solid tumors for more effective tumor cell killing while producing a large number of memory T cells.

[用 IL-21 和 CCL19 修饰可提高 NKP30 CAR-T 细胞在肺癌中的杀伤效率和肿瘤浸润】。]
目的方法:在NKP30 CAR-T细胞的基础上构建表达IL-21和CCL19融合基因的IL-21-CCL19修饰NKP30 CAR-T细胞,并用CD3CD28抗体和IL刺激:方法:以NKP30 CAR-T细胞为基础,用CD3CD28抗体和IL-2刺激,构建表达IL-21和CCL19融合基因的IL-21-CCL19修饰NKP30 CAR-T细胞。使用流式细胞仪和迁移实验研究了细胞在 IL-21 存在下的免疫表型和迁移。乳酸脱氢酶(LDH)释放和小球形成实验用于评估CAR-T细胞的杀伤和浸润能力,酶联免疫吸附试验(ELISPOT)和酶联免疫吸附试验测定了IFN-γ、IL-21和CCL19的分泌水平。将肿瘤细胞显微注射到卵黄囊中,24小时后在同一部位注射免疫细胞,建立了携带HCG-27细胞异种移植的斑马鱼模型,并用荧光显微镜捕捉荧光信号:结果:在正常组织和血细胞中几乎检测不到的NKP30配体B7H6在肺癌细胞中高表达(超过90%)。与NKP30 CAR-T细胞和传统T细胞相比,IL-21-CCL19 NKP30 CAR-T细胞具有更强的增殖和迁移能力,并能形成中心记忆T细胞。构建的细胞中 CTLA4 和 PD1 的表达量减少,从而提高了对肺癌细胞的杀伤效率,同时 IFN-γ、IL-21 和 CCL19 的产生量也显著增加。在斑马鱼模型中,CAR-T 细胞比典型的 T 细胞表现出更强的细胞毒性和增殖能力,但这两种 CAR-T 细胞之间的差异没有统计学意义:结论:用 IL-21 和 CCL19 修饰 NKP30 CAR-T 细胞有助于它们进入实体瘤,更有效地杀死肿瘤细胞,同时产生大量记忆 T 细胞。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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来源期刊
南方医科大学学报杂志
南方医科大学学报杂志 Medicine-Medicine (all)
CiteScore
1.50
自引率
0.00%
发文量
208
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