Study on the effect of ATPIF1 on the anti-tumor activity of CAR-NK92 cells by regulating glycolytic capacity.
- Author:
Biao LIU
1
;
Xue GONG
1
;
Biliang HU
2
;
Chunlei GUO
1
;
Genshen ZHONG
3
Author Information
1. Henan Key Laboratory of Immunology and Targeted Drugs, School of Medical Technology, Xinxiang Medical University, Xinxiang 453003, China.
2. Hunan Siweikang Pharmaceutical Co., Ltd., Changsha 410017, China.
3. School of Biological and Chemical Engineering, Changsha University, Changsha 410022, China. *Corresponding author, E-mail: zhonggs@ccsu.edu.cn.
- Publication Type:Journal Article
- MeSH:
Humans;
Glycolysis;
Killer Cells, Natural/metabolism*;
Receptors, Chimeric Antigen/immunology*;
Granzymes/genetics*;
Hexokinase/metabolism*;
Cell Line, Tumor;
Interleukin-2/genetics*;
Cell Proliferation;
NK Cell Lectin-Like Receptor Subfamily K/genetics*;
Membrane Potential, Mitochondrial
- From:
Chinese Journal of Cellular and Molecular Immunology
2025;41(10):865-874
- CountryChina
- Language:Chinese
-
Abstract:
Objective To investigate the effect of ATP synthase inhibitory factor 1 (ATPIF1) on the antitumor activity of chimeric antigen receptor (CAR)-NK92 cells. Methods HER2-targeted CAR-NK92 cells with ATPIF1 overexpression or knockdown were constructed. CAR-positive expression rate was detected by flow cytometry. Cell proliferation capacity was measured using CCK-8 assay. Glycolytic capacity was analyzed by Seahorse metabolic analyzer. Mitochondrial membrane potential levels were detected using JC-1 probe. Target cell lysis rate was evaluated by firefly luciferase reporter assay. Expression levels of CD107a, natural-killer group 2 member D (NKG2D), granzyme B (GzmB), perforin, and interleukin 2 (IL-2) were detected via flow cytometry. Quantitative real-time PCR was used to measure the expression of interferon-induced protein with tetratricopeptide repeats 1 (IFIT1), tumor necrosis factor α (TNF-α), ATPIF1, and hexokinase 1 (HK1). The impact of glycolytic inhibition by 2-Deoxy-D-glucose (2-DG) on CAR-NK92 antitumor capacity was examined. Results Successfully generated HER2-targeting control CAR-NK92 cells, as well as ATPIF1-overexpressing and ATPIF1 knockdown CAR-NK92 cells. The ATPIF1-overexpressing CAR-NK92 cells showed significantly enhanced target cell lysis rate, elevated expression levels of NKG2D and CD107a, increased secretion capacities of Granzyme B (GzmB) and IL-2, and upregulated mRNA expression levels of IFIT1 and TNF-α, while ATPIF1-knockdown cells exhibited opposite effects. ATPIF1 overexpression induced metabolic reprogramming in CAR-NK92 cells, manifested by significantly decreased mitochondrial membrane potential (δpsim), markedly upregulated HK1 mRNA expression, and enhanced basal glycolysis and glycolytic capacity. After glycolysis inhibition with 2-DG (5 μmol/L), both ATPIF1-overexpressing and knockdown CAR-NK92 cells showed no significant differences in NKG2D and CD107a expression levels compared to control cells. Conclusion ATPIF1 regulates the antitumor activity of CAR-NK92 cells through modulating glycolytic metabolism. Overexpression of ATPIF1 can enhance the antitumor efficacy of CAR-NK92 cells.
