The hTERT and iCasp9 Transgenes Affect EOMES and T-BET Levels in NK Cells and the Introduction of Both Genes Improves NK Cell Proliferation in Response to IL2 and IL15 Stimulation

• Published in: Biomedicines Vol. 12; no. 3; p. 650
• Main Authors: Palamarchuk, Anastasia I, Kovalenko, Elena I, Streltsova, Maria A
• Format: Journal Article
• Language: English
• Published: Switzerland MDPI AG 01-03-2024; MDPI
• Subjects: Antibiotics; Apoptosis; Cancer; Cell culture; Cell proliferation; Cytokines; Cytotoxicity; EOMES; Gene expression; Genes; hTERT; iCasp9; Immune checkpoint; Interleukin 15; Killer cells; Natural killer cells; NK cells; Phenotypes; Plasmids; Signal transduction; stimulation; T-BET; Telomerase reverse transcriptase; Transcription factors; Transgenes; Japan; Russia; United States > US; EOMES; NK cells; survival; exhaustion; hTERT; T-BET; iCasp9; immune checkpoints; stimulation
• ISSN: 2227-9059; 2227-9059
• DOI: 10.3390/biomedicines12030650

The NK cell exhaustion state evolving during extensive and prolonged cultivation is still one of the limitations of NK cell approaches. In this research, we transduced NK cells with the
and
genes. hTERT overexpression can prevent the functional exhaustion of NK cells during long-term cultivation, but, still, the therapeutic use of such cells is unsafe without irradiation. To overcome this obstacle, we additionally transduced NK cells with the
transgene that enables the rapid elimination of modified cells. We compared the proliferative and functional activities of the hTERT- and/or iCasp9-modified NK cells, determined their exhaustion state and monitored the levels of EOMES and T-BET, the main NK cell transcription factors. The
and
genes were shown to affect the EOMES and T-BET levels differently in the NK cells. The EOMES
T-BET
phenotype characterized the functionally active NK cells during two months of culture upon stimulation with IL2 and K562-mbIL21 feeder cells, which induced the greatest expansion rates of the NK cells, independently of the transgene type. On the other hand, under cytokine stimulation, the hTERT-iCasp9-NK cells displayed improved proliferation over NK cells modified with iCasp9 alone and showed an increased proliferation rate compared to the untransduced NK cells under stimulation with IL2 and IL15, which was accompanied by reduced immune checkpoint molecule expression. The individual changes in the EOMES and T-BET levels strictly corresponded to the NK cell functional activity, the surface levels of activating and inhibitory receptors along with the expansion rate and expression levels of pro-survival and pro-apoptotic genes.