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
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 <i>hTERT</i> and <i>iCasp9</i> genes. hTERT overexpression can prevent the functional exha...
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MDPI AG
2024-03-01
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| author | Anastasia I. Palamarchuk Elena I. Kovalenko Maria A. Streltsova |
| author_facet | Anastasia I. Palamarchuk Elena I. Kovalenko Maria A. Streltsova |
| author_sort | Anastasia I. Palamarchuk |
| collection | DOAJ |
| description | 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 <i>hTERT</i> and <i>iCasp9</i> 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 <i>iCasp9</i> 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 <i>hTERT</i> and <i>iCasp9</i> genes were shown to affect the EOMES and T-BET levels differently in the NK cells. The EOMES<sup>+</sup>T-BET<sup>+</sup> 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. |
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| last_indexed | 2024-04-24T18:31:14Z |
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| spelling | doaj.art-6ead65204c8841af80adca78422cdae62024-03-27T13:23:02ZengMDPI AGBiomedicines2227-90592024-03-0112365010.3390/biomedicines12030650The 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 StimulationAnastasia I. Palamarchuk0Elena I. Kovalenko1Maria A. Streltsova2Shemyakin & Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Ul. Miklukho-Maklaya 16/10, 117997 Moscow, RussiaShemyakin & Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Ul. Miklukho-Maklaya 16/10, 117997 Moscow, RussiaShemyakin & Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Ul. Miklukho-Maklaya 16/10, 117997 Moscow, RussiaThe 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 <i>hTERT</i> and <i>iCasp9</i> 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 <i>iCasp9</i> 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 <i>hTERT</i> and <i>iCasp9</i> genes were shown to affect the EOMES and T-BET levels differently in the NK cells. The EOMES<sup>+</sup>T-BET<sup>+</sup> 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.https://www.mdpi.com/2227-9059/12/3/650NK cellsEOMEST-BET<i>hTERT</i><i>iCasp9</i>stimulation |
| spellingShingle | Anastasia I. Palamarchuk Elena I. Kovalenko Maria A. Streltsova 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 Biomedicines NK cells EOMES T-BET <i>hTERT</i> <i>iCasp9</i> stimulation |
| title | 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 |
| title_full | 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 |
| title_fullStr | 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 |
| title_full_unstemmed | 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 |
| title_short | 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 |
| title_sort | 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 |
| topic | NK cells EOMES T-BET <i>hTERT</i> <i>iCasp9</i> stimulation |
| url | https://www.mdpi.com/2227-9059/12/3/650 |
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