Telomerase Reverse Transcriptase Increases Proliferation and Lifespan of Human NK Cells without Immortalization

Abstract

NK cells are the first line of defense against viruses and malignant cells, and their natural functionality makes these cells a promising candidate for cancer cell therapy. The genetic modifications of NK cells, allowing them to overcome some of their inherent limitations, such as low proliferative potential, can enable their use as a therapeutic product. We demonstrate that hTERT-engineered NK cell cultures maintain a high percentage of cells in the S/G2 phase for an extended time after transduction, while the life span of NK cells is measurably extended. Bulk and clonal NK cell cultures pre-activated in vitro with IL-2 and K562-mbIL21 feeder cells can be transduced with hTERT more efficiently compared with the cells activated with IL-2 alone. Overexpressed hTERT was functionally active in transduced NK cells, which displayed upregulated expression of the activation marker HLA-DR, and decreased expression of the maturation marker CD57 and activating receptor NKp46. Larger numbers of KIR2DL2/3+ cells in hTERT-engineered populations may indicate that NK cells with this phenotype are more susceptible to transduction. The hTERT-modified NK cells demonstrated a high natural cytotoxic response towards K562 cells and stably expressed Ki67, a proliferation marker. Overall, our data show that ectopic hTERT expression in NK cells enhances their activation and proliferation, extends in vitro life span, and can be a useful tool in developing NK-based cancer cell therapies.

WT1 Pulsed Human CD141+ Dendritic Cell Vaccine Has High Potential in Solid Tumor-Targeted Immunotherapy

“…As a result, NK cells may address the limitations of autologous CAR T cell therapy. However, there are several potential drawbacks to the usage of NK cells, such as difficulty in cell culture, immediacy in the peak activity of cellular kinetics, and shorter intrinsic longevity as well as limited memory phenotype in the life span and response [ 34 , 37 , 38 ]. Other research involves combinational therapies using DC or T cells and immune checkpoint inhibitors such as anti-PD1 (programmed cell death protein1) [ 39 ] or anti-PD-L1 (programmed death-ligand1) [ 40 ].…”

Overcoming tumor resistance mechanisms in CAR-NK cell therapy

“…However, transfected NK cells were not able to grow autonomously in nonobese diabetic severe‐combined‐immunodeficient γc−/− (NSG) mice and still eventually developed delayed senescence in vitro ( 147 ). More recently, Streltsova and coworkers corroborated that stable hTERT ectopic expression, even when gamma-retrovirus is used, increases the proliferation and lifespan of expanded and activated (K562-mbIL21+IL2) NK cells rather than complete immortalization ( 168 ). The safety of this strategy is a critical question because so far, the inability of ectopically expressed hTERT to cause oncogenic transformation of NK cells has not been firmly established.…”

Multiple Actions of Telomerase Reverse Transcriptase in Cell Death Regulation

“…A number of studies have shown that cells with an increased level of TERT have greater resistance to external damaging factors and are more protected from cell death [ 8 , 33 , 35 , 41 , 49 , 50 , 52 , 77 , 107 ]. Such stability, according to recent data, is due not only to the intranuclear activity of TERT (lengthening of telomeric ends and regulation of gene expression), but also due to the its cytoplasmic function: cell death regulation through direct interaction with mitochondria and proteins of the apoptotic pathway.…”

“…Immune cells transferred into a recipient were found to rapidly downregulate their activity despite their accumulation within tumor sites [ 153 ]. It was shown that TERT overexpression in modified T [ 154 ] and NK cells [ 107 , 155 ] increases their replicative potential and overall functional activity after long-lasting cultivation. This approach might help to overcome the development of immunosuppression during cancer treatment [ 156 , 157 , 158 ].…”