Application of self-cleaving P2A peptide for the expansion of natural killer cells

doi:10.3877/cma.j.issn.2095-1221.2019.05.005

Abstract

Abstract:

Objective

The purpose of this study was to improve differentiation and expansion for nature killer cells in vitro using self-cleaving P2A peptide.

Methods

IL-15 and 4-1BBL genes were connected via P2A. Lentiviral vector and drug screening were used to produce K562 feeder cells which were stably expressing IL-15 and 4-1BBL. Subsequently, they were co-cultured with human PBMC as feeder cells for NK cell differentiation and proliferation. The results were expressed as ±s and compared by two-sample t-test.

Results

The fold of CD3^-CD16⁺56⁺ NK cell expansion was 4480.43+37.80 times in vitro. The proportion of CD3^-CD16⁺56⁺ NK cells was 94.79%. The percentage of IFN-γ and TNF-α positive NK cells was (63.07±3.37) % and (54.85±2.04) %, respectively, which were higher than that of the control group (16.28±2.86) % and (14.53±1.15) %, (t = 62.25 and t = 22.66 respectively, P < 0.01) . The concentrations of IFN-γ and TNF-α secreted by NK cells were (111.39±6.95) pg/ml and (32.76±3.23) pg/ml, respectively, and higher than that of the control group (44.99±4.74) g/ml and (11.09±2.45) pg/ml, (t = 20.56 and t = 7.21 respectively, P < 0.01) . In vitro, the killing rates of NK cells against K562 and H520 cells were (78.52±7.36) % and (65.03±3.27) %, respectively, which were higher than those of the control group (48.53±6.66) % and (35.85±3.99) %, t = 11.56 and t = 11.35 respectively, P < 0.01) .

Conclusion

We have successfully improved the transmembrane expression of IL-15 and 4-1BBL in K562 feeder cells, and established a better culture method to obtain NK cells from human PBMCs.

Key words: Self-cleaving P2A peptide, Natural killer cells, Feeder cells, Immunotherapy

Xuemei Chen, Xi Zhang, Jianwei Lin, Zhenzhi Xue, Tao Liu. Application of self-cleaving P2A peptide for the expansion of natural killer cells[J]. Chinese Journal of Cell and Stem Cell(Electronic Edition), 2019, 09(05): 282-291.

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Figures/Tables 14

References 25

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