Umbilical cord blood as a promising source of NK cells for immunotherapy

Currently, a large number of studies on genetic modification of cord blood NK cells (UCB-NK) are carried out at both clinical and preclinical levels. Immunotherapy based on UCB-NK cells has great potential for antitumor therapy. However, despite having known several advantages over peripheral blood NK cells (PB- NK), including a high concentration in cord blood and low virulence rate, UCB-NK cells are predominantly characterized in the scientific literature as immature and low-functioning NK cells. In this work, we studied the phenotypic characteristics of UCB-NK cells and the possibility of stimulatory compensation of the decreased functional activity of UCB-NK cells. Our studies revealed UCB-NK cells can be characterized as poorly differentiated and weakly activated cells with high level of inhibitory receptor NKG2A and low level of activating receptor NKG2C and HLA-DR, accordingly with the literature data. Two types of stimuli were chosen to stimulate freshly isolated UCB-NK cells: 1) 100 units of IL-2; 2) combinations of 100 units IL-2 and K-562 feeder cells expressing membrane-bound IL-21 (K562-mbIL21). It was shown the degranulation (LAMP-1) and proliferative activity was higher than for parallel cultured ex vivo PB-NK cells under the same conditions for UCB-NK cells stimulated for 7 days with IL-2 + K562-mbIL21. Moreover, stimulation in the way of IL-2 + K562-mbIL21 seemed to be a more perspective way to obtain a large number of proliferatively active UCB-NK cells compared to stimulation with IL-2 only. Since genetic modification of NK cells is a promising way to improve the antitumor properties of NK cells, retroviral transduction procedure was performed to study of the stimulated UCB-NK cells. UCB-NK cells stimulated with IL-2 + K562-mbIL21 were transduced on day 8 of cultivation. In this study, we used targeted overexpression of the adaptor molecule DAP12, which is involved in the signaling of activating NK cell receptors. PB-NK cells and UCB-NK cells were transduced under the equal experimental conditions in same volume of viral particles. As a result, the transduction efficiency was found to be more than 4-fold higher for UCB-NK cells compared to PB-NK cells. Thus, UCB-NK cells appear to be a promising tool for further research in cancer immunotherapy.

1. Alnabhan R., Madrigal A., Saudemont A. Differential activation of cord blood and peripheral blood natural killer cells by cytokines. Cytotherapy, 2015, Vol. 17, pp. 73-85.

2. Barker J.N., Wagner J.E. Umbilical cord blood transplantation: current practice and future innovations. Crit. Rev. Oncol. Hematol., 2003, Vol. 48, pp. 35-43.

3. Campbell K.S., Colonna M. DAP12: a key accessory protein for relaying signals by natural killer cell receptors. Int. J. Biochem. Cell Biol., 1999, Vol. 31, pp. 631-636.

4. Dalle J.H., Menezes J., Wagner E., Blagdon M., Champagne J., Champagne M.A., Duval M. Characterization of cord blood natural killer cells: implications for transplantation and neonatal infections. Pediatr. Res.,2005, Vol. 57, no, 5, Pt 1, pp. 649-655.

5. Erokhina S.A., Streltsova M.A., Kanevskiy L.M., Telford W.G., Sapozhnikov A.M., Kovalenko E.I. HLA- DR+ NK cells are mostly characterized by less mature phenotype and high functional activity. Immunol. Cell Biol., 2018, Vol. 96, 212. doi:10.1111/IMCB.1032.

6. Kobyzeva P.A., Streltsova M.A., Erokhina S.A., Kanevskiy L.M., Telford W.G., Sapozhnikov A.M., Kovalenko E.I. CD56dim CD57- NKG2C+ NK cells retaining proliferative potential are possible precursors of CD57+ NKG2C+ memory-like NK cells. J. Leukoc. Biol., 2020, Vol. 108, no. 4, pp. 1379-1395.

7. Ljunggren H.G., Karre K. In Search of the “Missing Self”: MHC molecules and NK cell recognition. Immunol. Today, 1990, Vol. 11, pp. 237-244.

8. Luevano M., Daryouzeh M., Alnabhan R., Querol S., Khakoo S., Madrigal A., Saudemont A. The unique profile of cord blood natural killer cells balances incomplete maturation and effective killing function upon activation. Hum. Immunol., 2012, Vol. 73, pp. 248-257.

9. Romee R., Rosario M., Berrien-Elliott M.M., Wagner J.A., Jewell B.A., Schappe T., Leong J.W., Abdel-Latif S., Schneider S.E., Willey S., Neal C.C., Yu L., Oh S.T., Lee Y.-S., Mulder A., Claas F., Cooper M.A., Fehniger T.A. Cytokine-induced memory-like natural killer cells exhibit enhanced responses against myeloid leukemia. Sci. Transl. Med., 2016, Vol. 8, 357ra123. doi: 10.1126/scitranslmed.aaf2341.

10. Sarvaria A., Jawdat D., Madrigal J.A., Saudemont A. Umbilical cord blood natural killer cells, their characteristics, and potential clinical applications. Front. Immunol., 2017, Vol. 8, 329. doi: 10.3389/fimmu.2017.00329.

11. Schonberg K., Fischer J.C., Kogler G., Uhrberg M. Neonatal nk-cell repertoires are functionally, but not structurally, biased toward recognition of self HLA Class I. Blood, 2011, Vol. 117, pp. 5152-5156.

12. Velichinskii R.A., Streltsova, M.A., Kust S.A., Sapozhnikov A.M., Kovalenko E.I. The Biological Role and Therapeutic potential of NK cells in hematological and solid tumors. Int. J. Mol. Sci., 2021, Vol. 22, no. 21, 11385. doi: 10.3390/ijms222111385.

13. Verneris M.R., Miller J.S. The phenotypic and functional characteristics of umbilical cord blood and peripheral blood Natural Killer cells. Br. J. Haematol., 2009, Vol. 147, pp. 185-191.

14. Vivier E., Raulet D.H., Moretta A., Caligiuri M.A., Zitvogel L., Lanier L.L., Yokoyama W.M., Ugolini S. Innate or adaptive immunity? The example of natural Killer cells. Science, 2011, Vol. 331, pp. 44-49.

15. Wang Y., Xu H., Zheng X., Wei H., Sun R., Tian Z. High Expression of NKG2A/CD94 and low expression of granzyme b are associated with reduced cord blood NK cell activity. Cell. Mol. Immunol., 2007, Vol. 4, pp. 377-382.