Interleukins of the IL-2 family have different activity against natural killer cells when used in combination with IL-18.
Authors: Shamova Tatiana V. , Kit Oleg I. , Mezhevova Irina V. , Filippova Svetlana Yu , Sitkovskaya Anastasia O. , Gnennaya Nadezhda V. , Timofeeva Sofia V. , Novikova Inna A. , Zlatnik Elena Yu , Bondarenko Elena S. , Vashchenko Larisa N. , Kechedzhieva Emma E. , Dashkova Irina R. , Ausheva Tatiana V. , Andreiko Elena А. , Pozdnyakova Viktoria V. 
Published: June 2022

e14511

Background: The main effort to improve adoptive cancer immunotherapy is aimed at overcoming the limitations associated with low MHC expression on tumor cells. Reinfusion of patient's ex vivo activated NK cells, which eliminate tumor cells, regardless of the MHC molecules presence on their surface, is among promising approaches. Natural killers fate is regulated by a whole set of cytokines, of which IL-18 and interleukins of the IL-2 family (IL-2, IL-15, IL-7, IL-21) are of particular importance. Despite the common structure of their receptor, interleukins of the IL-2 family may exert different influence on NK phenotype when combined with IL-18. The aim of the study was to investigate the effect of IL-18 combinations with interleukins of IL-2 family on activation markers expression on breast cancer patients’ peripheral blood NK cells. Methods: Peripheral blood NK cells were enriched by magnetic cell sorting from PBMC using the NK Cell Isolation Kit (#130-092-657, Miltenyi Biotec, Germany). Next, the sorted NK were incubated at 106 cells/ml in RPMI 1640 medium (Gibco, USA) with addition of cytokines at a concentration of 10 ng/ml in 5 variants: 1) IL-18; 2) IL-18 + IL-2; 3) IL-18 + IL-7; 4) IL-18 + IL-15; 5) IL-18 + IL-21. Cells were further incubated at 5.0% CO2 and 370C. After 48 hours of incubation, the expression of CD16, CD56 and CD25 markers on NK was assessed by flow cytometry. Results: A decrease in CD16++CD56+ subpopulation percentage up to 52.5% and 54.9% compared to 62.2% in the control was observed after incubation with IL-18 alone and when it was combined with IL-2, respectively. At the same time, the addition of IL-15 and IL-21 caused a further decrease to 38.7% and 39.1%, respectively. Nevertheless the combination of IL-18 and IL-7 led to an increase in CD16++CD56+ subpopulation percentage up to 71.8%. The percentage of CD25-positive NK cells on the whole followed the dynamics of CD16++CD56+ subpopulation with the IL-18 +IL-15 combination causing the maximum decrease (up to 7.6%). The proportion of NK cells with the CD16+CD56++ phenotype, on the contrary, increased after incubation with IL-18 alone and its combination with IL-2 from 34.5% in control to 46.9% and 44.3%, respectively. Interleukin 18 combinations with IL-15 and IL-21 exhibited even more prominent effect causing increase in CD16+CD56++ proportion to 59.6% and 60.4%, respectively. At the same time, incubation with IL-18+IL-7 caused only a slight decrease in the percentage of the CD16++CD56+ NK subpopulation (up to 27.5%). Conclusions: Thus, IL-18 stimulated the generation of the cytokine-producing NK fraction while suppressing the cytolytic fraction. Also the antagonistic effect of IL-7 with IL-18 and the synergistic effect of IL-15 and IL-21 with IL-18 were revealed on NK cells in vitro.