Interleukin 21 Enhances the Expansion and Anti-Tumor Cytotoxic Activity of Cytokine-Induced Killer Cells Derived from Both Peripheral Blood and Cord Blood In Vitro.
Authors: Zhao Mingfeng , Deng Qi , Li Yuming , Lin Xuemei , Geng Li , Li Jinglan , Wang Shujie , Xu Xinnv , Shao Zonghong 
Published: October 2019

Abstract
Interleukin 21(IL-21) is a new member of interleukin 2 cytokine families which was discovered in 2000. IL-21 is produced by activated CD4 positive cells, and is known to influence T, B, NK cells and DC, and has potent anti-tumor effects. For example, IL-21 can improve the proliferation of B lymphocytes, enhance the production of IgG1; improve the proliferation and enhance the anti-tumor activity of both NK and T cells. The cytokine-induced killer (CIK) cells, which are characterized with the phenotype of CD3+CD56+, are the effective cells on adoptive cellular immunotherapy against tumors. We hypothesize that IL-21 could also affect the proliferation and function of CIK cells, thus play a certain role in the anti-tumor immunotherapy by CIK cells. The peripheral blood mononuclear cells (PBMC) and cord blood mononuclear cells (CBMC) from healthy donors were stimulated with anti-CD3 (OKT3) monoclonal antibody and IFNgamma and then expanded with IL-2 and with/without IL-21(200ng/ml). CD3+CD56+ CIK cells were counted by flow cytometry. Net lactate dehydrogenase release from target cells incubated with CIK cells was used as an index of CIK cells cytotoxicity against chronic myeloid leukemia cell line K562 and a variety of tumor target cells from patients. The concentration of the IFNgamma in the culture supernatant was measured by enzyme-linked-immunoassay, the quantity of IFNgamma RNA was measured by RT-PCR assay, and the cytotoxic activity against K562 cells by the culture supernatant was also detected. Cultured with IL-21, at day 14, the quantity of CIK cells was increased from a median of 17.5% to 26.5% (PBMC original) and from 33.8% to 55.9% (CBMC original); The cytotoxic activity rates against K562 cells by CIK cells were increased from 24.0% to 52.2% (PBMC original) and from 35.1% to 79.7% (CBMC original); The concentration of IFNgamma in the culture supernatant was increased for 1.9-fold (PBMC original), and for 3.2-fold (CBMC original); The cytotoxic activity against K562 cells by the culture supernatant was increased for 1.8-fold (PBMC original) and for 2.7-fold (CBMC original); The expression of IFNgamma RNA in CIK cells was also markedly increased derived from both PBMC and CBMC when cultured with IL-21. Moreover, the cytotoxic activity against leukemia cells from 11 patients (6 with acute lymphoblastic leukemia, 5 with acute myeloid leukemia) by CIK cells derived from CBMC were also detected. The cytotoxic activity rates were at a median of 68.3% (range, 34.7%–86.4%) when CIK cells were cultured with IL-21, rates that contrasted drastically to the cytotoxic activity rates when CIK cells were cultured without IL-21, which were only at a median of 37.4% (range, 16.1%–60.0%). In conclusion, our data indicated that IL-21 could enhance the expansion of CIK cells and their anti-tumor activity derived from both PBMC and CBMC in vitro, IFNgamma was evolved in this course although the mechanism still need to be explored. These observations open up the possibility of imagining a future clinical application of IL-21 in the anti-tumor immunotherapy by CIK cells.