Abstract

Purpose: Glioblastoma is the most malignant primary adult brain tumor with a poor prognosis; therefore, novel therapies are needed. Natural Killer (NK) cells are promising candidates for immunotherapy because of their ability to eliminate tumor cells without prior sensitization. An inadequate number of NK cells remain a major obstacle. The primary purpose of this study was to develop NK cells from Cord Blood Hematopoietic Stem Cells (CB-HSCs) on Umbilical Cord Mesenchymal Stem Cells (UC-MSCs) feeder layer and analyse cytotoxic potential of NK cells against glioblastoma in vitro.

Methods: UC-MSCs were treated with mitomycin-C to be used as a feeder layer. CB-HSCs were co-cultured with UC-MSCs and differentiated into NK cells by using medium containing cytokines such as thrombopoietin, Flt3- ligand, stem cell factor, IL-6, IL-7, IL-15 and IL-2. NK cells were characterized by immunocytochemistry. The in vitro cytotoxic effect of NK cells on T98-glioblastoma cells was determined by annexin V‑fluorescein isothiocyanate/ propidium iodide staining followed by flow cytometric analysis. qRT-PCR was performed to measure gene expression levels of KRAS, TP53, TGFBR2 and NANOG in glioblastoma cells after treatment with NK cells.

Results: NK cells were successfully differentiated from CB-HSCs on the UC-MSC feeder layer with the strong expression of cytotoxic receptors after 6 weeks. They demonstrated potent cytotoxicity against glioblastoma in vitro. Additionally, the KRAS oncogene expression in glioblastoma cells decreased upon co-culture with NK cells.

Conclusion: NK cells differentiated from CB-HSCs on UC-MSC feeder-layer were capable of eliminating glioblastoma cells via apoptosis in vitro and warrant further investigation in vivo and clinical settings.

Keywords: Apoptosis; Tissues; Monocyte Chemo Attractant Protein; Brain tumor; Cancer; Immunotherapy