Changes in the Immunologic Phenotype of Human Malignant Glioma Cells after Passaging in Vitro

Abstract Although immunotherapeutic strategies against glioblastomas have been promising both in vitro and in animal models, similar successes have not been realized in human clinical trials. One reason may be that immunotherapeutic strategies are based on prior studies that primarily have used human glioblastoma cell lines passaged in vitro , which may not accurately reflect the in vivo properties of glioblastoma cells. In this report, we used flow cytometry to quantify the expression of immunological cell surface molecules on human glioblastomas directly ex vivo (prior to any in vitro culturing) and after varying passages in vitro . Furthermore, we used ELISA to quantitate cytokine secretion after various passages in vitro . We demonstrate that in vitro culturing of established cell lines led to increases in the cell surface expression of MHC class I and ICAM-1 and secretion of IL-6 and TGF-β 2 . Furthermore, there were significant changes in the expression of MHC class I, MHC class II, B7-2, ICAM-1, and FasL when comparing ex vivo tumor cells to those after a single passage in vitro . After passaging once in vitro , there were also significant changes in the secretion of TGF-β 2 and IL-10. This report indicates that in vitro culturing leads to significant changes in both cell surface molecules and secreted cytokines, which are known to affect the ability of immune cells to initiate an anti-tumor immune response. These changes in the immunological phenotype of glioblastomas after in vitro culturing may in part explain the limited success of immunotherapeutic strategies against glioblastomas in human clinical trials.