Immunobiological Characterization of Cancer Stem Cells Isolated from Glioblastoma Patients

Abstract

Purpose: Cancer stem cells (CSC) have been isolated from human tumors, including glioblastoma multiforme (GBM). The aims of this study were the immunobiological characterization of GBM CSCs and the assessment of whether these cells represent suitable targets for immunotherapy.Experimental Design: GBM CSC lines and their fetal bovine serum (FBS)-cultured non-CSC pair lines were generated and examined by flow cytometry for expression of known tumor antigens, MHC-I and MHC-II molecules, antigen-processing machinery components, and NKG2D ligands. In addition, immunogenicity and immunosuppression of such cell lines for autologous or allogeneic T lymphocytes were tested by cytokine secretion (ELISPOT) or proliferation (carboxyfluorescein diacetate succinimidyl ester) assays, respectively.Results: Both GBM CSC and FBS lines were weakly positive and negative for MHC-I, MHC-II, and NKG2D ligand molecules, respectively. Antigen-processing machinery molecules were also defective in both cell types. Upregulation of most molecules was induced by IFNs or 5-Aza deoxycytidine, although more efficiently in FBS than in CSCs. Patient T-cell responses, mediated by both TH1 and the TH2 subsets, against autologous CSC could be induced in vitro. In addition, CSC but not their paired FBS tumor lines inhibited T-cell proliferation of healthy donors. Notably, a differential gene signature that was confirmed at the protein levels for some immunologic-related molecules was also found between CSC and FBS lines.Conclusions: These results indicate lower immunogenicity and higher suppressive activity of GBM CSC compared with FBS lines. The immunogenicity, however, could be rescued by immune modulation leading to anti-GBM T cell-mediated immune response. Clin Cancer Res; 16(3); 800-13. ©2010 AACR.

Cancer-Initiating Cells from Colorectal Cancer Patients Escape from T Cell–Mediated Immunosurveillance In Vitro through Membrane-Bound IL-4

“…The MHC class I and class II typing of the patients was performed on their PBMCs by single-stranded oligonucleotide probe-PCR typing (10).…”

“…Part of the primary cell dissociation of CRC tissues was cultured in the presence of RPMI 1640 supplemented with 10% FBS (Biowittaker, Lonza, Treviglio, Italy), hereafter denominated FBS tumor cells (#1076, 1247, 111011, 1039, and 20299), and representing the differentiated tumor cell counterparts, as previously described (10,17,18); these cell lines were used in parallel with CICs. Additional CIC lines (#1, 2, and 3) isolated from primary CRC patients have been previously characterized (19).…”

“…The expression of Ag-processing machinery (APM) and MICA, MICB, and ULBPs molecules by these cell lines was determined using the previously described purified Abs provided by one of us (S.F.) and by Amgen (Thousand Oaks, CA), respectively (10). The expression of immunoregulatory molecules or cytokines and their receptors was assessed using the following Abs: anti-IL-4 3007 and anti-IL-4R 25463 (R&D Systems).…”

“…A possible explanation for these limited clinical results may lie in the failure to target CICs, a hypothesis corroborated by the evidence that an increase in "stemness" profile was obtained in postvaccinated glioblastoma patients and following antitumor T cell activity (8,9). We have previously characterized the immune profile of CICs isolated from glioblastoma multiforme (GBM) tissues and found low immunogenicity (10). Moreover, a preferential induction in vitro of Th2-type responses occurred following the coculture of CICs with autologous PBMCs (10).…”

“…We have previously characterized the immune profile of CICs isolated from glioblastoma multiforme (GBM) tissues and found low immunogenicity (10). Moreover, a preferential induction in vitro of Th2-type responses occurred following the coculture of CICs with autologous PBMCs (10). Cancer-testis Ags have been found to be expressed by mesenchymal stem cells and to regulate the epithelial-to-mesenchymal transition, a process involved in development of metastases; thus these Ags may possibly have a role in the stemness properties of cancer cells and could represent target molecules for immunotherapy (11).…”