Stimulation of TLR9 with CpG ODN enhances apoptosis of glioma and prolongs the survival of mice with experimental brain tumors

Abstract

Toll-like receptors (TLRs) recognize a set of conserved molecular structures, so called pathogen-associated molecular patterns, which allow them to sense and initiate innate and adaptive immune responses. In this study, we examined the expression of TLRs in both human and murine glioma. We then analyzed the change in TLR expression after treatment with synthetic phosphorothioate oligodeoxynucleotides (ODNs) containing unmethylated CpG dinucleotides (CpG ODNs), strong activators of both innate and adaptive immunity. In addition, we investigated the in vivo effect of CpG injection into C57BL/6 mice implanted with syngeneic GL261 glioma. Our results indicate that TLR9 is overexpressed in human and murine glioma cell lines and CpG stimulation prolongs the survival of mice with experimental brain tumors. CpGs induce TLR9 down-regulation, followed by apoptosis of GL261 cells in vitro as well as in vivo. Furthermore, the effects of CpG stimulation appear to enhance the antigen presenting capacity of microglia, shift the immune response toward CD8(+) T cells, and decrease the number of CD4(+)CD25(+) regulatory T cells. Taken together, our data support the role of CpG in glioma immunotherapy and provide a rationale for further clinical development of CpG therapy in patients with malignant glioma.

Transposon-based interferon gamma gene transfer overcomes limitations of episomal plasmid for immunogene therapy of glioblastoma

“…The results reported in the current study using CpG ODN 2006 are consistent with those of Andaloussi et al 25 using CpG ODN 1826 in the GL261 intracranial glioma model. Both studies found treatment with CpG did not result in sustained tumor regression or cure.…”

“…Although we cannot exclude this possibility, there was no difference in tumor-reactive lymphocytes measured by ELISPOT from IFN-g-treated mice, regardless of stimulation with parental GL261 or GL261-Luc cells (Figure 7b). Moreover, the survival of untreated animals bearing GL261-Luc tumors was nearly identical to that reported by Andaloussi et al 25 using the parental GL261 cell line. Regardless of these potential limitations, GL261-Luc may be useful to study the time dynamics of tumor regression in response to immunotherapy in vivo and in real-time.…”

“…19 CpG ODN has also been shown to have anti-glioma activity in rats bearing syngeneic intracranial glioma 24 and mice bearing GL261 glioma. 25 Based on these previous studies, we investigated if CpG ODN might be an effective adjuvant to IFN-g gene therapy.…”

TLR Ligands in the Local Treatment of Established Intracerebral Murine Gliomas

“…Expression profiling of RNA samples of the GL261 glioma cells used in this study showed that they express high levels of TLR3, significant levels of TLR2 and TLR4 mRNA, and low-to-undetectable levels of TLR5, TLR7, and TLR9 mRNA. The virtual absence of TLR9 mRNA is surprising, as other data have suggested the presence of TLR9 GL261 glioma cells (33). Although it is difficult to exclude differences in the GL261 glioma cells, we would like to emphasize that our RNA samples were treated with DNase I to avoid accidental amplification of contaminating genomic DNA.…”

Comparative Proteomics Study Reveals That Bacterial CpG Motifs Induce Tumor Cell Autophagy in Vitro and in Vivo

“…In addition, various kinds of tumor cells, including colon cancer cells, also express TLRs (11)(12)(13), and TLR9 agonists have been proposed to be involved in growth and survival of human myeloma cells (14,15) as well as in invasion of human astrocytoma, glioblastoma, and breast cancer cells in vitro (16). Inversely CpG-ODNs have been shown to have antiproliferative and proapoptotic effects in vitro in human lung (17), breast (18), prostate (19), and colon (20) cancer cells as well as in murine glioma cells (21). Hence CpG effects in non-immune cells, and particularly in tumor cells, are still controversial, and this is, however, a crucial point as various cancer gene therapy clinical trials involve plasmid DNA or CpG-ODNs.…”