Abstract
Solid tumors contain hypoxic cells that are resistant to radiotherapy and chemotherapy. The resistance in glioblastoma has been linked to the expression of antiapoptotic Bcl-2 family members. In this study, we found that in human glioblastoma cells hypoxia induces the phosphorylation of the Bcl-2 family protein Bad, thus protecting hypoxic cells from paclitaxel-induced apoptosis. Akt activation is required for the hypoxia-induced protection. In contrast, the extracellular signal-regulated kinase 1/2 activities have only a partial effect, being able to modulate Bad phosphorylation but not paclitaxel-induced apoptosis in hypoxia. We also demonstrated that the degradation of adenosine with adenosine deaminase, the knockdown of A3 adenosine receptor expression by gene silencing, and the blockade of this receptor through A3 receptor antagonists blocked the hypoxia-induced phosphorylation of Bad and the prolonged cell survival after treatment with paclitaxel in hypoxia. Thus, the adenosinergic signaling may be an essential component in the hypoxia survival pathway. These results suggest that hypoxia-induced chemoresistance of human glioblastoma cells may occur in a novel mechanism involving activation of adenosine-A3 receptor-Akt pathway, which mediates Bad inactivation and favors cell survival.
Footnotes
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ABBREVIATIONS: GBM, glioblastoma multiforme; ADA, adenosine deaminase; Cl-IB-MECA, N6(3-iodobenzyl)2-chloroadenosine-5′N-methyluronamide; ERK, extracellular signal-regulated kinase; HIF-1α, hypoxia-inducible factor-1α; MEK, mitogen-activated protein kinase kinase; MRE 3008F20, 5N-(4-methoxyphenyl-carbamoyl)amino-8-propyl-2-(2-furyl)-pyrazolo-[4,3e]1,2,4-triazolo[1,5c] pyrimidine; siRNA, small interfering RNA; siRNAA3, small interfering RNA that targets A3 receptor mRNA; siRNABad, small interfering RNA that targets Bad; siRNAHIF-1α, siRNA targeting the transcription factor hypoxia-inducible factor-1α; PBS, phosphate-buffered saline; RT-PCR, reverse transcription-polymerase chain reaction; ANOVA, analysis of variance; PI3K, phosphatidylinositide-3-OH kinase; U0126, 1,4-diamino-2,3-dicyano-1,4-bis(methylthio)butadiene; LY294002, 2-(4-morpholinyl)-8-phenyl-1(4H)-benzopyran-4-one hydrochloride; SH-5, d-3-deoxy-2-O-methyl-myoinositol 1-[(R)-2-methoxy-3-(octadecyloxy)propyl hydrogen phosphate].
- Received October 17, 2006.
- Accepted March 30, 2007.
- The American Society for Pharmacology and Experimental Therapeutics
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