Abstract
BackgroundAnti-angiogenic therapy inhibits tumor growth and is considered as a potential clinical therapy for malignant glioma. However, inevitable recurrences and unexpected tumor resistance, particularly increased invasion ability of glioma cell, were observed after anti-angiogenic treatment. The underlying mechanism remains undetermined. Focal adhesion kinase (FAK) and proline-rich tyrosine kinase 2 (Pyk2) are closely associated with cell migration; therefore, we investigated the possible role of these kinases in rat C6 glioma cell invasion induced by bevacizumab, a recombinant monoclonal antibody against vascular endothelial growth factor (VEGF).MethodsThe effects of bevacizumab on migration and invasion of C6 glioma cells were investigated in vitro and in vivo. The cells proliferation, migration, and invasion were determined by MTT assay, wound healing, and transwell assay, respectively. Invasive potential of glioma cells in vivo was assessed by counting vimentin-positive cells crossing the solid tumor rim by immunohistochemical staining. The total and phosphorylated protein levels of FAK and Pyk2 were detected by Western blotting.ResultsBevacizumab exposure increased migration and invasion of cultured C6 cells in a concentration-dependent manner. In addition, the continuous bevacizumab treatment also promoted tumor invasion in rat C6 intracranial glioma models. Bevacizumab treatment enhanced Pyk2 phosphorylation at Tyr402, but no effect on FAK phosphorylation at Tyr397 both in vitro and in vivo. Knockdown of Pyk2 by siRNA or inhibition of Pyk2 phosphorylation by Src kinase specific inhibitor PP1 partially inhibited bevacizumab-induced cell invasion in cultured C6 glioma cells. Furthermore, the combined administration of bevacizumab and PP1 significantly suppressed glioma cell invasion into surrounding brain tissues compared to bevacizumab treatment alone in experimental rats.ConclusionsThese results suggest that anti-VEGF treatment promotes glioma cell invasion via activation of Pyk2. Inhibition of Pyk2 phosphorylation might be a potential target to ameliorate the therapeutic efficiency of anti-VEGF treatment.
Highlights
Anti-angiogenic therapy inhibits tumor growth and is considered as a potential clinical therapy for malignant glioma
Bevacizumab treatment promoted migration and invasion of glioma cell To exclude the possible contribution of an imbalance in cell proliferation and viability to cell migration and invasion after anti-vascular endothelial growth factor (VEGF) treatment, cell proliferation and viability under different concentrations (0, 2.5, 5 and 10 mg/ ml) of bevacizumab were measured by MTT assay before invasion experiment
The administration of 10 mg/kg bevacizumab for 3 weeks resulted in a significant increase in invasive tumor cells outside the tumor rim, visualized by vimentin staining (Figure 3), suggesting that anti-VEGF treatment induced an increased ability of cell migration and invasion both in cultured C6 cells and in intracranial C6 glioma cell xenograft
Summary
Anti-angiogenic therapy inhibits tumor growth and is considered as a potential clinical therapy for malignant glioma. Inevitable recurrences and unexpected tumor resistance, increased invasion ability of glioma cell, were observed after anti-angiogenic treatment. With the increasing accumulation of knowledge regarding angiogenesis, anti-angiogenic therapy has been developed and considered as an optimistic strategy for glioma patients [3,4]. With the angiogenesis-targeted therapy widely accepted, inevitable recurrence and unexpected tumor resistance, especially increased ability of glioma cell invasion were observed after anti-VEGF treatment [11,12,13,14,15,16]. Unexpected tumor resistance to bevacizumab aroused the additional investigation on direct effects of anti-VEGF therapy on tumor cell migration and invasion. The exact mechanism underlying tumor resistance to bevacizumab remains to be elucidated
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