ABSTRACT
PURPOSE: To identify mediators of glioblastoma antiangiogenic therapy resistance and target these mediators in xenografts. EXPERIMENTAL DESIGN: We conducted microarray analysis comparing bevacizumab-resistant glioblastomas (BRG) with pretreatment tumors from the same patients. We established novel xenograft models of antiangiogenic therapy resistance to target candidate resistance mediator(s). RESULTS: BRG microarray analysis revealed upregulation versus pretreatment of receptor tyrosine kinase c-Met, which underwent further investigation because of its prior biologic plausibility as a bevacizumab resistance mediator. BRGs exhibited increased hypoxia versus pretreatment in a manner correlating with their c-Met upregulation, increased c-Met phosphorylation, and increased phosphorylation of c-Met-activated focal adhesion kinase and STAT3. We developed 2 novel xenograft models of antiangiogenic therapy resistance. In the first model, serial bevacizumab treatment of an initially responsive xenograft generated a xenograft with acquired bevacizumab resistance, which exhibited upregulated c-Met expression versus pretreatment. In the second model, a BRG-derived xenograft maintained refractoriness to the MRI tumor vasculature alterations and survival-promoting effects of bevacizumab. Growth of this BRG-derived xenograft was inhibited by a c-Met inhibitor. Transducing these xenograft cells with c-Met short hairpin RNA inhibited their invasion and survival in hypoxia, disrupted their mesenchymal morphology, and converted them from bevacizumab-resistant to bevacizumab-responsive. Engineering bevacizumab-responsive cells to express constitutively active c-Met caused these cells to form bevacizumab-resistant xenografts. CONCLUSION: These findings support the role of c-Met in survival in hypoxia and invasion, features associated with antiangiogenic therapy resistance, and growth and therapeutic resistance of xenografts resistant to antiangiogenic therapy. Therapeutically targeting c-Met could prevent or overcome antiangiogenic therapy resistance.
Subject(s)
Angiogenesis Inhibitors/pharmacology , Drug Resistance, Neoplasm , Neovascularization, Pathologic/genetics , Proto-Oncogene Proteins c-met/genetics , Proto-Oncogene Proteins c-met/metabolism , Transcriptome , Angiogenesis Inhibitors/therapeutic use , Animals , Antibodies, Monoclonal, Humanized/pharmacology , Antibodies, Monoclonal, Humanized/therapeutic use , Bevacizumab , Cell Line, Tumor , Cell Survival/drug effects , Cell Survival/genetics , Cluster Analysis , Drug Resistance, Neoplasm/genetics , Enzyme Activation/genetics , Gene Expression Regulation, Neoplastic , Glioblastoma/drug therapy , Glioblastoma/genetics , Glioblastoma/metabolism , Glioblastoma/mortality , Humans , Mice , Neoplasm Invasiveness/genetics , Neovascularization, Pathologic/drug therapy , Proto-Oncogene Proteins c-met/antagonists & inhibitors , RNA Interference , Xenograft Model Antitumor AssaysABSTRACT
The precise subunit composition of synaptic ionotropic receptors in the brain is poorly understood. This information is of particular importance with regard to AMPA-type glutamate receptors, the multimeric complexes assembled from GluA1-A4 subunits, as the trafficking of these receptors into and out of synapses is proposed to depend upon the subunit composition of the receptor. We report a molecular quantification of synaptic AMPA receptors (AMPARs) by employing a single-cell genetic approach coupled with electrophysiology in hippocampal CA1 pyramidal neurons. In contrast to prevailing views, we find that GluA1A2 heteromers are the dominant AMPARs at CA1 cell synapses (approximately 80%). In cells lacking GluA1, -A2, and -A3, synapses are devoid of AMPARs, yet synaptic NMDA receptors (NMDARs) and dendritic morphology remain unchanged. These data demonstrate a functional dissociation of AMPARs from trafficking of NMDARs and neuronal morphogenesis. This study provides a functional quantification of the subunit composition of AMPARs in the CNS and suggests novel roles for AMPAR subunits in receptor trafficking.
