ABSTRACT
OBJECTIVE: Previous studies have shown that oxidized products of PAPC (Ox-PAPC) regulate cell transcription of interleukin-8, LDL receptor, and tissue factor. This upregulation takes place in part through the activation of sterol regulatory element-binding protein (SREBP) and Erk 1/2. The present studies identify vascular endothelial growth factor receptor 2 (VEGFR2) as a major regulator in the activation of SREBP and Erk 1/2 in endothelial cells activated by Ox-PAPC. METHODS AND RESULTS: Ox-PAPC induced the phosphorylation of VEGFR2 at Tyr1175 in human aortic endothelial cells. Inhibitors and siRNA for VEGFR2 decreased the transcription of interleukin-8, LDL receptor, and tissue factor in response to Ox-PAPC and the activation of SREBP and Erk 1/2, which mediate this transcription. We provide evidence that the activation of VEGFR2 is rapid, sustained, and c-Src-dependent. CONCLUSIONS: These data point to a major role of VEGFR2 in endothelial regulation by oxidized phospholipids which accumulate in atherosclerotic lesions and apoptotic cells.
Subject(s)
Endothelium, Vascular/drug effects , Endothelium, Vascular/physiology , Phosphatidylcholines/pharmacology , Vascular Endothelial Growth Factor Receptor-2/physiology , Cell Line , Cells, Cultured , Enzyme Activation/drug effects , Humans , Interleukin-8/genetics , Interleukin-8/physiology , Mitogen-Activated Protein Kinase 3/genetics , Mitogen-Activated Protein Kinase 3/physiology , Phosphorylation/drug effects , RNA, Small Interfering/pharmacology , Receptors, LDL/genetics , Receptors, LDL/physiology , Sterol Regulatory Element Binding Proteins/genetics , Sterol Regulatory Element Binding Proteins/physiology , Thromboplastin/genetics , Thromboplastin/physiology , Transcription, Genetic/drug effects , Transcription, Genetic/physiologyABSTRACT
The success of targeting kinases in cancer with small molecule inhibitors has been tempered by the emergence of drug-resistant kinase domain mutations. In patients with chronic myeloid leukemia treated with ABL inhibitors, BCR-ABL kinase domain mutations are the principal mechanism of relapse. Certain mutations are occasionally detected before treatment, suggesting increased fitness relative to wild-type p210 BCR-ABL. We evaluated the oncogenicity of eight kinase inhibitor-resistant BCR-ABL mutants and found a spectrum of potencies greater or less than p210. Although most fitness alterations correlate with changes in kinase activity, this is not the case with the T315I BCR-ABL mutation that confers clinical resistance to all currently approved ABL kinase inhibitors. Through global phosphoproteome analysis, we identified a unique phosphosubstrate signature associated with each drug-resistant allele, including a shift in phosphorylation of two tyrosines (Tyr253 and Tyr257) in the ATP binding loop (P-loop) of BCR-ABL when Thr315 is Ile or Ala. Mutational analysis of these tyrosines in the context of Thr315 mutations demonstrates that the identity of the gatekeeper residue impacts oncogenicity by altered P-loop phosphorylation. Therefore, mutations that confer clinical resistance to kinase inhibitors can substantially alter kinase function and confer novel biological properties that may impact disease progression.