title: The Vascular Endothelial Growth Factor Inhibitors Ranibizumab and Aflibercept Markedly Increase Expression of Atherosclerosis-Associated Inflammatory Mediators on Vascular Endothelial Cells.
abstract: BACKGROUND
  Recent studies have suggested that the VEGF inhibitors, Ranibizumab and Aflibercept may be associated with an excess of cardiovascular events, potentially driven by increasing atheroma instability, leading to plaque rupture and clinical events. Inflammation plays a key role in the progression of atherosclerotic plaque and particularly conversion to an unstable phenotype. Here, we sought to assess the in vitro effects of these drugs on the expression of key inflammatory mediators on endothelial cells.
  
  
  METHODS
  Human coronary artery endothelial cells were co-incubated for 16h with Ranibizumab (0.11nM) or Aflibercept (0.45nM), as determined by each drug's peak serum concentration (Cmax). Expression at protein (ELISA) and gene (RT-PCR) level of inflammatory chemokines CCL2, CCL5 and CXC3L1 as well as gene expression for the cell adhesion molecules VCAM-1, ICAM-1 and the key NF-κb protein p65 was assessed. VEGF-A protein levels were also determined.
  
  
  RESULTS
  Both drugs significantly increased chemokine, cell adhesion molecule (CAM) and p65 expression, while decreasing VEGF-A protein secretion. At equivalent Cmax concentrations, Aflibercept was significantly more pro-inflammatory than Ranibizumab. Reduction of secreted VEGF-A levels significantly attenuated inflammatory effects of both drugs, whereas blockade of the VEGF-A receptor or silencing of VEGF-A gene synthesis alone had no effect, suggesting that binding of drug to secreted VEGF-A is crucial in promoting inflammation. Finally, blockade of Toll-like receptor 4 significantly reduced inflammatory effects of both drugs.
  
  
  CONCLUSIONS
  We demonstrated here, for the first time, that both drugs have potent pro-inflammatory effects, mediated via activation of Toll-like receptor 4 on the endothelial cell surface by drug bound to VEGF-A. Further studies are required to investigate whether these effects are also seen in vivo.
authors.affiliation: Department of Cardiology, Royal Prince Alfred Hospital, Sydney, New South Wales, Australia.|Sydney Medical School, The University of Sydney, New South Wales, Australia.
authors.forename: Clare
authors.initials: C
authors.lastname: Arnott
authors.affiliation: Heart Research Institute, Sydney, New South Wales, Australia.
authors.forename: Gaya
authors.initials: G
authors.lastname: Punnia-Moorthy
authors.affiliation: Sydney Medical School, The University of Sydney, New South Wales, Australia.|Heart Research Institute, Sydney, New South Wales, Australia.
authors.forename: Joanne
authors.initials: J
authors.lastname: Tan
authors.affiliation: Sydney Medical School, The University of Sydney, New South Wales, Australia.|Heart Research Institute, Sydney, New South Wales, Australia.
authors.forename: Sara
authors.initials: S
authors.lastname: Sadeghipour
authors.affiliation: Sydney Medical School, The University of Sydney, New South Wales, Australia.|Heart Research Institute, Sydney, New South Wales, Australia.
authors.forename: Christina
authors.initials: C
authors.lastname: Bursill
authors.affiliation: Department of Cardiology, Royal Prince Alfred Hospital, Sydney, New South Wales, Australia.|Sydney Medical School, The University of Sydney, New South Wales, Australia.|Heart Research Institute, Sydney, New South Wales, Australia.
authors.forename: Sanjay
authors.initials: S
authors.lastname: Patel
mesh_terms: D050197:Atherosclerosis / Q000235:genetics / Q000378:metabolism*; D018925:Chemokines / Q000235:genetics / Q000378:metabolism; D003331:Coronary Vessels / Q000473:pathology; D042783:Endothelial Cells / Q000187:drug effects / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D018836:Inflammation Mediators / Q000378:metabolism*; D008024:Ligands; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D000069579:Ranibizumab / Q000494:pharmacology*; D040262:Receptors, Vascular Endothelial Growth Factor / Q000494:pharmacology*; D011993:Recombinant Fusion Proteins / Q000494:pharmacology*; D051197:Toll-Like Receptor 4 / Q000378:metabolism; D042461:Vascular Endothelial Growth Factor A / Q000037:antagonists & inhibitors* / Q000378:metabolism; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000378:metabolism
chemical_list: D018925:Chemokines; D018836:Inflammation Mediators; D008024:Ligands; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D051197:Toll-Like Receptor 4; D042461:Vascular Endothelial Growth Factor A; C533178:aflibercept; D040262:Receptors, Vascular Endothelial Growth Factor; D040301:Vascular Endothelial Growth Factor Receptor-2; D000069579:Ranibizumab