Novel atypical PKC inhibitors prevent vascular endothelial growth factor-induced blood-retinal barrier dysfunction.
Pro-inflammatory cytokines and growth factors such as VEGF (vascular endothelial growth factor) contribute to the loss of the BRB (blood-retinal barrier) and subsequent macular oedema in various retinal pathologies. VEGF signalling requires PKCβ [conventional PKC (protein kinase C)] activity; however, PKCβ inhibition only partially prevents VEGF-induced endothelial permeability and does not affect pro-inflammatory cytokine-induced permeability, suggesting the involvement of alternative signalling pathways. In the present study, we provide evidence for the involvement of aPKC (atypical PKC) signalling in VEGF-induced endothelial permeability and identify a novel class of inhibitors of aPKC that prevent BRB breakdown in vivo. Genetic and pharmacological manipulations of aPKC isoforms were used to assess their contribution to endothelial permeability in culture. A chemical library was screened using an in vitro kinase assay to identify novel small-molecule inhibitors, and further medicinal chemistry was performed to delineate a novel pharmacophore. We demonstrate that aPKC isoforms are both sufficient and required for VEGF-induced endothelial permeability. Furthermore, these specific, potent, non-competitive, small-molecule inhibitors prevented VEGF-induced tight junction internalization and retinal endothelial permeability in response to VEGF in both primary culture and in rodent retina. The results of the present study suggest that aPKC inhibition with 2-amino-4-phenyl-thiophene derivatives may be developed to preserve the BRB in retinal diseases such as diabetic retinopathy or uveitis, and the BBB (blood-brain barrier) in the presence of brain tumours.
Department of Cellular and Molecular Physiology, Penn State University College of Medicine, Hershey, PA 17033, USA. pmt132@psu.edu
Paul M
PM
Titchenell
Cheng-Mao
CM
Lin
Jason M
JM
Keil
Jeffrey M
JM
Sundstrom
Charles D
CD
Smith
David A
DA
Antonetti
D000818:Animals; D001813:Blood-Retinal Barrier / Q000378:metabolism*; D002417:Cattle; D003930:Diabetic Retinopathy / Q000378:metabolism; D006801:Humans; D008297:Male; D020033:Protein Isoforms / Q000037:antagonists & inhibitors / Q000378:metabolism; D011493:Protein Kinase C / Q000037:antagonists & inhibitors* / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012160:Retina / Q000166:cytology / Q000378:metabolism; D012171:Retinal Vessels / Q000166:cytology / Q000378:metabolism; D015398:Signal Transduction; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics / Q000378:metabolism*
D020033:Protein Isoforms; D047428:Protein Kinase Inhibitors; D011994:Recombinant Proteins; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; D011493:Protein Kinase C