Anti-angiogenic properties of a sulindac analogue.
OBJECTIVE
Angiopoietins (Ang) are crucial for new blood vessel formation and exert their effects by acting on the Tie2 receptor. We have recently described a sulindac analogue 2-((1E,Z)-1-benzylidene-5-bromo-2-methyl-1H-inden-3-yl)acetic acid; termed C-18 from now onwards) that inhibits Tie2 receptor activity in kinase assays in vitro. Here, we have assessed the ability of C-18 to inhibit angiogenesis-related properties of endothelial cells and tested its selectivity for the Tie2 receptor.


METHODS
For in vitro experiments human umbilical vein endothelial cells (HUVEC) were used. Proliferation was measured using the MTT assay; migration assays were performed in a modified Boyden chamber and tube-like structure formation was determined on matrigel. The effects of C-18 in vivo were evaluated in the chicken chorioallantoic membrane (CAM).


RESULTS
Pre-treatment of HUVEC with C-18 blocked Ang-1-stimulated migration, but also abolished vascular endothelial cell growth factor (VEGF)- and fibroblast growth factor 2-induced responses. Incubation with C-18 inhibited serum-induced proliferation in a concentration-dependent manner; C-18 was, however, without effect on Ang-1-induced survival. In addition, we observed that C-18 did not inhibit ligand-induced receptor phosphorylation of Tie2 or VEGFR2. On the other hand, C-18 blocked activation of members of the mitogen-activated protein kinase family and of the Ser/Thr kinase Akt induced by both VEGF and Ang-1. Furthermore, incubation of CAMs with C-18 led to a dose-dependent inhibition of vascular length.


CONCLUSIONS
C-18 did not act as a Tie2 inhibitor, as originally thought, but rather inhibited growth factor-stimulated signalling pathways that regulate endothelial cell migration and potently reduces neovascularization in vivo.
Pyriochou
A
A
Laboratory of Molecular Pharmacology, Department of Pharmacy, University of Patras, Patras, Greece.
Tsigkos
S
S
Vassilakopoulos
T
T
Cottin
T
T
Zhou
Z
Z
Gourzoulidou
E
E
Roussos
C
C
Waldmann
H
H
Giannis
A
A
Papapetropoulos
A
A
D020533:Angiogenesis Inhibitors / Q000494:pharmacology*; D042683:Angiopoietin-1 / Q000037:antagonists & inhibitors; D000818:Animals; D002465:Cell Movement / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D002642:Chick Embryo; D049033:Chorioallantoic Membrane / Q000187:drug effects / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D042783:Endothelial Cells / Q000187:drug effects* / Q000378:metabolism; D016222:Fibroblast Growth Factor 2 / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D020928:Mitogen-Activated Protein Kinases / Q000187:drug effects / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D017346:Protein Serine-Threonine Kinases / Q000187:drug effects / Q000378:metabolism; D042787:Receptor, TIE-2 / Q000187:drug effects* / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D013467:Sulindac / Q000008:administration & dosage / Q000494:pharmacology / Q000031:analogs & derivatives; D014471:Umbilical Veins; D042461:Vascular Endothelial Growth Factor A / Q000187:drug effects / Q000378:metabolism
C526537:2-(1-benzylidene-5-bromo-2-methyl-1H-inden-3-yl)acetic acid; D020533:Angiogenesis Inhibitors; D042683:Angiopoietin-1; D042461:Vascular Endothelial Growth Factor A; D016222:Fibroblast Growth Factor 2; D013467:Sulindac; D042787:Receptor, TIE-2; D017346:Protein Serine-Threonine Kinases; D020928:Mitogen-Activated Protein Kinases