Angiosuppressive and angiostimulatory effects exerted by synthetic partial sequences of endostatin.
OBJECTIVE
Endostatin, a peptide derived from proteolysis of collagen XVIII, is an endogenous inhibitor of angiogenesis and tumor growth. We have synthesized five peptide fragments designed to cover the whole length of the endostatin molecule (containing 40-50 amino acids each) with the aim of exploring the possibility that a specific sequence within the molecule might be responsible for its antiangiogenic effects.


METHODS
The five peptide sequences, termed fragments I, II, III, IV, and IVox, the latter bearing the original disulfide bond Cys(135)-Cys(165), were investigated for their effects on cultured endothelial cells, on enzyme activities related to angiogenesis, on tube formation in three-dimensional gel matrices, in vivo in the avascular rabbit cornea assay, and in an experimental tumor burden paradigm.


RESULTS
Both the fragment covering the COOH-terminal endostatin region, fragment IV, and particularly fragment IVox, retained the angioinhibitory effects of endostatin. Fragment IVox strongly inhibited endothelial cell migration and proliferation, in vitro tube formation, and in vivo angiogenesis, displaying a potency comparable with that of endostatin. When tested in vivo on tumor growth, fragment IVox demonstrated to be more effective than full-length endostatin. Unexpectedly, fragment III exhibited proangiogenic activity, increasing endothelial cell migration, producing neovascularization to an extent similar to that of vascular endothelial growth factor, and enhancing the angiogenic response to vascular endothelial growth factor in the cornea assay. Peptides encompassing the NH(2)-terminal region of endostatin (fragments I and II) had negligible effects on angiogenesis.


CONCLUSIONS
In view of these results, which show strikingly distinct profiles of endostatin fragments, we propose that the amino acid sequence of endostatin contains both angiosuppressive and angiostimulatory domains.
Morbidelli
Lucia
L
Pharmacology Section, Department of Molecular Biology, University of Siena, Siena, Italy.
Donnini
Sandra
S
Chillemi
Francesco
F
Giachetti
Antonio
A
Ziche
Marina
M
D020533:Angiogenesis Inhibitors / Q000138:chemical synthesis / Q000494:pharmacology*; D000818:Animals; D002417:Cattle; D002455:Cell Division / Q000187:drug effects; D002465:Cell Movement / Q000187:drug effects; D002478:Cells, Cultured; D003315:Cornea / Q000098:blood supply / Q000378:metabolism; D043169:Endostatins / Q000138:chemical synthesis / Q000494:pharmacology*; D004730:Endothelium, Vascular / Q000378:metabolism*; D005260:Female; D005347:Fibroblasts / Q000378:metabolism; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D009374:Neoplasms, Experimental / Q000098:blood supply* / Q000473:pathology; D009389:Neovascularization, Pathologic / Q000139:chemically induced* / Q000473:pathology / Q000517:prevention & control*; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000494:pharmacology*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt; D011817:Rabbits; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism
D020533:Angiogenesis Inhibitors; D043169:Endostatins; D010446:Peptide Fragments; D011518:Proto-Oncogene Proteins; D042461:Vascular Endothelial Growth Factor A; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt