ANTIPROLIFERATIVE AND ANTITUMOR ACTIVITIES OF D-REVERSE PEPTIDES DERIVED FROM THE 2ND-TYPE-1 REPEAT OF THROMBOSPONDIN-1

The extracellular matrix glycoprotein thrombospondin-1 (TSP1) inhibits angiogenesis, endothelial cell growth, motility and adhesion. Peptide s from the type I repeats of TSP1 mimic the adhesive and growth inhibi tory activities of the intact protein and specifically interact with h eparin and transforming growth factor-beta (TGF beta). To define the s tructural basis for the antiangiogenic activities of these peptides, w e prepared analogs of the TSP1 peptide KRFKQDGGWSHWSPWSSC. L-forward, L-reverse, and D-reverse (retro-inverso) analogs displayed identical a ctivities for binding to heparin, demonstrating a lack of stereospecif icity for heparin binding. The L-reverse and D-reverse peptides, howev er, had somewhat decreased abilities to activate latent TGF beta. Conj ugation of the forward peptides through a C-terminal thioether and the reverse peptides through an N-terminal thioether to polysucrose aboli shed the adhesive activity of the peptides and enhanced their antiprol iferative activities for endothelial and breast carcinoma cells stimul ated by fibroblast growth factor-2. Their antiproliferative activities were independent of latent TGF beta activation, because substitution of an Ala residue for the essential Phe residue in the TSP1 type-1 rep eat peptide increased their potency for inhibiting TSP1 binding to hep arin and for inhibiting endothelial cell proliferation, Although the c onjugated peptides were inactive in vivo, an unconjugated retro-invers o analog of the native TSP peptide inhibited breast tumor growth in a mouse xenograft model. Thus, these TSP-derived peptide analogs antagon ize endothelial growth through their heparin-binding activity rather t han through activation of latent TGF beta or increasing cell adhesion. These stable analogs may therefore be useful as therapeutic inhibitor s of angiogenesis stimulated by fibroblast growth factor-2. (C) Munksg aard 1997.