Antiproliferative and antitumor activities of D‐reverse peptides derived from the second type‐1 repeat of thrombospondin‐1

Abstract

The extracellular matrix glycoprotein thrombospondin‐1 (TSP1) inhibits angiogenesis, endothelial cell growth, motility and adhesion. Peptides from the type I repeats of TSPl mimic the adhesive and growth inhibitory activities of the intact protein and specifically interact with heparin and transforming growth factor‐β (TGFβ). To define the structural basis for the antiangiogenic activities of these peptides, we prepared analogs of the TSPl peptide KRFKQDGGWSHWSPWSSC. l‐forward, l‐reverse, and d‐reverse (retro‐inverso) analogs displayed identical activities for binding to heparin. demonstrating a lack of stereospecificity for heparin binding. The l‐reverse and d‐reverse peptides, however, had somewhat decreased abilities to activate latent TGFβ. Conjugation of the forward peptides through a C‐terminal thioether and the reverse peptides through an N‐terminal thioether to polysucrose abolished the adhesive activity of the peptides and enhanced their antiproliferative activities for endothelial and breast carcinoma cells stimulated by fibroblast growth factor‐2. Their antiproliferative activities were independent of latent TGFβ activation, because substitution of an Ala residue for the essential Phe residue in the TSPl type‐1 repeat peptide increased their potency for inhibiting TSPl binding to heparin and for inhibiting endothelial cell proliferation. Although the conjugated peptides were inactive in vivo, an unconjugated retro‐inverso analog of the native TSP peptide inhibited breast tumor growth in a mouse xenograft model. Thus, these TSP‐derived peptide analogs antagonize endothelial growth through their heparin‐binding activity rather than through activation of latent TGFβ or increasing cell adhesion. These stable analogs may therefore be useful as therapeutic inhibitors of angiogenesis stimulated by fibroblast growth factor‐2. © Munksgaard 1997.