Chemically synthesized matrix metalloproteinase and angiogenesis-inhibiting peptides as anticancer agents.

By connection of Regasepin2, a heptapeptide inhibitor of matrix metalloproteinases (MMPs), to the N- or C-terminus of ES-2, an anti-angiogenic peptide of 11 residues, two designed peptides CPU1 and CPU2 were generated. Unexpectedly, CPU2 inhibited MMP-8 and MMP-9 activity in the nanomolar range, whereas CPU1 displayed a weaker inhibitory profile than Regasepin2 against TACE, MMP-8 and MMP-9. CPU1 showed a higher affinity than CPU2 with integrin α5β1 in a HUVEC adhesion assay. In an in vitro angiogenesis model of HUVEC migration, CPU1 showed higher inhibition potency than CPU2 (85% inhibition for CPU1 at a concentration of 0.8 μM versus 17% inhibition for CPU2 at the same concentration). In an in vivo angiogenesis model in chicken egg chorioallantoic membranes, 1.37 μM CPU1 showed a significant inhibition in the formation of new blood vessels, whereas CPU2 and Regasepin2 had no effect on angiogenesis. Furthermore, CPU1 significantly inhibited B16F10 melanoma growth in a syngeneic mouse model (inhibition rate of 56.91% by tumor weight analysis at a dose of 20 mg/kg/d), whereas CPU2 and Regasepin2 did not show any inhibitory effect. In view of recent findings that MMP-9 is pro-angiogenic, our results indicated that the combination of MMP inhibitors and anti-angiogenic peptides may generate novel molecules with potent in vivo anti-tumor effects.