A CYCLIC HEXAPEPTIDE IS A POTENT ANTAGONIST OF ALPHA(4) INTEGRINS

The alpha(4) integrins mediate leukocyte adhesion to specific counter- receptors, including vascular cell adhesion molecule-1 (VCAM-1), the f ibronectin splice variant containing connecting segment 1 (CS1), and m ucosal addressin cell adhesion molecule-1. A series of cyclized peptid es based on the LDV sequence of CS1 were synthesized and assayed for i nhibition of alpha(4) integrin binding. The most potent peptide, CWLD VC (where * indicates disulfide-linked residues), inhibited alpha(4) beta(1)-dependent binding of lymphocytes to VCAM-1. and CS1 with half- maximal inhibition achieved at 1 to 3 mu M of peptide. The peptide pro ved more potent when the lymphocytes were activated with 1 mM MnCl2; h alf-maximal inhibition was reached at 0.4 and 0.05 mu M for VCAM-1 and CS1, respectively. This represents a 100- to 800-fold increase in pot ency over a linear CS1 peptide in these same assays. CWLDVC* also inh ibited alpha(4) beta(7)-dependent lymphocyte binding to the ligands mu cosal addressin cell adhesion molecule-1, VCAM-1 and CS1. Immunoprecip itation of radiolabeled integrin indicated that the peptide could bind alpha(4) beta(1) and alpha(4) beta(7) directly and elute alpha(4) bet a(1) from a CS1-conjugated agarose resin. The peptide showed selectivi ty for alpha(4) integrins in that it effectively inhibited alpha(4) be ta(1)-dependent, but not alpha(5) beta(1)-dependent, binding of cells to intact fibronectin. Due to its small size and potency, CWLDVC* may serve as a useful tool for the study of alpha(4) integrin biology and the development of small molecule therapeutics.