Chemokines mediate leukocyte migration and homeostasis and are key targets in inflammatory diseases including atherosclerosis, cytokine storm, and chronic autoimmune disease. Chemokine redundancy and ensuing network robustness has frustrated therapeutic development. Salivary evasins from ticks bind multiple chemokines to overcome redundancy and are effective in several preclinical disease models. Their clinical development has not progressed because of concerns regarding potential immunogenicity, parenteral delivery, and cost. Peptides mimicking protein activity can overcome the perceived limitations of therapeutic proteins. Here we show that peptides possessing multiple chemokine-binding and anti-inflammatory activities can be developed from the chemokine-binding site of an evasin. We used hydrogen-deuterium exchange MS to map the binding interface of the evasin P672 that physically interacts with C-C motif chemokine ligand (CCL) 8 and synthesized a 16-mer peptide (BK1.1) based on this interface region in evasin P672. Fluorescent polarization and native MS approaches showed that BK1.1 binds CCL8, CCL7, and CCL18 and disrupts CCL8 homodimerization. We show that a BK1.1 derivative, BK1.3, has substantially improved ability to disrupt P672 binding to CCL8, CCL2, and CCL3 in an AlphaScreen assay. Using isothermal titration calorimetry, we show that BK1.3 directly binds CCL8. BK1.3 also has substantially improved ability to inhibit CCL8, CCL7, CCL2, and CCL3 chemotactic function in vitro We show that local as well as systemic administration of BK1.3 potently blocks inflammation in vivo Identification and characterization of the chemokine-binding interface of evasins could thus inspire the development of novel anti-inflammatory peptides that therapeutically target the chemokine network in inflammatory diseases.
J Biol Chem
10926 - 10939
C–C motif chemokine ligand (CCL), chemokine, chemotaxis, evasin, host–pathogen interaction, hydrogen exchange mass spectrometry, immune response, inflammation, innate immunity, mass spectrometry, peptide interaction, peptides, protein structure, protein–protein interaction, tick, Amino Acid Sequence, Animals, Anti-Inflammatory Agents, Chemokine CCL8, Dimerization, Humans, Mass Spectrometry, Peptides, Protein Binding, Protein Engineering, Receptors, Chemokine, Sequence Homology, Amino Acid, Ticks