Discovery of Selective\nCyclic d‑Sulfopeptide\nLigands of the Chemokine CCL22 via Mirror-Image mRNA Display with\nGenetic Reprogramming

Chemokines are small proteins involved in recruiting\nleukocytes\nto sites of inflammation via interactions with specific cell surface\nreceptors. CCL22 is a chemokine known to play a critical role in inflammatory\ndiseases such as atopic dermatitis and asthma; inhibition of this\nchemokine therefore represents an attractive therapeutic strategy.\nHerein, we describe the discovery of cyclic d-sulfopeptide\ninhibitors of CCL22 identified through mirror-image mRNA display with\ngenetic reprogramming. Chemical synthesis of mirror-image d-CCL22 enabled screening of a cyclic peptide library comprised of\nall l-amino acids, with reprogramming of l-sulfotyrosine\nto mimic the presence of this post-translational modification on native\nchemokine receptors. Enriched macrocyclic peptides were prepared in\ntheir mirror-image d-form and assessed for binding against\nnative l-CCL22. The most potent ligand, a plasma-stable d-cyclic peptide bearing four d-sulfotyrosine residues,\nexhibited nanomolar affinity for CCL22, high selectivity over other\nchemokines, and nanomolar inhibition of CCL22 signaling through CCR4.\nThis work highlights the vast potential of mirror-image mRNA display\ntechnology for discovering proteolytically stable d-peptide\ninhibitors of protein–protein interactions relevant across\na range of therapeutic indications.