Epitope‐Targeted Macrocyclic Peptide Ligand with Picomolar Cooperative Binding to Interleukin‐17F

The IL‐17 cytokine family is associated with multiple immune and autoimmune diseases and comprises important diagnostic and therapeutic targets. This work reports the development of epitope‐targeted ligands designed for differential detection of human IL‐17F and its closest homologue IL‐17A. Non‐overlapping and unique epitopes on IL‐17F and IL‐17A were identified by comparative sequence analysis of the two proteins. Synthetic variants of these epitopes were utilized as targets for in situ click screens against a comprehensive library of synthetic peptide macrocycles with 5‐mer variable regions. Single generation screens yielded selective binders for IL‐17F and IL‐17A with low cross‐reactivity. Macrocyclic peptide binders against two distinct IL‐17F epitopes were coupled using variable length chemical linkers to explore the physical chemistry of cooperative binding. The optimized linker length yielded a picomolar affinity binder, while retaining high selectivity. The presented method provides a rational approach towards targeting discontinuous epitopes, similar to what is naturally achieved by many B cell receptors. Cooperative binding: Macrocyclic peptide ligands were developed against distinct and discontinuous epitopes of two interleukin‐17 isoforms. These ligands achieved differential detection of the targeted isoform. Knowledge of the targeted epitopes informed a potentially general strategy for conjoining two ligands by a chemical linker to yield a cooperative biligand with picomolar binding affinity.