Automated Rapid Synthesis of High-Purity Head-to-Tail Cyclic Peptides via a Diaminonicotinic Acid Scaffold

Cyclic peptides uniquely integrate the structural stability and target specificity of biologics with the synthetic versatility of small molecules. However, their chemical synthesis has historically lagged behind ribosomal production in both speed and efficiency. Herein, we present <i>CycloBot</i>, a robotic platform designed for fully automated, minute-scale assembly of cyclopeptides, approaching the throughput of ribosomal biosynthesis while accommodating both natural and non-natural amino acids. Central to this advancement is a diaminonicotinic acid (DAN) scaffold enabling "one-click", concomitant cyclization/cleavage head-to-tail cyclization directly on resin, affording macrocycles of varying sizes with yields up to 93%, crude purities exceeding 95%, and minimal epimerization. The platform's utility is exemplified by the rapid generation of a 20-member antimicrobial cyclic peptide library in 1 day, from which a lead candidate exhibited 100-fold enhanced antibacterial activity against <i>S. aureus</i> and <i>B. subtilis</i> relative to penicillin. By bridging the speed of ribosomal synthesis and the structural diversity accessible via chemical synthesis, this technology establishes a new paradigm for accelerated discovery and development of personalized cyclic peptide therapeutics.