Fmoc-Based Synthesis of Disulfide-Rich Cyclic Peptides

Disulfide-rich cyclic peptides have exciting potential as leads or frameworks in drug discovery; however, their use is faced with some synthetic challenges, mainly associated with construction of the circular backbone and formation of the correct disulfides. Here we describe a simple and efficient Fmoc solid-phase peptide synthesis (SPPS)-based method for synthesizing disulfide-rich cyclic peptides. This approach involves SPPS on 2-chlorotrityl resin, cyclization of the partially protected peptide in solution, cleavage of the side-chain protecting groups, and oxidization of cysteines to yield the desired product. We illustrate this method with the synthesis of peptides from three different classes of cyclic cystine knot motif-containing cyclotides: Möbius (M), trypsin inhibitor (T), and bracelet (B). We show that the method is broadly applicable to peptide engineering, illustrated by the synthesis of two mutants and three grafted analogues of kalata B1. The method reduces the use of highly caustic and toxic reagents and is better suited for high-throughput synthesis than previously reported methods for producing disulfide-rich cyclic peptides, thus offering great potential to facilitate pharmaceutical optimization of these scaffolds

Cyclic Penta- and Hexaleucine Peptides without <i>N</i>‑Methylation Are Orally Absorbed

Development of peptide-based drugs has been severely limited by lack of oral bioavailability with less than a handful of peptides being truly orally bioavailable, mainly cyclic peptides with <i>N</i>-methyl amino acids and few hydrogen bond donors. Here we report that cyclic penta- and hexa-leucine peptides, with no <i>N</i>-methylation and five or six amide NH protons, exhibit some degree of oral bioavailability (4–17%) approaching that of the heavily <i>N</i>-methylated drug cyclosporine (22%) under the same conditions. These simple cyclic peptides demonstrate that oral bioavailability is achievable for peptides that fall outside of rule-of-five guidelines without the need for <i>N</i>-methylation or modified amino acids