N-Methylation of Peptides: A New Perspective in Medicinal Chemistry

Abstract

The potential of peptides as drug candidates is limited by their poor pharmacokinetic properties. Many peptides have a short half-life in vivo and a lack of oral availability. Inspired by the excellent pharmacokinetic profile of cyclosporine, a natural, multiply N-methylated cyclic peptide, we envisioned multiple N-methylation as a promising way to rationally improve key pharmacokinetic characteristics. In this Account, we summarize our efforts toward modulating the properties of peptides by multiple N-methylation. As a first step, we simplified the synthesis of N-methylated amino acids in solution, by employing very mild conditions that could be tolerated by the diverse protecting groups required when working with naturally occurring amino acids. We also report the rapid and inexpensive syntheses of N-methylated peptides on a solid support; this facilitated the N-methyl scanning of bioactive peptides. Because of a lack of information regarding the conformational behavior of multiply N-methylated peptides, a complete library of N-methylated cyclic alanine pentapeptides was synthesized. The library provided valuable insight into the conformational modulation of cyclic peptides by N-methylation. This information is extremely valuable for the design of bioactive peptides and spatial screening of cyclic N-methylated peptides. To demonstrate the applicability of N-methylation to highly active but poorly bioavailable peptides, we performed a full N-methyl scan of the cyclopeptidic somatostatin analog cyclo(-PFwKTF-), known as the Veber-Hirschmann peptide. We show here for the first time that the simple approach of multiple N-methylation can drastically improve the metabolic stability and intestinal permeability of peptides, for example, resulting in 10% oral bioavailability for a tri-N-methylated Veber-Hirschmann peptide analog. In addition, we also describe a designed approach to N-methylated peptide library synthesis, which can accelerate the screening of N-methylated bioactive peptides. Finally, we find that multiple N-methylation of a cyclic hexapeptide integrin antagonist of GPIIb-IIIa (alphaIIb beta3 integrin), cyclo(-GRGDfL-), increases the selectivity of this peptide toward different integrin subtypes. This result demonstrates the utility of multiple N-methylation in elucidating the bioactive conformation of peptides.

Peptidic modulators of protein‐protein interactions: Progress and challenges in computational design

“…17 It has become clear that proteinpeptide interactions are very abundant in the cell and estimates suggest they represent 15-40% of all interactions, 18 making peptide discovery and development of wide applicability and interest. Cyclic peptides [19][20][21][22] and other modified peptides 23,24 can be highly potent as well as selective, providing a compromise between structural preorganization and sufficient flexibility to mold to a target surface and maximize binding interactions. There is much evidence that despite having molecular masses well above Lipinski's rule constraints, cyclic peptides and other macrocycles can possess drug-like physicochemical and pharmacokinetic properties such as good solubility, lipophilicity, metabolic stability and bioavailability.…”

Evolution of multiple, mutually orthogonal prolyl-tRNA synthetase/tRNA pairs for unnatural amino acid mutagenesis in Escherichia coli

“…N-modified amino acids are known to impart unique properties to peptides, including distinctive conformational propensities to the peptide backbone, increased cellular permeability, and resistance to protease mediated degradation (9)(10)(11)(12)(13). The generation of an orthogonal suppressor prolyl-tRNA (tRNA Pro )/prolyl-tRNA synthase (ProRS) pair may facilitate the site-specific incorporation of UAAs with backbone modifications into proteins, which has been difficult to achieve using existing tRNA/aaRS pairs.…”

Mutant cycle analysis with modified saxitoxins reveals specific interactions critical to attaining high-affinity inhibition of hNa V 1.7

“…A collection of isomeric toxins bearing a methyl substituent group was targeted to ensure that relative differences in solvation energies between these compounds would be minimal. Variances in ligand affinity could thus be attributed to steric factors governing toxin-Na V interactions (32,33). Previous bis-guandinium toxinNa V mutant cycle analyses have been limited to naturally occurring derivatives (9,10).…”