Peptide Ligands Stabilized by Small Molecules

Abstract

Bicyclic peptides generated through directed evolution by using phage display offer an attractive ligand format for the development of therapeutics. Being nearly 100-fold smaller than antibodies, they promise advantages such as access to chemical synthesis, efficient diffusion into tissues, and needle-free application. However, unlike antibodies, they do not have a folded structure in solution and thus bind less well. We developed bicyclic peptides with hydrophilic chemical structures at their center to promote noncovalent intramolecular interactions, thereby stabilizing the peptide conformation. The sequences of the peptides isolated by phage display from large combinatorial libraries were strongly influenced by the type of small molecule used in the screen, thus suggesting that the peptides fold around the small molecules. X-ray structure analysis revealed that the small molecules indeed formed hydrogen bonds with the peptides. These noncovalent interactions stabilize the peptide-protein complexes and contribute to the high binding affinity.

The Interplay of Disulfide Bonds, α-Helicity, and Hydrophobic Interactions Leads to Ultrahigh Proteolytic Stability of Peptides

“…[7][8][9][10] Many strategies have thus been developed to improve resistance of peptides to proteolysis, including backbone/side-chain modification, unnatural residue substitution, and cyclization. [10][11][12][13][14] These approaches, however, can be synthetically laborious or awkward to implement. As a consequence, simple strategies to effectively stabilize peptides from enzymatic digestion that do not have recourse to unnatural amino acids, extensive post-translational chemistry, or formulation strategies are of exceptional interest.…”

Bridged Analogues for p53-Dependent Cancer Therapy Obtained by S-Alkylation

“…Notably, the use of multi-thiol-reactive linkers has a remarkably long tradition, as an application for this purpose of the aromatic derivative, 1,3,5-tris(bromomethyl)benzene, was initially reported by Kemp & McNamara in 1985 (Kemp and McNamara, 1985) and use of its bis-reactive homologs, 1,2-bis(bromomethyl)benzene, and 1,3-bis(bromomethyl)benzene was described only few years later (Szewczuk et al, 1992). This methodology was also successfully applied in peptide drug development (Timmerman et al, 2007), including phage display (Angelini et al, 2012a; Baeriswyl et al, 2012; Baeriswyl et al, 2013; Baeriswyl and Heinis, 2013a; Baeriswyl and Heinis, 2013b; Bellotto et al, 2014; Chen et al, 2012; Chen et al, 2013; Chen et al, 2014b; Chen et al, 2014a; Heinis et al, 2009; Rentero-Rebollo et al, 2014; Timmerman et al, 2007) as well as peptide-albumin (Angelini et al, 2012c; Pollaro et al, 2014) and peptide-antibody drug conjugates (ADCs) (Angelini et al, 2012b). …”

Small cyclic agonists of iron regulatory hormone hepcidin

“…In these harsh conditions the pentaerythritol scaffold remained intact proving that the proposed linker is relatively stable and perhaps require more attention, especially that it efficiently produced tricyclic, sterically hindered compound containing various bulky side chains in the vicinity of S- alkylation sites. Tricyclic analogs obtained via this method would be a logical extension of previously done work on therapeutic bicyclic analogs 31 offering potentially even larger diversity, especially if coupled with phage display approach 32–42 . Notably, the pentaerythritol scaffold was used previously for combinatorial chemistry 54 and in the synthesis of polymers and nanomaterials using an atom transfer radical polymerization (ATRP) 55;56 but not as an S- alkylating component of peptides’ constraining scaffold(s).…”

“…Moreover, all reactions can be carried out in solution without any protecting groups. Notably, this approach was already applied in peptide drug development 31 , including phage display 32–42 as well as peptide-albumin 43;44 and peptide-antibody drug conjugates (ADCs) 45 .…”

“…Such assumption is plausible since substitution of 3 His for fairly similar Phe or (D)His residues has limited effects on overall activity 60 and S-alkylation of Cys residue with aromatic linker(s) would produce substituted phenylalanine homolog(s) (see Figure 2). Except aromatic bridging moieties we also tested 2 linear and more flexible linkers afforded by bis -reactive Michael type compound DVS and 2-chloro-N-(2-(2-chloro-acetyloamino)-ethyl)-acetamide, Notably, other thiol(s)-multireactive Michael type compounds, including aromatic derivatives, were also reported 32;61 .…”