2.2 CuAAC in Peptidomimetics and Protein Mimics

The tremendous recent developments in click chemistry, including the impressive developments of strain-promoted cycloaddition reagents, all started with the copper-catalyzed azide–alkyne cycloaddition (CuAAC) reaction conceived by Meldal et al. and Sharpless et al. This led to a revolution of extremely important applications in the chemical, biological, medical, and materials sciences. It is fair to state that, especially in the synthesis of multifunctional and complex small-to-large biomolecular constructs, CuAAC has been indispensable. This has been particularly evident in the area of peptides, peptidomimetics, and protein mimics. These biomolecules play key roles in the various peptide–peptide, peptide–protein, and protein–protein interactions that are involved in many diseases and disorders, and peptide-based therapeutics can be important in this context. However, it is often important to improve the bioactivity and overall stability, and modulate the spatial structure, of peptide-based therapeutics. The incorporation of the 1,4-disubstituted 1,2,3-triazole moiety as a non-native structural element using CuAAC is explored in this chapter. The resulting incorporated triazole moiety can lead to structural surrogates of the amide bond and disulfide bond. As a consequence, CuAAC can be utilized toward introducing conformational constraints and stabilizing secondary structures of α-helices, β-sheets/turns, or loop-like structures. In addition, CuAAC can be used to combine various peptide sequences with molecular scaffolds to develop protein mimics that can find applications as synthetic vaccines and antibodies.

Small and Simple, yet Sturdy: Conformationally Constrained Peptides with Remarkable Properties

The sheer size and vast chemical space (i.e., diverse repertoire and spatial distribution of functional groups) underlie peptides’ ability to engage in specific interactions with targets of various structures. However, the inherent flexibility of the peptide chain negatively affects binding affinity and metabolic stability, thereby severely limiting the use of peptides as medicines. Imposing conformational constraints to the peptide chain offers to solve these problems but typically requires laborious structure optimization. Alternatively, libraries of constrained peptides with randomized modules can be screened for specific functions. Here, we present the properties of conformationally constrained peptides and review rigidification chemistries/strategies, as well as synthetic and enzymatic methods of producing macrocyclic peptides. Furthermore, we discuss the in vitro molecular evolution methods for the development of constrained peptides with pre-defined functions. Finally, we briefly present applications of selected constrained peptides to illustrate their exceptional properties as drug candidates, molecular recognition probes, and minimalist catalysts.

Rational Design of CYP3A4 Inhibitors: A One-Atom Linker Elongation in Ritonavir-Like Compounds Leads to a Marked Improvement in the Binding Strength

Inhibition of the major human drug-metabolizing cytochrome P450 3A4 (CYP3A4) by pharmaceuticals and other xenobiotics could lead to toxicity, drug–drug interactions and other adverse effects, as well as pharmacoenhancement. Despite serious clinical implications, the structural basis and attributes required for the potent inhibition of CYP3A4 remain to be established. We utilized a rational inhibitor design to investigate the structure–activity relationships in the analogues of ritonavir, the most potent CYP3A4 inhibitor in clinical use. This study elucidated the optimal length of the head-group spacer using eleven (series V) analogues with the R1/R2 side-groups as phenyls or R1–phenyl/R2–indole/naphthalene in various stereo configurations. Spectral, functional and structural characterization of the inhibitory complexes showed that a one-atom head-group linker elongation, from pyridyl–ethyl to pyridyl–propyl, was beneficial and markedly improved Ks, IC50 and thermostability of CYP3A4. In contrast, a two-atom linker extension led to a multi-fold decrease in the binding and inhibitory strength, possibly due to spatial and/or conformational constraints. The lead compound, 3h, was among the best inhibitors designed so far and overall, the strongest binder (Ks and IC50 of 0.007 and 0.090 µM, respectively). 3h was the fourth structurally simpler inhibitor superior to ritonavir, which further demonstrates the power of our approach.

Encapsulation of Tricopper Cluster in a Protein-like Cavitand Enables Facile Redox Processes from CuICuICuI to CuIICuIICuII States

One-pot reaction of tris(2-aminoethyl)amine (TREN), [CuI (MeCN)4]PF6, and paraformaldehyde affords a mixedvalent [TREN4CuIICuICuI (3-OH)](PF6)3 complex. The macrocyclic azacryptand TREN4 contains four TREN motifs, three of which provide a bowl-shape binding pocket for the [Cu3(3-OH)]3+ core. The fourth TREN caps on top of the tricopper cluster to form a cavitand, imposing conformational constraints and preventing solvent interaction. Contrasting the limited redox capability of synthetic tricopper complexes reported so far, [TREN4CuIICuICuI (3-OH)](PF6)3 exhibits several reversible single-electron redox events. The distinct electrochemical behaviors of [TREN4CuIICuICuI (3-OH)](PF6)3 and its solvent-exposed analog [TREN3CuIICuIICuII (3-O)](PF6)4 suggest that isolation of tricopper core in a protein-like cavitand enables facile electron transfer, allowing potential application of synthetic tricopper complexes as redox catalysts. Indeed, the fully reduced [TREN4CuICuICuI (3- OH)](PF6)2 can reduce O2 under acidic conditions. The geometric constraints provided by the cavitand are reminiscent of Nature’s multicopper oxidases (MCOs). For the first time, a synthetic tricopper cluster was isolated and fully characterized at CuICuICuI (4a), CuIICuICuI (4b), and CuIICuIICuI (4c) state, providing structural and spectroscopic models for many intermediates in MCOs. Fast electron transfer rates (105 - 106 M −1 s −1 ) were observed for both CuICuICuI /CuIICuICuI and CuIICuICuI /CuIICuIICuI redox couples, approaching the rapid electron transfer rates of copper sites in MCO.

Encapsulation of Tricopper Cluster in a Protein-like Cavitand Enables Facile Redox Processes from CuICuICuI to CuIICuIICuII States

One-pot reaction of tris(2-aminoethyl)amine (TREN), [CuI (MeCN)4]PF6, and paraformaldehyde affords a mixedvalent [TREN4CuIICuICuI (3-OH)](PF6)3 complex. The macrocyclic azacryptand TREN4 contains four TREN motifs, three of which provide a bowl-shape binding pocket for the [Cu3(3-OH)]3+ core. The fourth TREN caps on top of the tricopper cluster to form a cavitand, imposing conformational constraints and preventing solvent interaction. Contrasting the limited redox capability of synthetic tricopper complexes reported so far, [TREN4CuIICuICuI (3-OH)](PF6)3 exhibits several reversible single-electron redox events. The distinct electrochemical behaviors of [TREN4CuIICuICuI (3-OH)](PF6)3 and its solvent-exposed analog [TREN3CuIICuIICuII (3-O)](PF6)4 suggest that isolation of tricopper core in a protein-like cavitand enables facile electron transfer, allowing potential application of synthetic tricopper complexes as redox catalysts. Indeed, the fully reduced [TREN4CuICuICuI (3- OH)](PF6)2 can reduce O2 under acidic conditions. The geometric constraints provided by the cavitand are reminiscent of Nature’s multicopper oxidases (MCOs). For the first time, a synthetic tricopper cluster was isolated and fully characterized at CuICuICuI (4a), CuIICuICuI (4b), and CuIICuIICuI (4c) state, providing structural and spectroscopic models for many intermediates in MCOs. Fast electron transfer rates (105 - 106 M −1 s −1 ) were observed for both CuICuICuI /CuIICuICuI and CuIICuICuI /CuIICuIICuI redox couples, approaching the rapid electron transfer rates of copper sites in MCO.

Synthesis of Medium-Sized Heterocycles by Transition-Metal-Catalyzed Intramolecular Cyclization

Medium-sized heterocycles (with 8 to 11 atoms) constitute important structural components of several biologically active natural compounds and represent promising scaffolds in medicinal chemistry. However, they are under-represented in the screening of chemical libraries as a consequence of being difficult to access. In particular, methods involving intramolecular bond formation are challenging due to unfavorable enthalpic and entropic factors, such as transannular interactions and conformational constraints. The present review focuses on the synthesis of medium-sized heterocycles by transition-metal-catalyzed intramolecular cyclization, which despite its drawbacks remains a straightforward and attractive synthesis strategy. The obtained heterocycles differ in their nature, number of heteroatoms, and ring size. The methods are classified according to the metal used (palladium, copper, gold, silver), then subdivided according to the type of bond formed, namely carbon–carbon or carbon–heteroatom.