Fragment-Based Drug Design Facilitated by Protein-Templated Click Chemistry: Fragment Linking and Optimization of Inhibitors of the Aspartic Protease Endothiapepsin

Abstract

There is an urgent need for the development of efficient methodologies that accelerate drug discovery. We demonstrate that the strategic combination of fragment linking/optimization and protein-templated click chemistry is an efficient and powerful method that accelerates the hit-identification process for the aspartic protease endothiapepsin. The best binder, which inhibits endothiapepsin with an IC₅₀ value of 43 μm, represents the first example of triazole-based inhibitors of endothiapepsin. Our strategy could find application on a whole range of drug targets.

Keywords: click chemistry; drug design; enzymes; inhibitors; liquid chromatography.

Figure 1

Schematic representation of protein‐templated click chemistry leading to a triazole‐based inhibitor starting from a library of azides and alkynes.

Figure 2

X‐ray crystal structure of endothiapepsin in complex with fragments 1 and 2 (PDB code: 3PBZ and 3PLD, respectively) and a modeled potential triazole inhibitor in the active site.36 Color code: protein skeleton: C: gray, O: red, and N: blue; fragment skeleton: C: purple, yellow and green, N: blue, O: red, Cl: green. Hydrogen bonds below 3.0 Å are shown as black, dashed lines.38

Scheme 1

a) Structures and retrosynthetic analysis of the designed triazole inhibitors starting from fragments 1 and 2; b) structures of the azides 3–11 and the alkynes 12–15.

Figure 3

Structure of the triazoles (17–20) identified using PTCC, inactive triazole 21.

Figure 4

Moloc‐generated modeled structures of: a) 17, and b) (S)‐18 in the active site of endothiapepsin. Color code: inhibitor skeleton: C: green, violet, N: blue, O: red; enzyme skeleton: C: gray. H bonds below 3.2 Å are shown as black, dashed lines.