Drug discovery in leishmaniasis using protein lipidation as a target

Abstract

The leishmaniases are infectious diseases caused by a number of species of obligate intracellular protozoa of the genus Leishmania with disease manifesting as cutaneous, mucocutaneous and visceral forms. Despite being endemic in more than 80 countries and its being the cause of high morbidity and mortality, leishmaniasis remains a neglected tropical disease. Chemotherapy is the frontline treatment, but drugs in current use suffer from toxic side effects, difficulties in administration and extended treatment times - moreover, resistance is emerging. New anti-leishmanial drugs are a recognised international priority. Here, we review investigations into N-myristoyltransferase (NMT) as a potential drug target. NMT catalyses the co-translational transfer of a C₁₄ fatty acid from myristoyl-CoA onto the N-terminal glycine residue of a significant subset of proteins in eukaryotic cells. This covalent modification influences the stability and interactions of substrate proteins with lipids and partner proteins. Structure-guided development of new lead compounds emerging from high-throughput screening campaigns targeting Leishmania donovani NMT has led to the discovery of potent inhibitors which have been used to gain insights into the role of protein myristoylation in these parasites and to validate NMT as a drug target.

Keywords: Leishmaniasis inhibitor discovery; N-myristoyltransferase; Neglected tropical disease; Protein structure.

Fig. 1

Chemical structures of a selection of clinically important anti-leishmanial drugs in current use

Fig. 2

Structure and ligand binding to Leishmania NMT. A Scheme showing the ordered binding of substrates and the ordered release of products in the NMT-catalysed reaction. B Key step in the catalytic mechanism. Following binding of the substrates and deprotonation of the substrate protein’s α-amino group by the carboxylate of Leu421, there is nucleophilic attack of the amino group of the glycine on the carbonyl carbon of the thioester of myristoyl-CoA. C The crystal structure of the ternary complex of NMT from L. major (PDB code 4c7h) with myristoyl-CoA and a peptidomimetic inhibitor bound in the active site. The protein is represented as a ribbon colour-ramped from the N-terminus (blue) to the C-terminus (red). The C-terminus is additionally labelled with an asterisk. The ligands are shown in sphere representation with atoms coloured by type: carbon, grey for peptidomimetic and green for myristoyl-CoA; oxygen, red; nitrogen, blue; sulphur, yellow; phosphorus, magenta. D Lower, zoom view of C with electrostatic surface rendering of the protein molecule emphasising the substrate binding groove. Upper, chemical structure of the peptidomimetic inhibitor (N-(10-aminodecyl)-L-seryl-N-(2-cyclohexyl)-L-lysinamide). E Structures of inhibitor molecules described in the text. In the centre, these molecules are displayed following least squares superposition of the protein Cα atoms in the respective structures of the LmNMT-myristoyl-CoA inhibitor ternary complexes. The inhibitors occupy overlapping volumes in the peptide-binding cavity. The inhibitors are coloured: aminoacylpyrrolidine, grey; pyrazolyl sulfonamide, blue; piperidinylindole, coral; quinoline, ice blue; thienopyrimidine, green; biphenyl, red

Fig. 3

The structure of the inhibitor binding pocket and protein–inhibitor interactions. A The thienopyrimidine (PDB Entry, 4cgo), B the aminoacylpyrrolidine (4cgn) and C the piperidinylindole (4cgm) ligands are shown in cylinder representation with grey carbon atoms with the surrounding protein in ball-and-stick and green carbon atoms. The myristoyl-CoA cofactor, which is situated to the left of the ligand in each case, has been omitted for clarity. D Hybridisation of the binding modes of the aminoacylpyrrolidine (grey carbons) and piperidinylindole inhibitors (coral carbons) to generate a more potent hybrid NMT inhibitor (magenta carbons). Other atoms are coloured by type: oxygen, red; nitrogen, blue; chlorine, white; fluorine, cyan