Discovery of high affinity inhibitors of Leishmania donovani N-myristoyltransferase

Abstract

N-Myristoyltransferase (NMT) is a potential drug target in Leishmania parasites. Scaffold-hopping from published inhibitors yielded the serendipitous discovery of a chemotype selective for Leishmania donovani NMT; development led to high affinity inhibitors with excellent ligand efficiency. The binding mode was characterised by crystallography and provides a structural rationale for selectivity.

Fig. 1. NMT affinity profile for 2,3-substituted benzo[b]thiophene 1: ^aEnzyme apparent K _i values are calculated from the IC₅₀ values using the Cheng–Prusoff equation (see ESI†) to enable cross-comparisons. IC₅₀ values were the mean of two or more independent determinations. Standard deviation is within 20% of reported IC₅₀. ^bHsNMT affinities reported in this work refer to HsNMT1; no significant difference in inhibition was observed between HsNMT1 and HsNMT2 isoforms.

Scheme 1. Synthesis of truncated inhibitor 5. Reagents and conditions: (a) 1-Boc-4-piperidinol, DIAD, PPh₃. THF, rt, 4 h, 94%; (b) NaOH, MeOH/H₂O, 50 °C, 2 h, 90%; (c) (3-methoxyphenyl)methanol, EDCI, HOBt, DIPEA, MeCN, rt, 18 h; (d) 10% TFA in DCM (v/v), rt, 2 h, 9% over 2 steps.

Scheme 2. Synthesis of oxadiazole derivatives 10a–k. Reagents: (a) BnBr, K₂CO₃, DMF, rt, 2 h, 72–89%; (b) DIAD, PPh₃, N-Boc-4-OH piperidine, THF, rt, 4 h; (c) NaOH, MeOH/H₂O, 50 °C, 2 h, 80–95% over two steps; (d) NH₂OH, EtOH, 80 °C, 6 h, 98%; (e) i). EDCI, HOBt, 9, DIPEA, CH₃CN, rt, 4 h; ii). 0.5 N NaOH, rt, 0.5 h; (f) 10% TFA in DCM, rt, 2 h, 20–60% over 2 steps.

Scheme 3. Synthesis of pyrazole derivative 13. Reagents and conditions: (a) 1-Boc-4-piperidinol, DIAD, PPh₃. THF, rt, 4 h, 46%; (b) hydrazine monohydrate, EtOH, reflux, 18 h, quantitative; (c) EDCI, HOBt, 2-(trimethyl-1H-pyrazol-4-yl)acetic acid, DIPEA, THF : DMF (4 : 1, v/v), rt, 18 h; (d) TsCl, 1,2,2,6,6-pentamethylpiperidine, DCM, rt, 3 h; (e) 10% TFA in DCM (v/v), rt, 2 h, 40% over three steps.

Fig. 2. Crystal structure of 13 (blue) bound to LmNMT (grey). (A) The chloro-substituent is buried within a hydrophobic pocket, and appears to have good shape complementarity with the enzyme active site. (B) In addition to the shape-complementarity of the scaffold, the compound appears to form a similar binding mode to that previously observed. (C) The trimethylpyrazole motif forms both a hydrogen bonding interaction with Ser330 and a hydrophobic stacking interaction with Phe90.

Fig. 3. Comparison of the crystal structure of 1 (pink) bound to PvNMT (yellow) and 13 (blue) bound to LmNMT (grey). (A) Overlay of 1 in PvNMT and 13 in LdNMT, with the enzymes removed for clarity; the phenyl ring of the benzo[b]thiophene scaffold protrudes in the direction of potential 3- and 4-substitutions of the phenyl series. (B) The surface of the active site of LmNMT (grey) shows a constrained, water filled back pocket in the same position as that occupied by the benzo[b]thiophene. (C) Alanine to methionine substitution in LmNMT results in a more constrained pocket and a potential clash with the bulkier benzothiophene scaffold.