Structure-based design of potent and selective Leishmania N-myristoyltransferase inhibitors

Abstract

Inhibitors of Leishmania N-myristoyltransferase (NMT), a potential target for the treatment of leishmaniasis, obtained from a high-throughput screen, were resynthesized to validate activity. Crystal structures bound to Leishmania major NMT were obtained, and the active diastereoisomer of one of the inhibitors was identified. On the basis of structural insights, enzyme inhibition was increased 40-fold through hybridization of two distinct binding modes, resulting in novel, highly potent Leishmania donovani NMT inhibitors with good selectivity over the human enzyme.

Chart 1. Leishmania-Selective Hits from Screening of a Subset of the Pfizer Compound File

Scheme 1. Synthesis of 1

Reagents and conditions: (a) pyrrolidine, N-Boc-4-piperidone, EtOH, rt, 3 days, 80%; (b) NH₄HCO₂, EtOH, Pd/C, 2 h, 96%; (c) para-fluorophenylacetyl chloride, Et₃N, THF, 2 h, 92%; (d) 6 M HCl, IPA, 2 h, 43%.

Scheme 2. Synthesis of Aminoacylpyrrolidine 2

Reagents and conditions: (a) 9, TFA, DCM, 0 °C to rt, 24 h, 88%; (b) (i) 1-chloroethyl chloroformate, toluene, 110 °C, 3 h, (ii) MeOH, reflux, 30 min; (c) 12, EDCI, HOBt, DIPEA, DMF, 4 h, 55% over 2 steps; (d) LiBH₄, THF, 3 h, 71%; (e) TFA, DCM, 2 h, 31%.

Figure 1

Inhibitor 1 (blue) bound in the peptide binding pocket of LmNMT (green). PDB code: 4cgn.

Figure 2

Inhibitor 2a (pink) bound in the peptide binding pocket of LmNMT (green). MyrCoA in blue. PDB code: 4cgl.

Scheme 3. Synthesis of 2a and 2b

Reagents and conditions: (a) (i) oxalyl chloride, DCM, DMF, 0 °C, 30 min, (ii) 17, Et₃N, LiCl, 15 h, 29%; (b) 9, TFA, DCM, 0 °C to rt, 24 h, 18a 36%, 18b 25%; (c) CO(OMe)₂, NaOMe, DCM, 15 h, 10a 48%, 10b 40%.

Figure 3

Overlay of I 2b (cyan) and 2a (pink) bound in the peptide binding pocket of LmNMT obtained by alignment of the protein main chain atoms (distances in Å). PDB code for 2b: 4cyn.

Figure 4

(a) Overlay of binding modes of 1 (blue) and 2a (pink). (b) Proposed hybrid structures 19 and 20.

Scheme 4. Synthesis of 19 and 20

Reagents and conditions: (a) 9, TFA, DCM, 0 °C to rt, 24 h, 79%; (b) SnCl₂, EtOH, 2 h, 87% (c) R = H, Ac₂O, Et₃N, DCM, 2 h, 65%, R = pF-Ph para-fluorophenylacetyl chloride, Et₃N, DCM, 2 h, 42%; (d) (i) 1-chloroethyl chloroformate, toluene, 110 °C, 3 h, (ii) MeOH, reflux, 30 min; (e) EDCI, HOBt, DIPEA, 12, DMF, 4 h, R = H 41% over 2 steps, R = cpF-Ph 43% over 2 steps; (f) LiBH₄, THF, 3 h; (g) TFA, DCM, 2 h, R = H 61% over 2 steps, R = pF-Ph 34% over two steps.

Figure 5

Overlay of hits 1 (blue), 2 (pink), and hybrid 20 (white; PDB code 4cyo) bound to LmNMT obtained by alignment of the protein main chain atoms.

Scheme 5. Synthesis of Acid 35

Reagents and conditions: (a) (i) Meldrum’s acid, pyridine, DCM, 0 °C, 30 min then rt, 15 h, (ii) EtOH, reflux, 2 h, 70%; (b) NaBH₄, MeOH, 0 °C to rt, 1.5 h, 40%; (c) LiOH, MeOH/H₂O 99%.

Scheme 6. Synthesis of Alcohol 43

Reagents and conditions: (a) (i) oxalyl chloride, DCM, DMF, 0 °C, 30 min, (ii) 16, Et₃N, LiCl, 15 h, 45%; (b) 9, TFA, DCM, 0 °C to rt, 24 h, 38a 40%; (c) CO(OMe)₂, NaOMe, DCM, 15 h 36%; (d) SnCl₂, EtOH, 2 h, quantitative; (e) para-fluorophenylacetyl chloride, Et₃N, DCM, 2 h 55%; (f) (i) 1-chloroethyl chloroformate, toluene, 110 °C, 3 h, (ii) MeOH, reflux, 30 min; (g) DMC, 35, Et₃N, DCM, 15 h, 27% over 2 steps; (h) LiBH₄, THF, 2 h, 25%.