Mechanistic and biological characterisation of novelN⁵-substituted paullones targeting the biosynthesis of trypanothione inLeishmania

Abstract

Trypanothione synthetase (TryS) produces N 1 ,N 8-bis(glutathionyl)spermidine (or trypanothione) at the expense of ATP. Trypanothione is a metabolite unique and essential for survival and drug-resistance of trypanosomatid parasites. In this study, we report the mechanistic and biological characterisation of optimised N 5-substituted paullone analogues with anti-TryS activity. Several of the new derivatives retained submicromolar IC 50 against leishmanial TryS. The binding mode to TryS of the most potent paullones has been revealed by means of kinetic, biophysical and molecular modelling approaches. A subset of analogues showed an improved potency (EC 50 0.5-10 mM) and selectivity (20-35) against the clinically relevant stage of Leishmania braziliensis (mucocutaneous leishmaniasis) and L. infantum (visceral leishmaniasis). For a selected derivative, the mode of action involved intracellular depletion of trypanothione. Our findings shed light on the molecular interaction of TryS with rationally designed inhibitors and disclose a new set of compounds with on-target activity against different Leishmania species.

Drug-like molecules with anti-trypanothione synthetase activity identified by high throughput screening

“…In both cases, the new analogues retained their on-target effect against the pathogen as confirmed by genetic 9 or metabolic approaches 13 . Kinetic, thermodynamic and computational approaches suggested that paullones occupy the polyamine and second glutathione binding site of TryS 13 . Yet, the identification of multi-spectrum TryS inhibitors with reasonable biological activity and selectivity proved unsuccessful.…”

“…Inhibition of trypanothione synthesis either by genetic 6 , 7 or chemical 9 , 12 , 13 approaches leads to an intracellular oxidative milieu due to exhaustion of the antioxidant system to cope with the elimination of endogenous peroxides caused by trypanothione depletion.…”

“…The most potent compounds targeted Li TryS and belonged to the scaffold N 5 -acetamide substituted, 3-chlorokenpaullone ( Figure 1 ). A series of analogues containing different substituents at position N 5 of the 3-chlorokenpaullone were synthetised and yielded TryS hits that gained activity against the T. cruzi enzyme and infective stage of the parasite 9 , 26 or showed improved selectivity for the intracellular form of L. infantum 13 . In both cases, the new analogues retained their on-target effect against the pathogen as confirmed by genetic 9 or metabolic approaches 13 .…”

“…A series of analogues containing different substituents at position N 5 of the 3-chlorokenpaullone were synthetised and yielded TryS hits that gained activity against the T. cruzi enzyme and infective stage of the parasite 9 , 26 or showed improved selectivity for the intracellular form of L. infantum 13 . In both cases, the new analogues retained their on-target effect against the pathogen as confirmed by genetic 9 or metabolic approaches 13 . Kinetic, thermodynamic and computational approaches suggested that paullones occupy the polyamine and second glutathione binding site of TryS 13 .…”

Synthesis and Antibacterial Evaluation of 3,4‐Dihydro‐1H‐benzo[b]azepine‐2,5‐dione Derivatives

“…Compound VI showed superior activity with IC 50 0.3 μM against Li TryS , thus inhibiting L. braziliensis (EC 50 0.2-0.4 μM) and L. infantum (EC 50 0.9 � 0.1 μM) amastigotes with good selectivity index. [18] Choi et al, isolated Marinoazepinone B and demonstrated a 16 mm zone of inhibition against the Pontibacillus sp. [19] Arora et al, reported multi-step synthesis of new Quinolino annulated azepinones (VIII) and its antibacterial properties against E. coli with MIC 1 μg/mL.…”