8-Methoxy-naphtho[2,3-b]thiophen-4,9-quinone, a non-competitive inhibitor of trypanothione reductase

Abstract

The enzyme trypanothione reductase is a recognised drug target in trypanosomatids and has been used in the search of new compounds with potential activity against diseases such as leishmaniasis, Chagas disease and African trypanosomiasis. thiophen-4,9-quinone was selected in a screening of natural and synthetic compounds using an in vitro assay with the recombinant enzyme from Trypanosoma cruzi. Its mode of inhibition fits a non-competitive model with respect to the substrate (trypanothione) and to the co-factor (NADPH),with respectively These parasites share a defence mechanism against oxidative stress based on the dithiol trypanothione [N 1 ,N 8 -bis(glutathionyl)-spermidine] and associated enzymes, specially trypanothione reductase (TR) (Fairlamb et al. 1985, Fairlamb & Cerami 1992, Flohe et al. 1999. This enzyme was shown to be essential for these parasites survival and infectivity and is recognised as an important target for the development of new drugs against these diseases (Fairlamb 1999, Iribarne et al. 2002.A lot of effort has been devoted to find TR inhibitors. Using either rational or empiric approaches, different classes of compounds, including peptides and peptoid, substituted polyamines, quinacrine and acridine analogues, 2-aminodiphenylsulfides, phenothiazine derivatives, substituted piperazines and nitrofuran derivatives, were studied for their inhibitory potential (see reviews by Werbovetz 2000, Augustyns et al. 2001, Schmidt & KrauthSiegel 2002. Investigation of natural products, although not so extensive, disclosed interesting inhibitors such as ajoene from garlic (Gallwitz et al. 1999), bisbenzylisoquinoline alkaloids (Fournet et al. 1998(Fournet et al. , 2000, and polyamines such as lunarine (Bond et al. 1999) and kukoamine (Ponasik et al. 1995).After the initial work of Henderson and co-workers (1988) naphthoquinone derivatives were further investigated for their action on TR (Jockers-Scherubl et al. 1989, Salmon-Chemin et al. 2000. In a previous work we reported the in vitro effect of several naphthothiophenquinone derivatives against epimastigote and trypomastigote forms of T. cruzi and the inhibition of the recombinant enzyme trypanothione reductase (Zani et al. 1997). We showed that quinone 8-Methoxy-naphtho [2,3-b]thiophen-4,9-quinone (TNQ2) was able to inhibit the enzyme activity by 87% at 100 µM after 30 min incubation. This compound was also one of the most active among 150 natural and synthetic compounds evaluated against TR (unpublished results). These findings stimulated us to carry out a more detailed investigation of the inhibition kinetics of this compound against TR and human glutathione reductase (hGR), the results of which are presented in this paper. [2,3-b]thiophen-4,9-quinoneThis compound was available from our previous work (Zani et al. 1997). Before use the material was recrystallized from MeOH-AcOEt to afford bright yellow needles with purity higher than 99.5%, as determined by reversed phase HPLC. A 100 mM stock solution was prepared in DMSO. This solution was dil...

Two Interacting Binding Sites for Quinacrine Derivatives in the Active Site of Trypanothione Reductase

“…Although TryR provides an attractive target for selective inhibition, purely competitive inhibitors of this enzyme may not be ideal as drug candidates, unless of low nanomolar affinity. Since there are extremely high concentrations of T[SH] 2 in the parasite cell (reaching 5 mM in L. donovani (43)), inhibition of TryR could result in the accumulation of sufficient T[S] 2 to partially reverse the effects of a competitive inhibitor (44). Indeed, more than 90% inhibition of TryR is required to kill T. brucei (45) and L. donovani can survive an 85% reduction in TryR activity with no effect upon either their viability or their ability to metabolise H 2 O 2 (8).…”

Inhibition of Disulfide Reductases as a Therapeutic Strategy

“…8-Methoxy-naphtho[2,3-b]thiophen-4,9-quinone inhibited a recombinant TryR enzyme from T. cruzi [97]. When tested against human Gor, this compound did not display any significant inhibition, indicating a good selectivity for the parasite enzymes [97].…”

“…8-Methoxy-naphtho[2,3-b]thiophen-4,9-quinone inhibited a recombinant TryR enzyme from T. cruzi [97]. When tested against human Gor, this compound did not display any significant inhibition, indicating a good selectivity for the parasite enzymes [97]. 1,4-Naphthoquinone derivatives with substituents in positions 2 and 3 inhibited TryR from T. cruzi and had potent antitrypanosomal activities in vitro in Trypanosoma brucei and T. cruzi cultures [98].…”

The Trypanocidal Activity of Naphthoquinones: A Review

“…Its mode of inhibition fits a non-competitive model with respect to the substrate (trypanothione) and to the co-factor (NADPH). In addition, when tested on human glutathione reductase, this compound did not display any significant inhibition, indicating a good selectivity for the parasite enzyme [ 118 ].…”