Improved Inhibitors of Trypanothione Reductase by Combination of Motifs: Synthesis, Inhibitory Potency, Binding Mode, and Antiprotozoal Activities

Trypanothione reductase (TR) is an essential enzyme in the trypanothione‐based redox metabolism of trypanosomatid parasites. This system is absent in humans and, therefore, offers a promising target for the development of selective new drugs against African sleeping sickness and Chagas' disease. Over the past two decades, a variety of nonpeptidic small‐molecule ligands of the parasitic enzyme were discovered. A current goal is to decipher the binding mode of these known inhibitors in order to optimize their structures. We analyzed the binding mode of recently reported 1‐(1‐(benzo[b]thiophen‐2‐yl)cyclohexyl)piperidine (BTCP) analogues using computer modeling methods. This led us to conclude that the analogues occupy a different region of the active site than the diaryl sulfide‐based class of inhibitors. A combination of the two motifs significantly increased affinity for the enzyme compared to the respective parent compounds. The newly synthesized conjugates exhibit Kic values for TR as low as 0.51±0.1 μM and high selectivity for the parasitic enzyme over the related human glutathione reductase (hGR), as was predicted by our molecular modeling studies. In vitro studies showed IC50 values in the low micromolar to submicromolar range against Trypanosoma brucei rhodesiense, often in combination with low cytotoxicity against mammalian cells. Interestingly, even stronger activities were found against Plasmodium falciparum.

[1]  J. Coura,et al.  Chagas disease: a new worldwide challenge , 2010, Nature.

[2]  A. Fairlamb,et al.  Comparative structural, kinetic and inhibitor studies of Trypanosoma brucei trypanothione reductase with T. cruzi☆ , 2010, Molecular and biochemical parasitology.

[3]  F. Diederich,et al.  Synthesis, Inhibition Potency, Binding Mode, and Antiprotozoal Activities of Fluorescent Inhibitors of Trypanothione Reductase Based on Mepacrine‐Conjugated Diaryl Sulfide Scaffolds , 2009, ChemMedChem.

[4]  I. Gilbert,et al.  Synthesis and Evaluation of 1-(1-(Benzo[b]thiophen-2-yl)cyclohexyl)piperidine (BTCP) Analogues as Inhibitors of Trypanothione Reductase , 2009, ChemMedChem.

[5]  Carlos A. Montanari,et al.  Docking and molecular dynamics simulation of quinone compounds with trypanocidal activity , 2009, Journal of molecular modeling.

[6]  F. Diederich,et al.  Pentafluorosulfanyl as a Novel Building Block for Enzyme Inhibitors: Trypanothione Reductase Inhibition and Antiprotozoal Activities of Diarylamines , 2009, Chembiochem : a European journal of chemical biology.

[7]  F. Diederich,et al.  Diaryl sulfide-based inhibitors of trypanothione reductase: inhibition potency, revised binding mode and antiprotozoal activities. , 2008, Organic & biomolecular chemistry.

[8]  Simon Croft,et al.  Kinetoplastids: related protozoan pathogens, different diseases. , 2008, The Journal of clinical investigation.

[9]  A. Zhang,et al.  Regioselective Synthesis of 1-(2,6-Dichloro-4-Trifluoromethyl-phenyl)-4-Alkyl-1H-[1,2,3]-Triazoles , 2008, Molecules.

[10]  Chang H. Park,et al.  Design of new potent and selective secretory phospholipase A2 inhibitors. Part 5: synthesis and biological activity of 1-alkyl-4-[4,5-dihydro-1,2,4-[4H]-oxadiazol-5-one-3-ylmethylbenz-4'-yl(oyl)] piperazines. , 2008, Bioorganic & medicinal chemistry.

[11]  I. Gilbert,et al.  Target assessment for antiparasitic drug discovery. , 2007, Trends in parasitology.

[12]  W. Hunter.  Picking pockets to fuel antimicrobial drug discovery. , 2007, Biochemical Society transactions.

[13]  R. Gust,et al.  Structure-activity relationship study to understand the estrogen receptor-dependent gene activation of aryl- and alkyl-substituted 1H-imidazoles. , 2007, Journal of medicinal chemistry.

[14]  A. Fairlamb,et al.  Drug Resistance in Leishmaniasis , 2006, Clinical Microbiology Reviews.

[15]  V. Yardley,et al.  Antitrypanosomal, antileishmanial, and antimalarial activities of quaternary arylalkylammonium 2-amino-4-chlorophenyl phenyl sulfides, a new class of trypanothione reductase inhibitor, and of N-acyl derivatives of 2-amino-4-chlorophenyl phenyl sulfide. , 2005, Journal of medicinal chemistry.

[16]  R. L. Krauth-Siegel,et al.  Dithiolproteine als Hüter des intrazellulären Redoxmilieus bei Parasiten: alte und neue Wirkstoff‐Targets bei Trypanosomiasis und Malaria , 2005 .

[17]  R. Schirmer,et al.  Dithiol proteins as guardians of the intracellular redox milieu in parasites: old and new drug targets in trypanosomes and malaria-causing plasmodia. , 2005, Angewandte Chemie.

[18]  A. Fairlamb,et al.  Two Interacting Binding Sites for Quinacrine Derivatives in the Active Site of Trypanothione Reductase , 2004, Journal of Biological Chemistry.

[19]  A. Fairlamb.  Chemotherapy of human African trypanosomiasis: current and future prospects. , 2003, Trends in parasitology.

[20]  R. Krauth-Siegel,et al.  Parasite-specific trypanothione reductase as a drug target molecule , 2003, Parasitology Research.

[21]  A. Fairlamb,et al.  Ellman's-reagent-mediated regeneration of trypanothione in situ: substrate-economical microplate and time-dependent inhibition assays for trypanothione reductase. , 2003, The Biochemical journal.

[22]  M. Barrett,et al.  Chemotherapy of human African trypanosomiasis. , 2002, Current pharmaceutical design.

[23]  D. Marks,et al.  Use of an additional hydrophobic binding site, the Z site, in the rational drug design of a new class of stronger trypanothione reductase inhibitor, quaternary alkylammonium phenothiazines , 2000 .

[24]  F. Diederich,et al.  Tetrakis(phenylamidinium)-Substituted Resorcin[4]arene Receptors for the Complexation of Dicarboxylates and Phosphates in Protic Solvents , 2000 .

[25]  M. Barrett.  The fall and rise of sleeping sickness , 1999, The Lancet.

[26]  M. Berriman,et al.  Crystal structure of Trypanosoma cruzi trypanothione reductase in complex with trypanothione, and the structure-based discovery of new natural product inhibitors. , 1999, Structure.

[27]  A. Fairlamb,et al.  Phenothiazine inhibitors of trypanothione reductase as potential antitrypanosomal and antileishmanial drugs. , 1998, Journal of medicinal chemistry.

[28]  E. Davioud‐Charvet,et al.  New spermine and spermidine derivatives as potent inhibitors of Trypanosoma cruzi trypanothione reductase. , 1997, Bioorganic & medicinal chemistry.

[29]  S. J. Simpson,et al.  Glutathione reductase turned into trypanothione reductase: structural analysis of an engineered change in substrate specificity. , 1997, Biochemistry.

[30]  P. Karplus,et al.  Charge is the major discriminating factor for glutathione reductase versus trypanothione reductase inhibitors. , 1996, Bioorganic & medicinal chemistry.

[31]  W. Kabsch,et al.  Crystal structure of the Trypanosoma cruzi trypanothione reductase·mepacrine complex , 1996, Proteins.

[32]  C. Sergheraert,et al.  2-Amino diphenylsulfides as new inhibitors of trypanothione reductase. , 1995, International journal of antimicrobial agents.

[33]  Paul R. Gerber,et al.  MAB, a generally applicable molecular force field for structure modelling in medicinal chemistry , 1995, J. Comput. Aided Mol. Des..

[34]  B. de Costa,et al.  Synthesis and biological evaluation of 1-[1-(2-benzo[b]thienyl)cyclohexyl]piperidine homologues at dopamine-uptake and phencyclidine- and sigma-binding sites. , 1993, Journal of Medicinal Chemistry.

[35]  A. Fairlamb,et al.  Metabolism and functions of trypanothione in the Kinetoplastida. , 1992, Annual review of microbiology.

[36]  A. Fairlamb,et al.  Rationally designed selective inhibitors of trypanothione reductase. Phenothiazines and related tricyclics as lead structures. , 1992, The Biochemical journal.

[37]  C. George,et al.  Synthesis of isothiocyanato-1-[1-(2-benzo[b]thienyl)cyclohexyl]piperidines, potential irreversible ligands at the dopamine re-uptake site , 1992 .

[38]  Jockers-Scherübl Mc,et al.  Trypanothione reductase from Trypanosoma cruzi. Catalytic properties of the enzyme and inhibition studies with trypanocidal compounds. , 1989 .