title: The overexpression of genes of thiol metabolism contribute to drug resistance in clinical isolates of visceral leishmaniasis (kala azar) in India.
abstract: BACKGROUND
  Visceral leishmaniasis (VL), also called Kala Azar (KA) or black fever in India, claims around 20,000 lives every year. Chemotherapy remains one of the most important tools in the control of VL. Current chemotherapy for Kala Azar in India relies on a rather limited arsenal of drugs including sodium antimony gluconate and amphotericin B in addition to the very expensive drug miltefosine. Pentavalent antimonials have been used for more than half a century in the therapy of leishmaniasis as it is relatively safe and inexpensive, however, the spread of resistance to this drug is forcing clinicians in India to abandon this treatment. Consequently, improvement of antimonial chemotherapy has become a major challenging area of study by leishmaniacs worldwide. The alarming emergence of resistance to the commonly used antleishmanial drug, sodium antimony gluconate, in India, has led us to elucidate the resistance mechanism(s) in clinical isolates. Studies on laboratory mutants have shown that resistance to antimonials is highly dependent on thiol levels. The parasite evades cytotoxic effects of antimonial therapy by enhanced efflux of drug upon conjugation with thiols, through overexpressed membrane proteins belonging to the superfamily of ABC transporters.
  
  
  METHODS
  We have carried out functional studies to determine the activity of the efflux pumps in antimonial resistant clinical isolates collected from disease endemic areas in India and also carried out molecular characterization of thiol levels in these parasites.
  
  
  RESULTS
  Overexpression of the gene coding for γ glutamylcysteine synthetase was observed in these resistant clinical isolates thereby establishing that thiols represent the key determinants of antimonial resistance. The SbIII/thiol conjugates can be sequestered by ABC transporter multidrug resistance protein A (MRPA) into intracellular organelles or can be directly pumped out by an uncharacterized transporter.
  
  
  CONCLUSIONS
  Our studies investigating antimonial resistance in different L. donovani clinical isolates suggest that over functioning of MRP plays a role in generation of antimony resistance phenotype in some L. donovani clinical isolates.
authors.affiliation: Central Drug Research Institute, Jankipuram Extension, Lucknow, 226031, India. neeloo888@yahoo.com.
authors.forename: Neeloo
authors.initials: N
authors.lastname: Singh
authors.affiliation: Department of Pharmacology, Institute of Post Graduate Medical Education and Research, Kolkata, India. ilatimc@gmail.com.
authors.forename: Mitali
authors.initials: M
authors.lastname: Chatterjee
authors.affiliation: Banaras Hindu University, Varanasi, India. drshyamsundar@hotmail.com.
authors.forename: Shyam
authors.initials: S
authors.lastname: Sundar
mesh_terms: D000666:Amphotericin B / Q000627:therapeutic use; D000818:Animals; D000965:Antimony / Q000627:therapeutic use; D000981:Antiprotozoal Agents / Q000627:therapeutic use*; D004351:Drug Resistance / Q000235:genetics*; D005721:Glutamate-Cysteine Ligase / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D007194:India; D007893:Leishmania donovani / Q000235:genetics* / Q000502:physiology; D007898:Leishmaniasis, Visceral / Q000188:drug therapy / Q000469:parasitology*; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D010767:Phosphorylcholine / Q000031:analogs & derivatives / Q000627:therapeutic use; D015800:Protozoan Proteins / Q000235:genetics / Q000378:metabolism; D013438:Sulfhydryl Compounds / Q000032:analysis / Q000378:metabolism*
chemical_list: D000981:Antiprotozoal Agents; D026901:Membrane Transport Proteins; D015800:Protozoan Proteins; D013438:Sulfhydryl Compounds; D010767:Phosphorylcholine; C039128:miltefosine; D000666:Amphotericin B; D000965:Antimony; D005721:Glutamate-Cysteine Ligase