title: Mechanism of amphotericin B resistance in clinical isolates of Leishmania donovani.
abstract: The clinical value of amphotericin B, the mainstay therapy for visceral leishmaniasis in sodium antimony gluconate-nonresponsive zones of Bihar, India, is now threatened by the emergence of acquired drug resistance, and a comprehensive understanding of the underlying mechanisms is the need of the hour. We have selected an amphotericin B-resistant clinical isolate which demonstrated 8-fold-higher 50% lethal doses (LD(50)) than an amphotericin B-sensitive strain to explore the mechanism of amphotericin B resistance. Fluorimetric analysis demonstrated lower anisotropy in the motion of the diphenylhexatriene fluorescent probe in the resistant strain, which indicated a higher fluidity of the membrane for the resistant strain than for the sensitive strain. The expression patterns of the two transcripts of S-adenosyl-l-methionine:C-24-Δ-sterol methyltransferase and the absence of ergosterol, replaced by cholesta-5,7,24-trien-3β-ol in the membrane of the resistant parasite, indicate a decreased amphotericin B affinity, which is evidenced by decreased amphotericin B uptake. The expression level of MDR1 is found to be higher in the resistant strain, suggesting a higher rate of efflux of amphotericin B. The resistant parasite also possesses an upregulated tryparedoxin cascade and a more-reduced intracellular thiol level, which helps in better scavenging of reactive oxygen species produced by amphotericin B. The resistance to amphotericin B was partially reverted by the thiol metabolic pathway and ABC transporter inhibitors. Thus, it can be concluded that altered membrane composition, ATP-binding cassette transporters, and an upregulated thiol metabolic pathway have a role in conferring amphotericin B resistance in clinical isolates of Leishmania donovani.
authors: Department of Molecular Biology, Rajendra Memorial Research Institute of Medical Sciences, Agamkuan, Patna, Bihar, India.
authors: Bidyut
authors: B
authors: Purkait
authors: Ashish
authors: A
authors: Kumar
authors: Nilay
authors: N
authors: Nandi
authors: Abul Hasan
authors: AH
authors: Sardar
authors: Sushmita
authors: S
authors: Das
authors: Sudeep
authors: S
authors: Kumar
authors: Krishna
authors: K
authors: Pandey
authors: Vidyananda
authors: V
authors: Ravidas
authors: Manish
authors: M
authors: Kumar
authors: Tripti
authors: T
authors: De
authors: Dharmendra
authors: D
authors: Singh
authors: Pradeep
authors: P
authors: Das
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000378:metabolism; D000666:Amphotericin B / Q000494:pharmacology*; D000818:Animals; D000981:Antiprotozoal Agents / Q000494:pharmacology*; D002460:Cell Line; D004351:Drug Resistance; D005786:Gene Expression Regulation; D006801:Humans; D007194:India; D007893:Leishmania donovani / Q000187:drug effects* / Q000302:isolation & purification; D007898:Leishmaniasis, Visceral / Q000469:parasitology; D008560:Membrane Fluidity; D008563:Membrane Lipids / Q000737:chemistry; D021261:Parasitic Sensitivity Tests; D015800:Protozoan Proteins / Q000235:genetics / Q000378:metabolism; D013438:Sulfhydryl Compounds / Q000378:metabolism
chemical_list: D018528:ATP-Binding Cassette Transporters; D000981:Antiprotozoal Agents; D008563:Membrane Lipids; D015800:Protozoan Proteins; D013438:Sulfhydryl Compounds; D000666:Amphotericin B