Increased Glycolytic ATP Synthesis Is Associated with Tafenoquine Resistance in<i>Leishmania major</i>

Manzano, José Ignacio; Carvalho, Luís M.; Pérez‐Victoria, José M.; Castanys, Santiago; Gamarro, Francisco

doi:10.1128/aac.01545-10

Cited by 31 publications

(37 citation statements)

References 33 publications

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“…To determine the relationship between thiol levels and the sensitivity of the different Leishmania lines, 3 mM BSO (a ␥-glutamylcysteine synthetase inhibitor) was added at 28°C for 48 h before the sensitivity experiments. The sensitivity of intracellular Leishmania amastigotes to Sb(III) and Sb(V) (Pentostam) was determined after 72 h of incubation as described previously (28).…”

Section: Methodsmentioning

confidence: 99%

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A New ABC Half-Transporter in Leishmania major Is Involved in Resistance to Antimony

Manzano

García-Hernández

Castanys

et al. 2013

Antimicrob Agents Chemother

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The characterization of ABCI4, a new intracellular ATP-binding cassette (ABC) half-transporter in Leishmania major, is described. We show that ABCI4 is involved in heavy metal export, thereby conferring resistance to Pentostam, to Sb(III), and to As(III) and Cd(II). Parasites overexpressing ABCI4 showed a lower mitochondrial toxic effect of antimony by decreasing reactive oxygen species production and maintained higher values of both the mitochondrial electrochemical potential and total ATP levels with respect to controls. The ABCI4 half-transporter forms homodimers as determined by a coimmunoprecipitation assay. A combination of subcellular localization studies under a confocal microscope and a surface biotinylation assay using parasites expressing green fluorescent protein-and FLAG-tagged ABCI4 suggests that the transporter presents a dual localization in both mitochondria and the plasma membrane. Parasites overexpressing ABCI4 present an increased replication in mouse peritoneal macrophages. We have determined that porphyrins are substrates for ABCI4. Consequently, the overexpression of ABCI4 confers resistance to some toxic porphyrins, such as zinc-protoporphyrin, due to the lower accumulation resulting from a significant efflux, as determined using the fluorescent zinc-mesoporphyrin, a validated heme analog. In addition, ABCI4 has a significant ability to efflux thiol after Sb(III) incubation, thus meaning that ABCI4 could be considered to be a potential thiol-X-pump that is able to recognize metal-conjugated thiols. In summary, we have shown that this new ABC transporter is involved in drug sensitivity to antimony and other compounds by efflux as conjugated thiol complexes.

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Section: Methodsmentioning

confidence: 99%

“…Amastigotes were cultured for 24 and 72 h in infected peritoneal macrophages from BALB/c mice (Charles River, Ltd.) as described previously (28).…”

Section: Methodsmentioning

confidence: 99%

A New ABC Half-Transporter in Leishmania major Is Involved in Resistance to Antimony

Manzano

García-Hernández

Castanys

et al. 2013

Antimicrob Agents Chemother

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“…As described previously, intracellular pH of wild-type or mutant L. major strains was measured using the pH-sensitive fluorescent sensor, BCECF-AM (Invitrogen) (Pal et al, 2015;Manzano et al, 2011). Briefly, 1×10 7 late log phase L. major promastigotes were resuspended in 10 μM BCECF-AMcontaining buffer and incubated for 30 min.…”

Section: Intracellular Ph Measurementmentioning

confidence: 99%

Interplay between a cytosolic and a cell surface carbonic anhydrase in pH homeostasis and acid tolerance of Leishmania

Pal

Abbasi

Mondal

et al. 2017

Journal of Cell Science

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Leishmania parasites have evolved to endure the acidic phagolysosomal environment within host macrophages. How Leishmania cells maintain near-neutral intracellular pH and proliferate in such a proton-rich mileu remains poorly understood. We report here that, in order to thrive in acidic conditions, Leishmania major relies on a cytosolic and a cell surface carbonic anhydrase, LmCA1 and LmCA2, respectively. Upon exposure to acidic medium, the intracellular pH of the LmCA1 +/− , LmCA2 +/− and LmCA1 +/− :LmCA2 +/− mutant strains dropped by varying extents that led to cell cycle delay, growth retardation and morphological abnormalities. Intracellular acidosis and growth defects of the mutant strains could be reverted by genetic complementation or supplementation with bicarbonate. When J774A.1 macrophages were infected with the mutant strains, they exhibited much lower intracellular parasite burdens than their wild-type counterparts. However, these differences in intracellular parasite burden between the wild-type and mutant strains were abrogated if, before infection, the macrophages were treated with chloroquine to alkalize their phagolysosomes. Taken together, our results demonstrate that haploinsufficiency of LmCA1 and/or LmCA2 renders the parasite acid-susceptible, thereby unravelling a carbonic anhydrase-mediated pH homeostatic circuit in Leishmania cells.

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“…ATP was measured using a CellTiter-Glo luminescent assay, which generates a luminescent signal proportional to the amount of ATP present, as described previously (19). BSF parasites (10 7 per ml) were incubated in culture medium with or without 10 M TFQ for 10, 60, 120, and 180 min.…”

Section: Methodsmentioning

confidence: 99%

“…TFQ accumulation was determined fluorometrically by recording an emission spectrum in the range of 360 to 460 nm upon excitation at 340 nm using an Aminco-Bowman Series 2 spectrometer as described previously (19).…”

Section: Methodsmentioning

confidence: 99%

The Oral Antimalarial Drug Tafenoquine Shows Activity against Trypanosoma brucei

Carvalho

Martínez‐García

Pérez-Victoria

et al. 2015

Antimicrob Agents Chemother

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cThe protozoan parasite Trypanosoma brucei causes human African trypanosomiasis, or sleeping sickness, a neglected tropical disease that requires new, safer, and more effective treatments. Repurposing oral drugs could reduce both the time and cost involved in sleeping sickness drug discovery. Tafenoquine (TFQ) is an oral antimalarial drug belonging to the 8-aminoquinoline family which is currently in clinical phase III. We show here that TFQ efficiently kills different T. brucei spp. in the submicromolar concentration range. Our results suggest that TFQ accumulates into acidic compartments and induces a necrotic process involving cell membrane disintegration and loss of cytoplasmic content, leading to parasite death. Cell lysis is preceded by a wide and multitarget drug action, affecting the lysosome, mitochondria, and acidocalcisomes and inducing a depolarization of the mitochondrial membrane potential, elevation of intracellular Ca 2؉ , and production of reactive oxygen species. This is the first report of an 8-aminoquinoline demonstrating significant in vitro activity against T. brucei.

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Increased Glycolytic ATP Synthesis Is Associated with Tafenoquine Resistance inLeishmania major

Cited by 31 publications

References 33 publications

A New ABC Half-Transporter in Leishmania major Is Involved in Resistance to Antimony

A New ABC Half-Transporter in Leishmania major Is Involved in Resistance to Antimony

Interplay between a cytosolic and a cell surface carbonic anhydrase in pH homeostasis and acid tolerance of Leishmania

The Oral Antimalarial Drug Tafenoquine Shows Activity against Trypanosoma brucei

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