Browsing by Author "Widersten, Mikael"

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    Inhibition of Glutathione S-Transferases by antimalarial drugs possible implications for circumventing anticancer drug resistance.
    (Wiley-Liss, Inc., 2001-08-06) Mukanganyama, Stanley; Widersten, Mikael; Naik, Yogeshkumar S.; Mannervik, B.; Hasler, Julia A.
    A strategy to overcome multidrug resistance in cancer cells involves treatment with a combination of the antineoplastic agent and a chemomodulator that inhibits the activity of the resistance-causing protein. The aim of our study was to investigate the effects of antimalarial drugs on human recombinant glutathione S-transferase (GSTs) activity in the context of searching for effective and clinically acceptable inhibitors of these enzymes. Human recombinant GSTs heterologously expressed in Escherichia coli were used for inhibition studies. GST Al-l activity was inhibited by artemisinin with an IC,, of 6 pM, whilst GST MI-l was inhibited by quinidine and its diastereoisomer quinine with IC5,s of I2 pM and 17 pM, respectively. GST M3-3 was inhibited by tetracycline only with an IC,, of 47 pM. GST PI-l was the most susceptible enzyme to inhibition by antimalarials with IC,, values of I, 2, 1, 4, and 13 pM for pyrimethamine, arteniislnin, quinidine, quinine and tetracycline, respectively. The IC,, values obtained for artemisinin, quinine, quinidine and tetracycline are below peak plasma concentrations obtained during therapy of malaria with these drugs. It seems likely, therefore, that GSTs may be inhibited in vivo at doses normally used in clinical practice. Using the substrate ethacrynic acid, a diuretic drug also used as a modulator to overcome drug resistance in tumour cells, GST PI-l activity was inhibited by tetracycline, quinine, pyrimethamine and quinidine with IC,, values of 18, 27, 45 and 70 pM, respectively. The ubiquitous expression of GSTs in different malignancies suggests that the addition of nontoxic reversing agents such as antimalarials could enhance the efficacy of a variety of alkylating agents.
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    Proposed Reductive Metabolism of Artemisinin by Glutathione Transferases in vitro
    (The Harwood academic publishers imprint,, 2001-01-18) Naik, Yogeshkumar S.; Muganyama, Stanley; Hasler, Julia A.; Widersten, Mikael; Mannervik, B.
    Artemisinin is a sesquiterpene lactone containing an endoperoxide bridge. It is a promising new antimalarial and is particularly useful against the drug resistant strains of Plasmodium fakiparum. It has unique antimalarial properties since it acts through the generation of free radicals that alkylate parasite proteins. Since the antimalarial action of the drug is antagonised by glutathionc and ascorbate and has unusual pharmacokinetic properties in humans, we have investigated if the drug is broken down by a typical reductive reaction in the presence of glutathione transferases. Cytosolic glutathione transferases (GSTs) detoxlfy electrophilic xenobiotics by catalysing the formation of glutathione (GSH) conjugates and exhibit glutathione peroxidase activity towards hydroperoxides. Arternisinin was incubated with glutathione, NADPH and glutathione reductase and G~TiSn a coupled assay syste&analogous to the standard assav scheme with cumene hv,d rover- L oxide as a substrate df GSTS. Arternisinin was shown to stimulate NADPH oxidation in cytosols from rat liver, kidney, intestines and in affinity purified preparations of GSTs from rat liver. Using human recombinant GSTs hetelorogously expressed in Escherichia coli, artemisinin was similarly shown to stimulate NADPH oxidation with the highest activity observed with GST MI-1. Using recombinant GSTs the activity of GSTs with artemisinin was at least two fold higher than the reaction with CDNB. Considering these results, it is possible that GSTs may contribute to the metabolism of artemisinin in the presence of NADPH and GSSG-reductase We propose a model, based on the known reactions of GSTs and sesquiterpenes, in which (1) artemisinin reacts with GSH resulting in oxidised glutathione; (.2) the oxidised glutathione is then converted to reduced glutathione via glutathione reductase; and (3) the latter reaction may then result in the depletion of NADPH via GSSG-reductase. The ability of artemisinin to react with GSH in the presence of GST may be responsible for the NADPH utilisation observed in vitro and suggests that cytosolic GSTs are likely to be contributing to metabolism of artemisinin and related drugs in vivo