Interactions of prostaglandin A2 with the glutathione-mediated biotransformation system

The cyclopentenone prostaglandin Az (PGA,) is known to inhibit cell proliferation, and metabolism of this compound thus might be important in controlling its ultimate function. The glutathione-related metabolism of PGA(2) was therefore investigated both with purified glutathione S-transferase P1-1 (GSTP1-1) and with IGR-39 human melanoma cells. Firstly, the irreversible inhibition of human GSTP1-1 and its mutants C47S, C101S, and C47S/C101S was studied. PGA2 appeared to inhibit GSTP1-1 mainly by binding to the cysteine 47 moiety of the enzyme. This binding was reversed by a molar excess of GSH, indicating that retro-Michael cleavage occurs. Secondly, after exposing IGR-39 human melanoma cells to PGA(2), both diastereoisomers of die PGA(2)-glutathione conjugate are excreted into the medium, although with a clear excess of the S-form, due to its preferential formation by the GSTP1-1 present in the cells. Thirdly, the effect of PGA(2) on intracellular GST activity was determined by quantification of the excreted glutathione conjugate S-(2,4-dinitrophenyl)glutathione (DNPSG) after exposure to 1-chloro-2,4-dinitrobenzene. DNPSG excretion was inhibited after incubation with 10 or 20 mu M PGA(2) for 1 or 4 hr, as a result of glutathione depletion, reversible GST inhibition, and covalent modification of intracellular GST. Furthermore, PGA(2) also inhibited transport of DNPSG by the multidrug resistance-associated protein, an effect that was reversible and competitive. In conclusion, PGA(2) modulates all three aspects of the glutathione mediated biotransformation system, i.e. GSH levels, GSTP1-1 activity, and transport of GSH conjugates. A role for GSTP1-1 as a specific transport protein inside the cell is indicated. (C) 1999 Elsevier Science Inc.