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The effects of 4-hydroxytamoxifen (OHTAM) on the mitochondrial permeability transition (MPT) induced by Ca 2+ plus peroxynitrite (ONOO - ) or phenylarsine oxide (PhAsO) were studied to clarify its mechanisms of MPT inhibition. Ca 2+ plus ONOO - induced mitochondrial swelling, membrane potential (??) depolarization, and Ca 2+ release. OHTAM, when preincubated with mitochondria, prevents those events, and if added after their induction this drug promotes a time-dependent reversal of ?? depolarization and Ca 2+ release associated with MPT induction, because these events are also inhibited by cyclosporine A (CyA). Preincubation with OHTAM also inhibits thiol group oxidation associated with the MPT promoted by ONOO - and allows the NAD(P) + to recover their reduced state faster and in a higher extension. The mitochondrial swelling and Ca 2+ release after MPT induction with Ca 2+ plus PhAsO are inhibited by OHTAM; similarly to CyA, the oxidation of NAD(P)H induced by this combination is also inhibited. According to these data, the MPT inhibition by OHTAM may be related to its antioxidant capacity and the binding to target protein components of the MPT, preventing the oxidation of NAD(P)H and thiol groups, an event that increases the sensitivity to MPT induction. [ABSTRACT FROM AUTHOR]

The antiestrogen tamoxifen (TAM) inhibits the growth of different estrogen receptor (ER)-negative cells. Recently, multiple effects of TAM on mitochondrial bioenergetic functions have been pointed to explain its ER-independent cell death mechanisms. We have shown that TAM and its major active metabolite 4-hydroxytamoxifen (OHTAM) induce depolarization of the mitochondrial membrane potential (ΔΨ) and uncouple the mitochondrial respiration, depressing the oxidative phosphorylation efficiency. To clarify the biochemical mechanisms underlying the changes in the regulation of ATP synthesis and yield, in this work we evaluated the alterations of mitochondrial adenine nucleotides induced by both drugs and ascertained whether such changes could reflect a specific inhibition of either the adenine nucleotide translocase (ANT) or the phosphate carrier, as well as the activation of ATP hydrolysis due to ΔΨ depolarization. We found that both antiestrogens caused a concentration-dependent decrease in mitochondrial ATP levels. Mitochondrial ADP and AMP were concomitantly increased with a subsequent decrease in the ATP/ADP or ATP/AMP ratios. The total concentration of adenine nucleotides also changed. Additionally, both drugs decreased the ANT content of mitochondria, inhibited the phosphate carrier and induced ATP hydrolysis. However, the effects of TAM were more drastic than those induced by OHTAM. Therefore, the depletion of ATP might result from an activation of ATP catabolism, as well as from a decrease in the mitochondrial content of ANT and partial inhibition of the phosphate carrier. Our data may explain the ER-independent effects and cytotoxicity of both drugs and, in agreement with other previous studies, suggest that OHTAM is much less toxic to mitochondria than TAM. [Copyright &y& Elsevier]

The effects of 4-hydroxytamoxifen (OHTAM), the major active metabolite of the antiestrogen tamoxifen used in the breast cancer therapy, were studied on the mitochondrial permeability transition (MPT) and bioenergetic functions of mitochondria to evaluate the mechanisms underlying the cell death and toxic effects. The MPT was induced in vitro by incubating rat liver mitochondria with 1 mM inorganic phosphate plus Ca2+ and with tert-butyl hydroperoxide. OHTAM provides protection against the Ca2+-induced mitochondrial swelling, depolarization of the mitochondrial membrane potential (ΔΨ), loss of electrophoretic Ca2+ uptake capacity and uncoupling of respiration, similarly to cyclosporine A. The concentrations of OHTAM used do not significantly affect ΔΨ, respiratory control and adenosine diphosphate/oxygen ratios and induce repolarization and Ca2+ re-uptake, suggesting that such inhibitory effects of OHTAM were due to the prevention of the MPT induction and not due to the inhibition of the mitochondrial Ca2+ uniporter. Since the MPT induction has been linked to an oxidized shift in the mitochondrial redox state and/or increase in the generation of reactive oxygen species, the MPT prevention by OHTAM may be related to its high antioxidant capacity. [Copyright &y& Elsevier]