Adriamycin Aglycones Research Articles

The exogenous NADH dehydrogenase of heart mitochondria is the key enzyme responsible for selective cardiotoxicity of anthracyclines.

The molecular mechanism of the anthracycline-dependent development of cardiotoxicity is still far from being clear. However, it is generally accepted, that mitochondria play a significant role in triggering this organ specific injury. The results presented in this study demonstrate that, in contrast to liver mitochondria, isolated heart mitochondria shuttle single electrons to adriamycin, giving rise to oxygen radical formation via autoxidation of adriamycin semiquinones. This one electron reduction of anthracyclines is catalyzed by the exogenous NADH dehydrogenase associated with complex I of heart mitochondria, an enzyme which is lacking in liver mitochondria. Upon addition of NADH heart mitochondria generate significant amounts of adriamycin semiquinones while liver mitochondria were ineffective. Adriamycin semiquinones undergo both autoxidation leading to superoxide radical release and complex reactions under formation of adriamycin aglycone. Due to the high lipophilicity adriamycin aglycones accumulate in the inner mitochondrial membrane where they interfere with electron carriers of the respiratory chain. Adriamycin aglycone semiquinones emerging from an interaction with complex I were found to trigger homolytic cleavage of H2O2 which results in the formation of hydroxyl radicals. As demonstrated in this study the activation of adriamycin by the exogenous NADH dehydrogenase of cardiac mitochondria initiates a cascade of reaction steps leading to the establishment of oxidative stress. Our experiments suggest the exogenous NADH dehydrogenase of heart mitochondria to play a key role in the cardiotoxicity of adriamycin. This organ-specific enzyme initiates a sequence of one electron transfer reactions ending up in the establishment of oxidative stress.

Interactions of Adriamycin aglycones with mitochondria may mediate Adriamycin cardiotoxicity

Adriamycin and related anthracyclines are potent oncolytic agents, the clinical utility of which is limited by severe cardiotoxicity. Aglycone metabolites of Adriamycin (5–20 μM) induce a Ca 2+-dependent increase in the permeability of the inner mitochondrial membrane of both heart and liver mitochondria to small (< 1500 Da) solutes; this phenomenon is accompanied by release of mitochondrial Ca 2+, mitochondrial swelling, collapse of the membrane potential, oxidation of mitochondrial pyridine nucleotides [NAD(P)H], uncoupling, and a transition from the condensed to the orthodox conformation and is inhibited by ATP, dithiothreitol, the immunosuppressant cyclosporin A, and the ubiquitous polyamine spermine. Aglycones also modify mitochondrial sulfhydryl groups and induce a Ca 2+ independent oxidation of mitochondrial NAD(P)H which appears to reflect electron transport from NADH to oxygen, mediated by the aglycones and resulting in the production of Superoxide (O 2 −). Selenium deficiency and butylated hydroxytoluene inhibit aglycone-induced Ca 2+ release from liver, but not heart, mitochondria, suggesting that the interactions of the aglycones with mitochondria diner in these two tissues. It can be proposed that the effects of Adriamycin aglycones on heart mitochondria are responsible for the cardiotoxicity of the parent drug.

Interaction of adriamycin aglycones with isolated mitochondria: Effect of selenium deficiency

Adriamycin (AdM) aglycones have dramatic effects on isolated heart mitochondria, oxidizing pyridine nucleotides, modifying sulfhydryl groups, and triggering a permeability transition of the inner membrane that results in free passage of solutes smaller than 1500 Da. In this investigation, the role of glutathione (GSH) peroxidase in these actions of the aglycones was evaluated, by comparing mitochondria from selenium-deficient and selenium-supplemented rats, with the following results. Selenium deficiency was without effect on the permeability transition of heart mitochondria, followed via Ca 2+ release and triggered by AdM aglycone or by t-butyl hydroperoxide (TBH) or H 2O 2, both of which are authentic substrates of the peroxidase. The permeability transitio of liver mitochondria was delayed by selenium deficiency regardless of the triggering agent; however, substantial triggering by the aglycone and TBH persisted in mitochondria from selenium-deficient animals. Selenium deficiency inhibited thiol modification elicited by AdM aglycone and H 2O 2 in heart mitochondria and by the aglycone, TBH, and possibly H 2O 2 in liver mitochondria. It would thus appear that AdM aglycone, TBH, and H 2O 2 can induce the permeability transition of isolated heart mitochondria via a process (or processes) distinct from the catalytic activity of the peroxidase. Furthermore, even in liver, where involvement of the peroxidase is observed, mechanisms other than the GSH cycle can contribute to transition induction by the agylcone and by TBH. Finally, mitochondrial -SH group modification by the aglycones appeared not be causally linked to induction of the permeability transition. This laboratory has suggested that the effects of aglycone metabolites of AdM on mitochondria mediate the cardiotoxicity that limits use of the parent drug. The data presented in this paper argue against the involvement of GSH peroxidase in that process. They are in agreement with in vivo studies, which have generally failed to find evidence for amelioration of AdM cardiotoxicity in selenium-deficient animals.

Oxidation of mitochondrial pyridine nucleotides by aglycone derivatives of adriamycin

Adriamycin (AdM) and related anthracyclines are potent antineoplastic agents, the clinical utility of which is limited by severe cardiotoxicity. Aglycone derivatives of AdM have recently been reported to trigger the release of Ca 2+ from isolated, preloaded rat heart mitochondria and to modify mitochondrial sulfhydryl (SH) groups. Both mitochondrial Ca 2+ retention and SH status are sensitive to mitochondrial NAD(P) + NAD(P)H ratios. This investigation examined the effects of AdM and its aglycone derivatives on the pyridine nucleotide redox status of isolated, intact heart mitochondria with the following results. (i) AdM aglycones induced the slow, Ca 2+-independent oxidation of mitochondrial NAD(P)H. Oxidation was proportional to aglycone concentration between 5 and 60 μ m. (ii) In terms of potency, 7-deoxy AdM aglycone ⩾ 7-hydroxy AdM aglycone ⪢ AdM. (iii) Inhibitor data suggested that NAD(P)H oxidation reflects the rotenone-insensitive reduction of AdM aglycone and subsequent electron transfer to O 2 generating Superoxide. (iv) NAD(P)H oxidation mediated by AdM aglycone could be distinguished from the Ca 2+-dependent NAD(P)H oxidation associated with mitochondrial Ca 2+ release. This communication is the first to describe redox interactions of AdM with intact mitochondria.

Inhibition by cyclosporin A and butylated hydroxytoluene of the inner mitochondrial membrane permeability transition induced by Adriamycin aglycones

Effets de la cyclosporine A et du BHT (butyl-hydroxytoluene) sur la transition de permeabilite induite par la partie aglycone de l'adriamycine dans les membranes des mitochondries du foie et du coeur chez le rat

Mitochondrial sulfhydryl group modification by adriamycin aglycones

Induction of Ca 2+ release from isolated, preloaded rat heart mitochondria by low concentrations (<5 μM) of adriamycin aglycones, has recently been reported [(1988) Biochem. Pharmacol. 37, 803]. Ca 2+ release occurs via a generalized, Ca 2+-dependent increase in the permeability of the inner mitochondrial membrane to small molecules. The process is antagonized by dithiothreitol, suggesting thiol involvement. This communication demonstrates modification of mitochondrial sulfhydryl groups, detected as decreased 5,5′-dithiobis-(2-nitrobenzoic acid) (DTNB) reactivity, by adriamycin aglycones. Ca 2+ release and sulfhydryl modification are shown to depend similarly on aglycone concentration and on the C-7 substituent of the anthracycline ring. In addition, DTNB elicits Ca 2+ release. It can therefore be proposed that adriamycin aglycones alter mitochondrial membrane permeability by altering mitochondrial thiol status.

Na +-independent release of Ca 2+ from rat heart mitochondria : Induction by adriamycin aglycone

The effect of adriamycin aglycones on Ca 2+ retention by isolated, preloaded rat heart mitochondria was assessed. After an initial lag, which decreased with increasing drug concentration, the 7-hydroxy-aglycone (5–20 μM) triggered Ca 2+ release. Aglycone-induced Ca 2+ release was correlated with Ca 2+ -dependent mitochondrial swelling, Ca 2+-dependent collapse of the mitochondrial membrane potential, Ca 2+-dependent oxidation of mitochondrial pyridine nucleotides, and a transition from the condensed to the orthodox configuration. Aglycone-induced Ca 2+ release was inhibited by dibucaine, dithiothreitol, ATP, and bovine serum albumin. It can be concluded, therefore, that aglycone-induced Ca 2+ release reflects the Ca 2+-dependent increase in the permeability of the inner mitochondrial membrane to solutes of molecular weight < 1000 which has been observed with other triggering agents [R. A. Haworth and D. R. Hunter, Archs Biochem. Biophys. 195, 460 (1979); I. Al-Nasser and M. Crompton, Biochem. J. 239, 19 (1986)]. In particular, the 7-hydroxy-aglycone decreased the amount of Ca 2+ required to trigger the permeability increase. No effect of the aglycone on Ca 2+ uptake could be discerned. 7-Deoxy-adriamycin aglycone, the more prominent biological metabolite of adriamycin, was similarly effective in inducing Ca 2+ release, and both aglycones were substantially more effective than the parent drug. Adriamycin and related anthracyclines are potent antineoplastic agents, the clinical use of which is limited by severe cardiotoxicity. These results suggest that aglycone formation and the resultant disruption of both cellular Ca 2+ homeostasis and metabolite compartmentation may mediate anthracycline cardiotoxicity.

Effect of cyclophosphamide pretreatment on the short-term disposition and biliary excretion of adriamycin metabolites in rat.

The effect of pretreatment with cyclophosphamide 180 mg/kg upon the short-term disposition of adriamycin in anesthetized rat 4 days later was studied. There was a significant decrease in plasma adriamycin clearance, from 125 to 48 ml/min/kg, and a significant decrease in the apparent volume of the peripheral compartment of adriamycin distribution, from 51.7 to 25.6 l/kg, in cyclophosphamide-pretreated as against control rats. Biliary excretion of adriamycin over 2.5 h was increased significantly by 114% in cyclophosphamide-pretreated rats and there was a small but nonsignificant increase in biliary adriamycinol excretion and a decrease in excretion of adriamycin aglycones. Cyclophosphamide pretreatment was associated with an 83% increase in bile flow. Cyclophosphamide pretreatment had no significant effect upon the utilization of adriamycin or upon the formation of adriamycin metabolites by rat isolated hepatocytes. The results suggest that NADPH-cytochrome P-450 reductase, which is decreased 40% by cyclophosphamide pretreatment, is not rate-limiting in elimination of adriamycin. Biliary excretion of adriamycin is increased when plasma adriamycin clearance is decreased, suggesting that cyclophosphamide pretreatment affects a pathway besides biliary excretion that is responsible for the short-term removal of adriamycin from plasma.