Daunorubicin-Induced Cardiac Injury in the Rabbit: A Role for Daunorubicinol?

doi:10.1006/taap.1993.1023

Toxicology and Applied Pharmacology

Volume 118, Issue 2, February 1993, Pages 177-185

https://doi.org/10.1006/taap.1993.1023 Get rights and content

Abstract

This study evaluated potential contributions of daunorubicin and its principle metabolite, daunorubicinol, to the cardiotoxicity of daunorubicin therapy. Daunorubicin (15 mg/kg) or placebo (normal saline) was administered by iv bolus to New Zealand white rabbits and 3 to 4 days later, hearts were removed to measure contractility (dF/dt), concentrations of daunorubicin and daunorubicinol, and evidence of oxidative stress on glutathione and glutathione peroxidase. Contractile function of isolated atria and papillary muscles was depressed (p < 0.05). Daunorubicinol exceeded daunorubicin concentration in the heart (p < 0.005) with a ratio of metabolite to parent drug of 26 in atrial and 32 in ventricular tissue. There was a significant correlation between peak plasma (r = −0.63; p < 0.05) or cardiac concentration (r = −0.78; p < 0.02) of daunorubicinol, but not daunorubicin, and depression of dF/dt in papillary muscles. In separate in vitro studies, daunorubicinol at a concentration (5.5 μg/g tissue or 10 μM) approximating that observed ex vivo in heart inhibited Ca²⁺ uptake into cardiac sarcoplasmic reticulum vesicles by 39 ± 3%, whereas 10 μM daunorubicin (14-fold higher than actual ex vivo cardiac concentrations) did not demonstrate any detectable inhibition. Daunorubicin treatment failed to significantly alter concentrations of GSH or GSSG or activities of glutathione peroxidase in the heart. Thus, cardiac dysfunction observed 3 to 4 days after a single dose of daunorubicin did not clearly relate to oxidative stress, but was associated with a cardiac concentration of daunorubicinol that appeared sufficiently high to impair Ca²⁺ metabolism.

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Cited by (52)

Inhibition of human anthracycline reductases by emodin - A possible remedy for anthracycline resistance
2016, Toxicology and Applied Pharmacology
Citation Excerpt :
All other reductases are most likely to play minor roles in DOX reduction. There is growing evidence that the C-13 hydroxy metabolites DAUNOL and DOXOL are the main trigger for chronic cardiotoxicity of anthracyclines (Bains et al., 2013; Forrest et al., 2000; Licata et al., 2000; Mordente et al., 2001; Olson et al., 1988; Propper and Maser, 1997) and, most importantly, that inhibition of specific anthracycline reductases may increase the chemotherapeutic efficacy and decrease the cardiotoxicity of anthracyclines (Cusack et al., 1993; Forrest & Gonzalez, 2000; Gavelová et al., 2008; Hofman et al., 2014; Olson et al., 2003; Tanaka et al., 2005; Zhong et al., 2011). In line with this, positive effects of the co-administration of curcumin, an inhibitor for CBR1 (Hintzpeter et al., 2014), AKR1B1 and AKR1B10 (Matsunaga et al., 2009; Muthenna et al., 2009), with DAUN or DOX have been reported in rat models (Swamy et al., 2012; Venkatesan, 1998; Venkatesan et al., 2000) and in numerous human cancer cell lines (Hosseinzadeh et al., 2011; Qian et al., 2011; Wang et al., 2011).
The clinical application of anthracyclines, like daunorubicin and doxorubicin, is limited by two factors: dose-related cardiotoxicity and drug resistance. Both have been linked to reductive metabolism of the parent drug to their metabolites daunorubicinol and doxorubicinol, respectively. These metabolites show significantly less anti-neoplastic properties as their parent drugs and accumulate in cardiac tissue leading to chronic cardiotoxicity. Therefore, we aimed to identify novel and potent natural inhibitors for anthracycline reductases, which enhance the anticancer effect of anthracyclines by preventing the development of anthracycline resistance.
Human enzymes responsible for the reductive metabolism of daunorubicin were tested for their sensitivity towards anthrachinones, in particular emodin and anthraflavic acid. Intense inhibition kinetic data for the most effective daunorubicin reductases, including IC₅₀- and K_i-values, the mode of inhibition, as well as molecular docking, were compiled. Subsequently, a cytotoxicity profile and the ability of emodin to reverse daunorubicin resistance were determined using multiresistant A549 lung cancer and HepG2 liver cancer cells. Emodin potently inhibited the four main human daunorubicin reductases in vitro. Further, we could demonstrate that emodin is able to synergistically sensitize human cancer cells towards daunorubicin at clinically relevant concentrations.
Therefore, emodin may yield the potential to enhance the therapeutic effectiveness of anthracyclines by preventing anthracycline resistance via inhibition of the anthracycline reductases. In symphony with its known pharmacological properties, emodin might be a compound of particular interest in the management of anthracycline chemotherapy efficacy and their adverse effects.
Curcumin is a tight-binding inhibitor of the most efficient human daunorubicin reductase - Carbonyl reductase 1
2015, Chemico-Biological Interactions
Citation Excerpt :
Convincing evidence supports the idea that the C-13 hydroxy metabolites of DAUN and DOX, daunorubicinol (DAUNOL) and doxorubicinol (DOXOL), respectively, are the main trigger for chronic cardiotoxicity [16–21]. Thus, inhibition of CBR1 may increase the efficacy and decrease cardiotoxicity of anthracyclines [7,22,23]. Indeed, in various cell culture models it has been demonstrated that inhibition of CBR1 by known inhibitors, e.g. hydroxy-PP (3-(1-tert-butyl-4-amino-1H-pyrazolo[3,4-d]pyrimidin-3-yl)phenol), enhanced the effectiveness and decreased the cardiotoxicity of the anticancer drug DAUN by preventing its reduction to DAUNOL [24].
Curcumin is a major component of the plant Curcuma longa L. It is traditionally used as a spice and coloring in foods and is an important ingredient in curry. Curcuminoids have anti-oxidant and anti-inflammatory properties and gained increasing attention as potential neuroprotective and cancer preventive compounds. In the present study, we report that curcumin is a potent tight-binding inhibitor of human carbonyl reductase 1 (CBR1, K_i = 223 nM). Curcumin acts as a non-competitive inhibitor with respect to the substrate 2,3-hexandione as revealed by plotting IC₅₀-values against various substrate concentrations and most likely as a competitive inhibitor with respect to NADPH. Molecular modeling supports the finding that curcumin occupies the cofactor binding site of CBR1. Interestingly, CBR1 is one of the most effective human reductases in converting the anthracycline anti-tumor drug daunorubicin to daunorubicinol. The secondary alcohol metabolite daunorubicinol has significantly reduced anti-tumor activity and shows increased cardiotoxicity, thereby limiting the clinical use of daunorubicin. Thus, inhibition of CBR1 may increase the efficacy of daunorubicin in cancer tissue and simultaneously decrease its cardiotoxicity. Western-blots demonstrated basal expression of CBR1 in several cell lines. Significantly less daunorubicin reduction was detected after incubating A549 cell lysates with increasing concentrations of curcumin (up to 60% less with 50 μM curcumin), suggesting a beneficial effect in the co-treatment of anthracycline anti-tumor drugs together with curcumin.
Understanding the binding of daunorubicin and doxorubicin to NADPH-dependent cytosolic reductases by computational methods
2012, European Journal of Medicinal Chemistry
Citation Excerpt :
Among them, a prevailing hypothesis implicates the C-13 hydroxy metabolite, doxorubicinol (DOXol) and daunorubicinol (DAUNol), compounds that lack the antiproliferative activity of DOX and DAUN, as a major component in the development of chronic cardiomyopathy [7]. The accumulation of the metabolite in the heart has been described suggesting that the C-13 hydroxy metabolite might be transported and selectively trapped by cardiac cells [8]. Several cytosolic reductases of the short chain dehydrogenase class (SDR) and of the aldo–keto reductase class (AKR) metabolize DOX to DOXol and DAUN to DAUNol (Fig. 1) implicating these enzymes in the development of cardiotoxicity [9,10].
The anthracycline anticancer agents daunorubicin (DAUN) and doxorubicin (DOX) are reduced by different NADPH-dependent cytosolic reductases into their corresponding alcohol metabolites daunorubicinol (DAUNol) and doxorubicinol (DOXol), which have been implicated in the development of chronic cardiomyopathy. To better understand the individual importance of each enzyme in the reduction and to provide deeper insight into the binding at atomic level we performed molecular docking and dynamics simulations of DAUN and DOX into the active sites of human carbonyl reductase 1 (CBR1) and human aldehyde reductase (AKR1A1). Such simulations evidenced a different behavior between the reductases with respect to DAUN and DOX suggesting major contribution of CBR1 in the reduction. The results are in agreement with available experimental data and for each enzyme and anthracycline pair provided the identification of key residues involved in the interactions. The structural models that we have derived could serve as a useful tool for structure-guided drug design studies.
Synthesis of 3-[(N-carboalkoxy)ethylamino]-indazole-dione derivatives and their biological activities on human liver carbonyl reductase
2010, Bioorganic and Medicinal Chemistry
A series of indazole-dione derivatives were synthesized by the 1,3-dipolar cycloaddition reaction of appropriate substituted benzoquinones or naphthoquinones and N-carboalkoxyamino diazopropane derivatives. These compounds were evaluated for their effects on human carbonyl reductase. Several of the analogs were found to serve as substrates for carbonyl reductase with a wide range of catalytic efficiencies, while four analogs display inhibitory activities with IC₅₀ values ranging from 3–5 μM. Two of the inhibitors were studied in greater detail and were found to be noncompetitive inhibitors against both NADPH and menadione with K_I values ranging between 2 and 11 μM. Computational studies suggest that conformation of the compounds may determine whether the indazole-diones bind productively to yield product or nonproductively to inhibit the enzyme.
Effects of angiotensin II receptor blocker (candesartan) in daunorubicin-induced cardiomyopathic rats
2006, International Journal of Cardiology
Daunorubicin is an anthracycline anti-tumor agent; anthracycline chemotherapy in cancer can cause severe cardiomyopathy leading to a frequently fatal congestive heart failure; the first-line treatment is diuretics and digoxin. Recently, angiotensin-converting enzyme inhibitors have been shown to be effective in the treatment of such toxicity. The purpose of this study was to investigate the effects of angiotensin II type-1 receptor antagonist (candesartan) in a rat model of daunorubicin-induced cardiomyopathy.
Rats were treated with a cumulative dose of 9 mg/kg body weight daunorubicin (i.v.). 28 days later, after the development of cardiomyopathy, animals were randomly assigned to candesartan-treated (5 mg/kg/day, p.o.) or vehicle-treated groups; age-matched normal rats were used as the control group. Candesartan treatment was continued for 28 days. Hemodynamic and echocardiographic parameters were measured, cardiac protein and mRNA were analyzed, and histopathological analyses of myocardial fibrosis, cell size and apoptosis were conducted.
Following cardiomyopathy, left ventricular end diastolic pressure and left ventricular systolic dimension were significantly elevated; while % fractional shortening and Doppler E/A ratio were significantly reduced. Cardiomyopathic hearts showed significant increases in % fibrosis, % apoptosis, and myocyte diameter/body weight ratio; candesartan treatment reversed these changes. Fas-L protein overexpression in myopathic hearts was significantly suppressed by treatment with candesartan. Moreover, SERCA2 mRNA and protein expression were both down-regulated in myopathic hearts and restored to normal by candesartan treatment, significantly.
Our findings suggest that candesartan treatment significantly improved the left ventricular function and reversed the myocardial pathological changes investigated in this model of daunorubicin-induced cardiomyopathy; suggesting its potentials in limiting daunorubicin cardiotoxicity.
Anthracycline secondary alcohol metabolite formation in human or rabbit heart: Biochemical aspects and pharmacologic implications
2003, Biochemical Pharmacology
Citation Excerpt :
There are of course potential arguments against the hypothesis that secondary alcohol metabolites mediate chronic cardiomyopathy. Some investigators reported that cardiac dysfunction induced in rabbits after systemic administration of DRN correlated with cardiac concentration of DNRol but not with that of DNR [41]. This would be consistent with the alcohol metabolite hypothesis.
Clinical use of the anticancer anthracyclines doxorubicin (DOX) and daunorubicin (DNR) is limited by development of cardiotoxicity upon chronic administration. Secondary alcohol metabolites, formed after two-equivalent reduction of a carbonyl group in the side chain of DOX or DNR, have been implicated as potential mediators of chronic cardiotoxicity. In the present study we characterized how human heart converted DOX or DNR to their alcohol metabolites DOXol or DNRol. Experiments were carried out using post-mortem myocardial samples obtained by ethically-acceptable procedures, and results showed that DOXol and DNRol were formed by flavin-independent cytoplasmic reductases which shared common features like pH-dependence and requirement for NADPH, but not NADH, as a source of reducing equivalents. However, studies performed with inhibitors exhibiting absolute or mixed specificity toward best known cytoplasmic reductases revealed that DOX and DNR were metabolized to DOXol or DNRol through the action of distinct enzymes. Whereas DOX was converted to DOXol by aldehyde-type reductase(s) belonging to the superfamily of aldo-keto reductases, DNR was converted to DNRol by carbonyl reductase(s) belonging to the superfamily of short-chain dehydrogenase/reductases. This pattern changed in cardiac cytosol derived from rabbit, a laboratory animal often exploited to reproduce cardiotoxicity induced by anthracyclines and to develop protectants for use in cancer patients. In fact, only carbonyl reductases were involved in metabolizing DOX and DNR in rabbit cardiac cytosol, although with different K_m and V_max. Collectively, these results demonstrate that human myocardium convert DOX and DNR to DOXol or DNRol by virtue of different reductases, an information which may be of value to prevent alcohol metabolite formation during the course of anthracycline-based anticancer therapy. These results also raise caution on the preclinical value of animal models of anthracycline cardiotoxicity, as they demonstrate that the metabolic routes leading to DOXol in a laboratory animal may not be the same as those occurring in patients.

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Regular ArticleDaunorubicin-Induced Cardiac Injury in the Rabbit: A Role for Daunorubicinol?

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Regular Article
Daunorubicin-Induced Cardiac Injury in the Rabbit: A Role for Daunorubicinol?