Autophagy (but not metabolism) is a key event in mitoxantrone-induced cytotoxicity in differentiated AC16 cardiac cells.

Mitoxantrone (MTX) is an antineoplastic agent used to treat advanced breast cancer, prostate cancer, acute leukemia, lymphoma and multiple sclerosis. Although it is known to cause cumulative dose-related cardiotoxicity, the underlying mechanisms are still poorly understood. This study aims to compare the cardiotoxicity of MTX and its' pharmacologically active metabolite naphthoquinoxaline (NAPHT) in an in vitro cardiac model, human-differentiated AC16 cells, and determine the role of metabolism in the cardiotoxic effects. Concentration-dependent cytotoxicity was observed after MTX exposure, affecting mitochondrial function and lysosome uptake. On the other hand, the metabolite NAPHT only caused concentration-dependent cytotoxicity in the MTT reduction assay. When assessing the effect of different inhibitors/inducers of metabolism, it was observed that metyrapone (a cytochrome P450 inhibitor) and phenobarbital (a cytochrome P450 inducer) slightly increased MTX cytotoxicity, while 1-aminobenzotriazole (a suicide cytochrome P450 inhibitor) decreased fairly the MTX-triggered cytotoxicity in differentiated AC16 cells. When focusing in autophagy, the mTOR inhibitor rapamycin and the autophagy inhibitor 3-methyladenine exacerbated the cytotoxicity caused by MTX and NAPHT, while the autophagy blocker, chloroquine, partially reduced the cytotoxicity of MTX. In addition, we observed a decrease in p62, beclin-1, and ATG5 levels and an increase in LC3-II levels in MTX-incubated cells. In conclusion, in our in vitro model, neither metabolism nor exogenously given NAPHT are major contributors to MTX toxicity as seen by the residual influence of metabolism modulators used on the observed cytotoxicity and by NAPHT's low cytotoxicity profile. Conversely, autophagy is involved in MTX-induced cytotoxicity and MTX seems to act as an autophagy inducer, possibly through p62/LC3-II involvement.

Chemobrain: mitoxantrone-induced oxidative stress, apoptotic and autophagic neuronal death in adult CD-1 mice.

Mitoxantrone (MTX) is a topoisomerase II inhibitor used to treat a wide range of tumors and multiple sclerosis but associated with potential neurotoxic effects mediated by hitherto poorly understood mechanisms. In adult male CD-1 mice, the underlying neurotoxic pathways of a clinically relevant cumulative dose of 6 mg/kg MTX was evaluated after biweekly administration for 3 weeks and sacrifice 1 week after the last administration was undertaken. Oxidative stress, neuronal damage, apoptosis, and autophagy were analyzed in whole brain, while coronal brain sections were used for a closer look in the hippocampal formation (HF) and the prefrontal cortex (PFC), as these areas have been signaled out as the most affected in 'chemobrain'. In the whole brain, MTX-induced redox imbalance shown as increased endothelial nitric oxide synthase and reduced manganese superoxide dismutase expression, as well as a tendency to a decrease in glutathione levels. MTX also caused diminished ATP synthase ß expression, increased autophagic protein LC3 II and tended to decrease p62 expression. Postsynaptic density protein 95 expression decreased in the whole brain, while hyperphosphorylation of Tau was seen in PFC. A reduction in volume was observed in the dentate gyrus (DG) and CA1 region of the HF, while GFAP-ir astrocytes increased in all regions of the HF except in the DG. Apoptotic marker Bax increased in the PFC and in the CA3 region, whereas p53 decreased in all brain areas evaluated. MTX causes damage in the brain of adult CD-1 mice in a clinically relevant cumulative dose in areas involved in memory and cognition.

Ratiometric Delivery of Mitoxantrone and Berberine Co-encapsulated Liposomes to Improve Antitumor Efficiency and Decrease Cardiac Toxicity.

Combination therapy is one of the most common clinical practices in the treatment of malignancies. Synergistic effects, however, are produced only when optimal ratios of combined drugs were delivered to tumor cells. Thus, carriers co-encapsulating of multiple drugs are widely utilized for coordinated delivery. Herein, co-encapsulated pegylated liposomal formulation of mitoxantrone (MIT) and berberine (BER) at an optimal ratio has been developed (MBL) with high encapsulation efficiency (EE) and drug loading in order to achieve the purpose of ratiometric loading and delivery. MBL can not only extend blood circulation but also enhance tumor accumulation for both MIT and BER. More importantly, MBL can maintain the originally desired drug ratio in tumors within 48 h of intravenous injection for synergistic therapy. Compared with the liposomal formulation of MIT-treated group (ML), the progression of tumor growth was inhibited significantly in murine 4T1 breast tumor model after the treatment of MBL, as well as a lower cardiac toxicity. In addition, MBL evidently prolonged the survival of mice with L1210 ascitic tumor model. In summary, such a strategy of co-encapsulated liposomes could improve the clinical applications against multiple cancers.

Mitoxantrone impairs proteasome activity and prompts early energetic and proteomic changes in HL-1 cardiomyocytes at clinically relevant concentrations.

Mitoxantrone (MTX) is used to treat several types of cancers and to improve neurological disability in multiple sclerosis. Unfortunately, cardiotoxicity is a severe and common adverse effect in MTX-treated patients. Herein, we aimed to study early and late mechanisms of MTX-induced cardiotoxicity using murine HL-1 cardiomyocytes. Cells were exposed to MTX (0.1, 1 or 10 µM) during short (2, 4, 6, or 12 h) or longer incubation periods (24 or 48 h). At earlier time points, (6 and 12 h) cytotoxicity was already observed for 1 and 10 µM MTX. Proteomic analysis of total protein extracts found 14 proteins with higher expression and 26 with lower expression in the cells exposed for 12 h to MTX (pH gradients 4-7 and 6-11). Of note, the expression of the regulatory protein 14-3-3 protein epsilon was increased by a factor of two and three, after exposure to 1 and 10 µM MTX, respectively. At earlier time-points, 10 µM MTX increased intracellular ATP levels, while decreasing media lactate levels. At later stages (24 and 48 h), MTX-induced cytotoxicity was concentration and time-dependent, according to the MTT reduction and lactate dehydrogenase leakage assays, while caspase-9, -8 and -3 activities increased at 24 h. Regarding cellular redox status, total glutathione increased in 1 µM MTX (24 h), and that increase was dependent on gamma-glutamylcysteine synthetase activity. Meanwhile, for both 1 and 10 µM MTX, oxidized glutathione was significantly higher than control at 48 h. Moreover, MTX was able to significantly decrease proteasomal chymotrypsin-like activity in a concentration and time-independent manner. In summary, MTX significantly altered proteomic, energetic and oxidative stress homeostasis in cardiomyocytes at clinically relevant concentrations and our data clearly demonstrate that MTX causes early cardiotoxicity that needs further study.

Toxicity Evaluation and Biomarker Selection with Validated Reference Gene in Embryonic Zebrafish Exposed to Mitoxantrone.

Notwithstanding the widespread use and promising clinical value of chemotherapy, the pharmacokinetics, toxicology, and mechanism of mitoxantrone remains unclear. To promote the clinical value in the treatment of human diseases and the exploration of potential subtle effects of mitoxantrone, zebrafish embryos were employed to evaluate toxicity with validated reference genes based on independent stability evaluation programs. The most stable and recommended reference gene was gapdh, followed by tubα1b, for the 48 h post fertilization (hpf) zebrafish embryo mitoxantrone test, while both eef1a1l1 and rpl13α were recommended as reference genes for the 96 hpf zebrafish embryo mitoxantrone test. With gapdh as an internal control, we analyzed the mRNA levels of representative hepatotoxicity biomarkers, including fabp10a, gclc, gsr, nqo1, cardiotoxicity biomarker erg, and neurotoxicity biomarker gfap in the 48 hpf embryo mitoxantrone test. The mRNA levels of gclc, gsr, and gfap increased significantly in 10 and 50 µg/L mitoxantrone-treated 48 hpf embryos, while the transcript levels of fabp10a decreased in a dose-dependent manner, indicating that mitoxantrone induced hepatotoxicity and neurotoxicity. Liver hematoxylin⁻eosin staining and the spontaneous movement of embryos confirmed the results. Thus, the present research suggests that mitoxantrone induces toxicity during the development of the liver and nervous system in zebrafish embryos and that fabp10a is recommended as a potential biomarker for hepatotoxicity in zebrafish embryos. Additionally, gapdh is proposed as a reference gene for the 48 hpf zebrafish embryo mitoxantrone toxicity test, while eef1a1l1 and rpl13α are proposed as that for the 96 hpf test.

Naphthoquinoxaline metabolite of mitoxantrone is less cardiotoxic than the parent compound and it can be a more cardiosafe drug in anticancer therapy.

Mitoxantrone (MTX) is an antineoplastic agent used to treat several types of cancers and on multiple sclerosis, which shows a high incidence of cardiotoxicity. Still, the underlying mechanisms of MTX cardiotoxicity are poorly understood and the potential toxicity of its metabolites scarcely investigated. Therefore, this work aimed to synthesize the MTX-naphthoquinoxaline metabolite (NAPHT) and to compare its cytotoxicity to the parent compound in 7-day differentiated H9c2 cells using pharmacological relevant concentrations (0.01-5 µM). MTX was more toxic in equivalent concentrations in all cytotoxicity tests performed [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide reduction, neutral red uptake, and lactate dehydrogenase release assays] and times tested (24 and 48 h). Both MTX and NAPHT significantly decreased mitochondrial membrane potential in 7-day differentiated H9c2 cells after a 12-h incubation. However, energetic pathways were affected in a different manner after MTX or NAPHT incubation. ATP increased and lactate levels decreased after a 24-h incubation with MTX, whereas for the same incubation time and concentrations, NAPHT did not cause any significant effect. The increased activity of ATP synthase seems responsible for MTX-induced increases in ATP levels, as oligomycin (an inhibitor of ATP synthase) abrogated this effect on 5 µM MTX-incubated cells. 3-Methyladenine (an autophagy inhibitor) was the only molecule to give a partial protection against the cytotoxicity produced by MTX or NAPHT. To the best of our knowledge, this was the first broad study on NAPHT cardiotoxicity, and it revealed that the parent drug, MTX, caused a higher disruption in the energetic pathways in a cardiac model in vitro, whereas autophagy is involved in the toxicity of both compounds. In conclusion, NAPHT is claimed to largely contribute to MTX-anticancer properties; therefore, this metabolite should be regarded as a good option for a safer anticancer therapy since it is less cardiotoxic than MTX.

Evaluation of microRNAs-208 and 133a/b as differential biomarkers of acute cardiac and skeletal muscle toxicity in rats.

Conventional circulating biomarkers of cardiac and skeletal muscle (SKM) toxicity lack specificity and/or have a short half-life. MicroRNAs (miRNAs) are currently being assessed as biomarkers of tissue injury based on their long half-life in blood and selective expression in certain tissues. To assess the utility of miRNAs as biomarkers of cardiac and SKM injury, male Sprague-Dawley rats received a single dose of isoproterenol (ISO); metaproterenol (MET); allylamine (AAM); mitoxantrone (MIT); acetaminophen (APAP) or vehicle. Blood and tissues were collected from rats in each group at 4, 24 and 48h. ISO, MET, and AAM induced cardiac and SKM lesions and APAP induced liver specific lesions. There was no evidence of tissue injury with MIT by histopathology. Serum levels of candidate miRNAs were compared to conventional serum biomarkers of SKM/cardiac toxicity. Increases in heart specific miR-208 only occurred in rats with cardiac lesions alone and were increased for a longer duration than cardiac troponin and FABP3 (cardiac biomarkers). ISO, MET and AAM induced increases in MyL3 and skeletal muscle troponin (sTnl) (SKM biomarkers). MIT induced large increases in sTnl indicative of SKM toxicity, but sTnl levels were also increased in APAP-treated rats that lacked SKM toxicity. Serum levels of miR-133a/b (enriched in cardiac and SKM) increased following ISO, MET, AAM and MIT treatments but were absent in APAP-treated rats. Our results suggest that miR-133a/b are sensitive and specific markers of SKM and cardiac toxicity and that miR-208 used in combination with miR-133a/b can be used to differentiate cardiac from SKM toxicity.

Pathways of cardiac toxicity: comparison between chemotherapeutic drugs doxorubicin and mitoxantrone.

Anthracyclines, e.g., doxorubicin (DOX), and anthracenediones, e.g., mitoxantrone (MTX), are drugs used in the chemotherapy of several cancer types, including solid and non-solid malignancies such as breast cancer, leukemia, lymphomas, and sarcomas. Although they are effective in tumor therapy, treatment with these two drugs may lead to side effects such as arrhythmia and heart failure. At the same clinically equivalent dose, MTX causes slightly reduced cardiotoxicity compared with DOX. These drugs interact with iron to generate reactive oxygen species (ROS), target topoisomerase 2 (Top2), and impair mitochondria. These are some of the mechanisms through which these drugs induce late cardiomyopathy. In this review, we compare the cardiotoxicities of these two chemotherapeutic drugs, DOX and MTX. As described here, even though they share similarities in their modes of toxicant action, DOX and MTX seem to differ in a key aspect. DOX is a more redox-interfering drug, while MTX induces energy imbalance. In addition, DOX toxicity can be explained by underlying mechanisms that include targeting of Top2 beta, mitochondrial impairment, and increases in ROS generation. These modes of action have not yet been demonstrated for MTX, and this knowledge gap needs to be filled.

Dual photo- and pH-responsive supramolecular nanocarriers based on water-soluble pillar[6]arene and different azobenzene derivatives for intracellular anticancer drug delivery.

Two novel types of supramolecular nanocarriers fabricated by the amphiphilic host-guest inclusion complex formed from water-soluble pillar[6]arene (WP6) and azobenzene derivatives G1 or G2 have been developed, in which G1 is structurally similar to G2 but has an extra phenoxy group in its hydrophobic region. Supramolecular micelles can be initially formed by WP6 with G1, which gradually transform into layered structures with liquid-crystalline properties, whereas stable supramolecular vesicles are obtained from WP6 and G2, which exhibit dual photo- and pH-responsiveness. Notably, the resulting WP6⊃G2 vesicles can efficiently encapsulate anticancer drug mitoxantrone (MTZ) to achieve MTZ-loaded vesicles, which maintain good stability in a simulated normal physiological environment, whereas in an acid environment similar to that of tumor cells or with external UV irradiation, the encapsulated drug is promptly released. More importantly, cytotoxicity assay indicates that such vesicles have good biocompatibility and the MTZ-loaded vesicles exhibit comparable anticancer activity to free MTZ, especially with additional UV stimulus, whereas its cytotoxicity for normal cells was remarkably reduced. Flow cytometric analysis further confirms that the cancer cell death caused by MTZ-loaded vesicles is associated with apoptosis. Therefore, the dual pH- and UV-responsive supramolecular vesicles are a potential platform for controlled release and targeted anticancer drug delivery.

Subcellular Localization of Proteins Responding to Mitoxantrone-Induced DNA Damage in Leukaemic Cells.

The aim of the present study was to investigate the subcellular localization of proteins participating in the double-strand break response pathway - p53, Mdm2, p21 and Chk2. MOLT-4 cells were pre-treated with mitoxantrone in concentrations 1 nmol/l and 5 nmol/l. The trypan blue technique was used to determine cell viability and proliferation. Western blotting was used to evaluate changes in p53, Mdm2 and Chk2 protein expression and sandwich ELISA was used to evaluate changes in the p21 protein amount. After 1 nmol/l mitoxantrone cells did not die, but their ability to proliferate was decreased. The p53 protein was activated and phosphorylated at serines 15 and 392 and accumulated in the nucleus after 24 and 48 h. The Mdm2 protein was present in the cytoplasm with its maximal level after 8 and 16 h. The p21 protein was detected in the nucleus after 24 and 48 h. Increased levels of phosphorylated Chk2 at threonine 68 were observed in the cytoplasmic fraction after 24 and 48 h of mitoxantrone treatment. We used mitoxantrone as an inducer of double-strand breaks to bring new data about the subcellular distribution of proteins responding to DNA damage. In MOLT-4 cells, the p53 protein was activated. p53 was phosphorylated at serines 15 and 392 and accumulated in the nucleus. The Mdm2 protein was activated in advance to p53 and occurred in the cytoplasm. The p21 protein was present in the nucleus. Chk2 kinase was activated by the phosphorylation at threonine 68 and we observed increased levels of this protein in the cytoplasmic fraction.

The N-terminal pro-brain natriuretic peptide as a marker of mitoxantrone-induced cardiotoxicity in multiple sclerosis patients.

BACKGROUND AND PURPOSE: Mitoxantrone (MTX) has been shown to reduce progression of disability and number of clinical exacerbations in patients with progressive multiple sclerosis (MS). Prolonged administration of MTX, however, is limited by the risk of cardiotoxicity. Cardiac monitoring in MTX-treated patients includes usually measurement of left ventricular ejection fraction (LVEF) by means of echocardiography. The N-terminal pro-brain natriuretic peptide (NT-proBNP) represents a novel diagnostic tool in the assessment of heart failure. This study was aimed to evaluate the usefulness of NT-proBNP for early detection of MTX-induced cardiotoxicity in MS patients. MATERIALS AND METHODS: We measured the NT-proBNP plasma levels in 45 MS patients who completed 24-month MTX therapy and in 37 MS patients of control group. RESULTS: The median NT-proBNP plasma value was 15.12pg/mL. In 12 MTX-treated patients (27%), NT-proBNP plasma values were elevated, though this subgroup of patients neither clinical showed evidence of myocardial damage nor had the LVEF value <50%. In five patients with normal NT-proBNP, we observed LVEF decline >10%. We did not observe correlations between the NT-proBNP levels and patient age, MS duration, relapses index, Extended Disability Status Scale (EDSS), MTX single dose and the total cumulative dose of MTX. In 8 patients (22%) from control group, NT-proBNP plasma levels were also elevated. CONCLUSIONS: The results of our study confirm that MTX therapy is safe for carefully selected and closely monitored MS patients. We believe that serial evaluation of NT-proBNP levels (before, during and after MTX therapy) can identify MS patients at high risk for MTX-induced cardiotoxicity.

Synthesis and biological evaluation of new cytotoxic indazolo[4,3-gh]isoquinolinone derivatives.

A series of indazolo[4,3-gh]isoquinolinones derivatives have been synthesized to decrease cardiotoxic side effects in comparison to Mitoxantrone. The antiproliferative effects of different side chains were investigated and tested on at least four different cell lines of cervix, ovarian, CNS, NSCLC (non-small-cell lung cancer) and colon carcinoma. In addition to antiproliferative activities, influence on cell cycle and intercalation behavior have been tested.

The metabolic profile of mitoxantrone and its relation with mitoxantrone-induced cardiotoxicity.

Mitoxantrone (MTX) is an antitumor agent that causes cardiotoxicity in 18 % patients. The metabolic profile of MTX was assessed after incubation of 100 µM MTX with hepatic S9 fraction isolated from rats. The presence of MTX and its metabolites was also assessed in vivo through the analysis of liver and heart extracts of MTX-treated rats. The cytotoxic effects of MTX and MTX metabolites were evaluated in the H9c2 cells after 24-h incubation with MTX alone and MTX + metabolites. The influence of CYP450- and CYP2E1-mediated metabolism for the cytotoxicity of MTX was assessed after 96-h incubation with MTX (100 nM and 1 µM) in the presence/absence of CYP450 or CYP2E1 inhibitors. After 4-h incubation in supplemented S9 fraction, the MTX content was 35 % lower and 5 metabolites were identified: an acetoxy ester derivative (never described before), two glutathione conjugates, a monocarboxylic acid derivative, and the naphtoquinoxaline, the later commonly related to MTX pharmacological effects. The presence of MTX and naphtoquinoxaline metabolite was evidenced in vivo in liver and heart of MTX-treated rats. The cytotoxicity caused by MTX + metabolites was higher than that observed in the H9c2 cells incubated with non-metabolized MTX group. The co-incubation of MTX with CYP450 and CYP2E1 inhibitors partially prevented the cytotoxicity observed in the MTX groups incubated with H9c2 cells, highlighting that the metabolism of MTX is relevant for its undesirable effects. The naphtoquinoxaline metabolite is described in heart and liver in vivo, highlighting that this metabolite accumulates in these tissues. It was demonstrated that MTX P450-mediated metabolism contributed to MTX toxicity.

Genotoxic stress causes the accumulation of the splicing regulator Sam68 in nuclear foci of transcriptionally active chromatin.

DNA-damaging agents cause a multifaceted cellular stress response. Cells set in motion either repair mechanisms or programmed cell death pathways, depending on the extent of the damage and on their ability to withstand it. The RNA-binding protein (RBP) Sam68, which is up-regulated in prostate carcinoma, promotes prostate cancer cell survival to genotoxic stress. Herein, we have investigated the function of Sam68 in this cellular response. Mitoxantrone (MTX), a topoisomerase II inhibitor, induced relocalization of Sam68 from the nucleoplasm to nuclear granules, together with several other RBPs involved in alternative splicing, such as TIA-1, hnRNP A1 and the SR proteins SC35 and ASF/SF2. Sam68 accumulation in nuclear stress granules was independent of signal transduction pathways activated by DNA damage. Using BrU labelling and immunofluorescence, we demonstrate that MTX-induced nuclear stress granules are transcriptionally active foci where Sam68 and the phosphorylated form of RNA polymerase II accumulate. Finally, we show that MTX-induced relocalization of Sam68 correlates with changes in alternative splicing of its mRNA target CD44, and that MTX-induced CD44 splicing depends on Sam68 expression. These results strongly suggest that Sam68 is part of a RNA-mediated stress response of the cell that modulates alternative splicing in response to DNA damage.

An application of p-sulfonatocalix[6]arenes to attenuate cardiotoxicity of mitoxantrone in vitro: preparation, characterization and evaluation.

OBJECTIVES: In this study, p-sulfonatocalix[6]arenes (SCA6) was proposed to construct a host-guest complexation to carry mitoxantrone (MIT) to maintain its anti-proliferation effect on HepG2 cells as well as to attenuate cardiotoxicity on H9C2 cells as a nano-size drug delivery system. METHODS: SCA6 binding to MIT evidenced through competitive fluorescence titration method. The complex was characterized using UV-visible, Fourier transform infrared, and proton nuclear magnetic resonance (1H-NMR) spectroscopies and differential scanning calorimetry analysis. The cytotoxicity was examined by a cell counting kit-8 assay on six cells. High content analysis, cell apoptosis and cell cycle experiments were measured to investigate the mechanism of detoxification in H9C2. KEY FINDINGS: The host-guest complexation was formed with a stoichiometry ratio of 1:1. Cytotoxicity study demonstrated that MIT/SCA6 complex could improve the cell viability on H9C2, MCF-7, A549, Hek293 and L02 cells and remained cytotoxicity effect on HepG2 cells. High content analysis showed that MIT/SCA6 complex could enhance the cell viability, mitochondrial mass and mitochondrial membrane potential and ameliorate the nuclear swelling on H9C2 cells. Moreover, the complex were arrested in G0/G1 phase of the cell cycle and same with MIT, while the detoxication was attributed to reducing early apoptosis. CONCLUSIONS: The host-guest complexation between SCA6 and MIT had the ability to attenuate cardiotoxicity and provided a potential strategy for the application of soluble calixarenes in chemotherapy.

[Severe skin reaction with mucous membrane inflammation during MINE chemotherapy]. / Réaction cutanée sévère avec mucite au cours d'une chimiothérapie de type MINE.

BACKGROUND: MINE chemotherapy is used to treat refractory Hodgkin's disease. Cutaneous adverse effects of MINE regimen are uncommon and chiefly consist of erythema and oedema of the extremities. More recently, a number of cases of panniculitis and subcutaneous inflammatory oedema have been described. OBSERVATION: We report the case of a 17-year-old girl developing acute and painful oedema of the limbs with panniculitis of the trunk. This incident was associated with inflammatory lesions of mucous membrane, in particularly in the genital area and on the tongue. These signs occurred 7 days after initiation of MINE chemotherapy, with no other drugs being introduced. A drug-induced reaction was suspected due to the absence of any other aetiology, particularly infectious disease. The condition gradually improved with symptomatic pain therapy. The patient's chemotherapy was subsequently modified. DISCUSSION: The chronology of the symptoms, spontaneous improvement after the end of treatment, and the absence of other potential causative factors resulted in a hypothesis of a cutaneous adverse reaction to the MINE regimen. The signs could be due to capillary leak syndrome resulting from the toxicity of vinorelbine on endothelial cells. Dermatologists should be aware of these cutaneous adverse effects and of the inflammatory lesions of mucous membrane newly described herein.

Estrone-targeted liposomes for mitoxantrone delivery via estrogen receptor: In vivo targeting efficacy, antitumor activity, acute toxicity and pharmacokinetics.

Estrogen receptor (ER) is a potential target receptor for ER-positive cancer therapy including breast cancers, gastric cancers, and human acute myeloblastic leukaemia. In order to reduce the side-effects of mitoxantrone (MTO), estrone-targeted liposomes for MTO delivery via ER were designed for selectively targeting cancer cells. In previous studies, MTO-loaded estrogen receptor targeted and sterically stabilized liposome (ES-SSL-MTO; ES: estrone, is known to bind the ER) had been synthesized and showed a very high antiproliferative effect with IC50 value of 0.7 ng/mL. Based on these, further studies including in vivo targeting efficacy and antitumor activity, acute toxicity and pharmacokinetics of MTO liposomes were carried out. The results showed SSL (sterically stabilized liposome, PEGylated liposome, PEG: Polyethylene Glycol) could reduce drug metabolism, improve the stability of liposomes, prolong in vivo circulation time of drugs, reduce the toxicity of MTO. But SSL could not be enriched in tumor tissues. However, estrone (ES)-targeted liposomes could be delivered to tumor sites. ES-SSL could effectively enter into ER-expressing tumor cellsand be accumulated, prolong the circulation time in vivo, reduce side effects of drug. ES-SSL-MTO could provide higher bioavailability than MTO, enhance the anti-tumor effect and the safety of MTO, reduce the toxicity and side effects of MTO and improve the therapeutic effect of MTO. These facts proved ES-SSL is a useful tumor-targeting drug delivery system for MTO.

Subchronic administration of mitoxantrone and the influence of enzyme inhibitors on its induced cardiotoxicity in mice: role of NRF-2/CYP2E1.

OBJECTIVE: Mitoxantrone (MTX)- induced cardiotoxicity is a clinical concern that is limiting its use. The aim of this paper, therefore, was to investigate the subchronic administration of MTX plus nonspecific/specific inhibitors of CYP450/2E1, to assess the extent of oxidative-induced injury by measuring levels of oxidative cardiac and injury biomarkers in mice and to evaluate the effects of CYP2E1 on caspase 3 activity and nuclear factor erythroid 2-related factor-2 (NRF-2). MATERIALS AND METHODS: Mice (n = 32) were divided into four treatment groups of eight: control, MTX, MTX + 4-methlypyrazole (4MP) and MTX + disulfiram (Disf). After 6 weeks of treatments, blood and heart samples were collected. RESULTS: Liquid chromatography-mass spectrometry (LCMS) analysis of MTX-treated plasma samples revealed several metabolites with different retention times. Cardiac antioxidant enzymes and creatine kinase (CK) levels were not significantly different among the groups. However, cardiac troponin and caspase 3 activity were significantly raised, with increased CYP2E1 expressions and reduced NRF-2 expression. Tissue damage was observed in all the treatment groups, including MTX, leading to the conclusion that MTX-induced cardiotoxicity was mediated by CYP2E1 activity, which initiated caspase 3 production, and decreased NRF-2 expression. CONCLUSIONS: Therefore, agents that inhibit CPY2E1 expression might attenuate MTX-induced cardiotoxicity by increasing NRF-2 expression.

Exploring the aging effect of the anticancer drugs doxorubicin and mitoxantrone on cardiac mitochondrial proteome using a murine model.

Current cancer therapies are successfully increasing the lifespan of cancer patients. Nevertheless, cardiotoxicity is a serious chemotherapy-induced adverse side effect. Doxorubicin (DOX) and mitoxantrone (MTX) are cardiotoxic anticancer agents, whose toxicological mechanisms are still to be identified. This study focused on DOX and MTX's cardiac mitochondrial damage and their molecular mechanisms. As a hypothesis, we also sought to compare the cardiac modulation caused by 9 mg/kg of DOX or 6 mg/kg of MTX in young adult mice (3 months old) with old control mice (aged control, 18-20 months old) to determine if DOX- and MTX-induced damage had common links with the aging process. Cardiac homogenates and enriched mitochondrial fractions were prepared from treated and control animals and analyzed by immunoblotting and enzymatic assays. Enriched mitochondrial fractions were also characterized by mass spectrometry-based proteomics. Data obtained showed a decrease in mitochondrial density in young adults treated with DOX or MTX and aged control, as assessed by citrate synthase (CS) activity. Furthermore, aged control had increased expression of the peroxisome proliferator-activated receptor γ coactivator 1 α (PGC1α) and manganese superoxide dismutase (MnSOD). Regarding the enriched mitochondrial fractions, DOX and MTX led to downregulation of proteins related to oxidative phosphorylation, fatty acid oxidation, amino acid metabolic process, and tricarboxylic acid cycle. MTX had a greater impact on malate dehydrogenase (MDH2) and pyruvate dehydrogenase E1 component subunit α (PDHA1). No significant proteomic changes were observed in the enriched mitochondrial fractions of aged control when compared to young control. To conclude, DOX and MTX promoted changes in several mitochondrial-related proteins in young adult mice, but none resembling the aged phenotype.

Lack of ABCG2 expression and side population properties in human pluripotent stem cells.

The multidrug transporter ABCG2 in cell membranes enables various stem cells and cancer cells to efflux chemicals, including the fluorescent dye Hoechst 33342. The Hoechst(-) cells can be sorted out as a side population with stem cell properties. Abcg2 expression in mouse embryonic stem cells (ESCs) reduces accumulation of DNA-damaging metabolites in the cells, which helps prevent cell differentiation. Surprisingly, we found that human ESCs do not express ABCG2 and cannot efflux Hoechst. In contrast, trophoblasts and neural epithelial cells derived from human ESCs are ABCG2(+) and Hoechst(-). Human ESCs ectopically expressing ABCG2 become Hoechst(-), more tolerant of toxicity of mitoxantrone, a substrate of ABCG2, and more capable of self-renewal in basic fibroblast growth factor (bFGF)-free condition than control cells. However, Hoechst(low) cells sorted as a small subpopulation from human ESCs express lower levels of pluripotency markers than the Hoechst(high) cells. Similar results were observed with human induced pluripotent stem cells. Conversely, mouse ESCs are Abcg2(+) and mouse trophoblasts, Abcg2(-). Thus, absence of ABCG2 is a novel feature of human pluripotent stem cells, which distinguishes them from many other stem cells including mouse ESCs, and may be a reason why they are sensitive to suboptimal culture conditions.