RESUMEN
Anthracyclines, such as doxorubicin (adriamycin), daunorubicin, or epirubicin, rank among the most effective agents in classical anticancer chemotherapy. However, cardiotoxicity remains the main limitation of their clinical use. Topoisomerase IIß has recently been identified as a plausible target of anthracyclines in cardiomyocytes. We examined the putative topoisomerase IIß selective agent XK469 as a potential cardioprotective and designed several new analogs. In our experiments, XK469 inhibited both topoisomerase isoforms (α and ß) and did not induce topoisomerase II covalent complexes in isolated cardiomyocytes and HL-60, but induced proteasomal degradation of topoisomerase II in these cell types. The cardioprotective potential of XK469 was studied on rat neonatal cardiomyocytes, where dexrazoxane (ICRF-187), the only clinically approved cardioprotective, was effective. Initially, XK469 prevented daunorubicin-induced toxicity and p53 phosphorylation in cardiomyocytes. However, it only partially prevented the phosphorylation of H2AX and did not affect DNA damage measured by Comet Assay. It also did not compromise the daunorubicin antiproliferative effect in HL-60 leukemic cells. When administered to rabbits to evaluate its cardioprotective potential in vivo, XK469 failed to prevent the daunorubicin-induced cardiac toxicity in either acute or chronic settings. In the following in vitro analysis, we found that prolonged and continuous exposure of rat neonatal cardiomyocytes to XK469 led to significant toxicity. In conclusion, this study provides important evidence on the effects of XK469 and its combination with daunorubicin in clinically relevant doses in cardiomyocytes. Despite its promising characteristics, long-term treatments and in vivo experiments have not confirmed its cardioprotective potential.
Asunto(s)
Antraciclinas , Quinoxalinas , Inhibidores de Topoisomerasa II , Ratas , Animales , Conejos , Inhibidores de Topoisomerasa II/toxicidad , Inhibidores de Topoisomerasa II/uso terapéutico , Antraciclinas/toxicidad , Antraciclinas/uso terapéutico , Cardiotoxicidad , Daunorrubicina/toxicidad , Daunorrubicina/uso terapéutico , Doxorrubicina/toxicidad , Antibióticos Antineoplásicos/toxicidad , ADN-Topoisomerasas de Tipo II/metabolismo , ADN-Topoisomerasas de Tipo II/uso terapéutico , Daño del ADNRESUMEN
Ferritin, a naturally occurring iron storage protein, has gained significant attention as a drug delivery platform due to its inherent biocompatibility and capacity to encapsulate therapeutic agents. In this study, we successfully genetically engineered human H ferritin by incorporating 4 or 6 tryptophan residues per subunit, strategically oriented towards the inner cavity of the nanoparticle. This modification aimed to enhance the encapsulation of hydrophobic drugs into the ferritin cage. Comprehensive characterization of the mutants revealed that only the variant carrying four tryptophan substitutions per subunit retained the ability to disassemble and reassemble properly. As a proof of concept, we evaluated the loading capacity of this mutant with ellipticine, a natural hydrophobic indole alkaloid with multimodal anticancer activity. Our data demonstrated that this specific mutant exhibited significantly higher efficiency in loading ellipticine compared to human H ferritin. Furthermore, to evaluate the versatility of this hydrophobicity-enhanced ferritin nanoparticle as a drug carrier, we conducted a comparative study by also encapsulating doxorubicin, a commonly used anticancer drug. Subsequently, we tested both ellipticine and doxorubicin-loaded nanoparticles on a promyelocytic leukemia cell line, demonstrating efficient uptake by these cells and resulting in the expected cytotoxic effect.
Asunto(s)
Antineoplásicos , Elipticinas , Nanopartículas , Humanos , Ferritinas/genética , Ferritinas/química , Apoferritinas/genética , Triptófano , Antineoplásicos/farmacología , Antineoplásicos/química , Sistemas de Liberación de Medicamentos , Portadores de Fármacos/química , Doxorrubicina/farmacología , Doxorrubicina/química , Nanopartículas/química , Interacciones Hidrofóbicas e Hidrofílicas , Línea Celular TumoralRESUMEN
Labile redox-active iron ions have been implicated in various neurodegenerative disorders, including the Parkinson's disease (PD). Iron chelation has been successfully used in clinical practice to manage iron overload in diseases such as thalassemia major; however, the use of conventional iron chelators in pathological states without systemic iron overload remains at the preclinical investigative level and is complicated by the risk of adverse outcomes due to systemic iron depletion. In this study, we examined three clinically-used chelators, namely, desferrioxamine, deferiprone and deferasirox and compared them with experimental agent salicylaldehyde isonicotinoyl hydrazone (SIH) and its boronate-masked prochelator BSIH for protection of differentiated PC12 cells against the toxicity of catecholamines 6-hydroxydopamine and dopamine and their oxidation products. All the assayed chelating agents were able to significantly reduce the catecholamine toxicity in a dose-dependent manner. Whereas hydrophilic chelator desferrioxamine exerted protection only at high and clinically unachievable concentrations, deferiprone and deferasirox significantly reduced the catecholamine neurotoxicity at concentrations that are within their plasma levels following standard dosage. SIH was the most effective iron chelator to protect the cells with the lowest own toxicity of all the assayed conventional chelators. This favorable feature was even more pronounced in prochelator BSIH that does not chelate iron unless its protective group is cleaved in disease-specific oxidative stress conditions. Hence, this study demonstrated that while iron chelation may have general neuroprotective potential against catecholamine auto-oxidation and toxicity, SIH and BSIH represent promising lead molecules and warrant further studies in more complex animal models.
Asunto(s)
Quelantes del Hierro , Sobrecarga de Hierro , Animales , Catecolaminas/farmacología , Deferasirox/farmacología , Deferiprona/farmacología , Deferoxamina/farmacología , Dopamina/farmacología , Hierro/farmacología , Quelantes del Hierro/farmacología , Estrés Oxidativo , Oxidopamina/farmacología , Células PC12 , RatasRESUMEN
Angiotensin-converting enzyme inhibitors (ACEis) have been used to treat anthracycline (ANT)-induced cardiac dysfunction, and they appear beneficial for secondary prevention in high-risk patients. However, it remains unclear whether they truly prevent ANT-induced cardiac damage and provide long-lasting cardioprotection. The present study aimed to examine the cardioprotective effects of perindopril on chronic ANT cardiotoxicity in a rabbit model previously validated with the cardioprotective agent dexrazoxane (DEX) with focus on post-treatment follow-up (FU). Chronic cardiotoxicity was induced by daunorubicin (DAU; 3 mg/kg/week for 10 weeks). Perindopril (0.05 mg/kg/day) was administered before and throughout chronic DAU treatment. After the completion of treatment, significant benefits were observed in perindopril co-treated animals, particularly full prevention of DAU-induced mortality and prevention or significant reductions in cardiac dysfunction, plasma cardiac troponin T (cTnT) levels, morphological damage, and most of the myocardial molecular alterations. However, these benefits significantly waned during 3 weeks of drug-free FU, which was not salvageable by administering a higher perindopril dose. In the longer (10-week) FU period, further worsening of left ventricular function and morphological damage occurred together with heart failure (HF)-related mortality. Continued perindopril treatment in the FU period did not reverse this trend but prevented HF-related mortality and reduced the severity of the progression of cardiac damage. These findings contrasted with the robust long-lasting protection observed previously for DEX in the same model. Hence, in the present study, perindopril provided only temporary control of ANT cardiotoxicity development, which may be associated with the lack of effects on ANT-induced and topoisomerase II ß (TOP2B)-dependent DNA damage responses in the heart.
Asunto(s)
Inhibidores de la Enzima Convertidora de Angiotensina/uso terapéutico , Cardiotoxicidad/prevención & control , Daunorrubicina/efectos adversos , Perindopril/uso terapéutico , Animales , Antibióticos Antineoplásicos , Cardiotoxicidad/tratamiento farmacológico , Cardiopatías/inducido químicamente , Cardiopatías/prevención & control , Insuficiencia Cardíaca/tratamiento farmacológico , Insuficiencia Cardíaca/mortalidad , Masculino , Conejos , Troponina T/sangreRESUMEN
BACKGROUND: Anthracycline-induced heart failure has been traditionally attributed to direct iron-catalyzed oxidative damage. Dexrazoxane (DEX)-the only drug approved for its prevention-has been believed to protect the heart via its iron-chelating metabolite ADR-925. However, direct evidence is lacking, and recently proposed TOP2B (topoisomerase II beta) hypothesis challenged the original concept. METHODS: Pharmacokinetically guided study of the cardioprotective effects of clinically used DEX and its chelating metabolite ADR-925 (administered exogenously) was performed together with mechanistic experiments. The cardiotoxicity was induced by daunorubicin in neonatal ventricular cardiomyocytes in vitro and in a chronic rabbit model in vivo (n=50). RESULTS: Intracellular concentrations of ADR-925 in neonatal ventricular cardiomyocytes and rabbit hearts after treatment with exogenous ADR-925 were similar or exceeded those observed after treatment with the parent DEX. However, ADR-925 did not protect neonatal ventricular cardiomyocytes against anthracycline toxicity, whereas DEX exhibited significant protective effects (10-100 µmol/L; P<0.001). Unlike DEX, ADR-925 also had no significant impact on daunorubicin-induced mortality, blood congestion, and biochemical and functional markers of cardiac dysfunction in vivo (eg, end point left ventricular fractional shortening was 32.3±14.7%, 33.5±4.8%, 42.7±1.0%, and 41.5±1.1% for the daunorubicin, ADR-925 [120 mg/kg]+daunorubicin, DEX [60 mg/kg]+daunorubicin, and control groups, respectively; P<0.05). DEX, but not ADR-925, inhibited and depleted TOP2B and prevented daunorubicin-induced genotoxic damage. TOP2B dependency of the cardioprotective effects was probed and supported by experiments with diastereomers of a new DEX derivative. CONCLUSIONS: This study strongly supports a new mechanistic paradigm that attributes clinically effective cardioprotection against anthracycline cardiotoxicity to interactions with TOP2B but not metal chelation and protection against direct oxidative damage.
Asunto(s)
Antraciclinas/farmacología , Cardiotoxicidad/prevención & control , Dexrazoxano/farmacología , Insuficiencia Cardíaca/tratamiento farmacológico , Inhibidores de Topoisomerasa II/metabolismo , Antraciclinas/efectos adversos , Antibióticos Antineoplásicos/efectos adversos , Antibióticos Antineoplásicos/farmacología , Cardiotoxicidad/tratamiento farmacológico , Cardiotoxicidad/metabolismo , ADN-Topoisomerasas de Tipo II/efectos adversos , ADN-Topoisomerasas de Tipo II/metabolismo , Daunorrubicina/metabolismo , Daunorrubicina/farmacología , Dexrazoxano/efectos adversos , Cardiopatías/tratamiento farmacológico , Humanos , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/metabolismo , Estrés Oxidativo/efectos de los fármacosRESUMEN
The anthracycline (ANT) anticancer drugs such as doxorubicin or daunorubicin (DAU) can cause serious myocardial injury and chronic cardiac dysfunction in cancer survivors. A bisdioxopiperazine agent dexrazoxane (DEX) has been developed as a cardioprotective drug to prevent these adverse events, but it is uncertain whether it is the best representative of the class. The present study used a rabbit model of chronic ANT cardiotoxicity to examine another bisdioxopiperazine compound called GK-667 (meso-(butane-2,3-diylbis(2,6-dioxopiperazine-4,1-diyl))bis(methylene)-bis(2-aminoacetate) hydrochloride), a water-soluble prodrug of ICRF-193 (meso-4,4'-(butan-2,3-diyl)bis(piperazine-2,6-dione)), as a potential cardioprotectant. The cardiotoxicity was induced by DAU (3 mg/kg, intravenously, weekly, 10 weeks), and GK-667 (1 or 5 mg/kg, intravenously) was administered before each DAU dose. The treatment with GK-667 was well tolerated and provided full protection against DAU-induced mortality and left ventricular (LV) dysfunction (determined by echocardiography and LV catheterization). Markers of cardiac damage/dysfunction revealed minor cardiac damage in the group co-treated with GK-667 in the lower dose, whereas almost full protection was achieved with the higher dose. This was associated with similar prevention of DAU-induced dysregulation of redox and calcium homeostasis proteins. GK-667 dose-dependently prevented tumor suppressor p53 (p53)-mediated DNA damage response in the LV myocardium not only in the chronic experiment but also after single DAU administration. These effects appear essential for cardioprotection, presumably because of the topoisomerase IIß (TOP2B) inhibition provided by its active metabolite ICRF-193. In addition, GK-667 administration did not alter the plasma pharmacokinetics of DAU and its main metabolite daunorubicinol (DAUol) in rabbits in vivo. Hence, GK-667 merits further investigation as a promising drug candidate for cardioprotection against chronic ANT cardiotoxicity.
Asunto(s)
Cardiomiopatías/prevención & control , Daño del ADN , Dicetopiperazinas/farmacología , Miocitos Cardíacos/efectos de los fármacos , Profármacos/farmacología , Inhibidores de Topoisomerasa II/farmacología , Disfunción Ventricular Izquierda/prevención & control , Función Ventricular Izquierda/efectos de los fármacos , Remodelación Ventricular/efectos de los fármacos , Animales , Cardiomiopatías/inducido químicamente , Cardiomiopatías/metabolismo , Cardiomiopatías/fisiopatología , Cardiotoxicidad , Enfermedad Crónica , Daunorrubicina , Modelos Animales de Enfermedad , Fibrosis , Células HL-60 , Humanos , Masculino , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/patología , Conejos , Proteína p53 Supresora de Tumor/metabolismo , Disfunción Ventricular Izquierda/metabolismo , Disfunción Ventricular Izquierda/fisiopatologíaRESUMEN
Cardioprotective activity of dexrazoxane (ICRF-187), the only clinically approved drug against anthracycline-induced cardiotoxicity, has traditionally been attributed to its iron-chelating metabolite. However, recent experimental evidence suggested that the inhibition and/or depletion of topoisomerase IIß (TOP2B) by dexrazoxane could be cardioprotective. Hence, we evaluated a series of dexrazoxane analogues and found that their cardioprotective activity strongly correlated with their interaction with TOP2B in cardiomyocytes, but was independent of their iron chelation ability. Very tight structure-activity relationships were demonstrated on stereoisomeric forms of 4,4'-(butane-2,3-diyl)bis(piperazine-2,6-dione). In contrast to its rac-form 12, meso-derivative 11 (ICRF-193) showed a favorable binding mode to topoisomerase II in silico, inhibited and depleted TOP2B in cardiomyocytes more efficiently than dexrazoxane, and showed the highest cardioprotective efficiency. Importantly, the observed ICRF-193 cardioprotection did not interfere with the antiproliferative activity of anthracycline. Hence, this study identifies ICRF-193 as the new lead compound in the development of efficient cardioprotective agents.
Asunto(s)
Cardiotónicos/uso terapéutico , Cardiotoxicidad/tratamiento farmacológico , Piperazinas/uso terapéutico , Inhibidores de Topoisomerasa II/uso terapéutico , Animales , Animales Recién Nacidos , Cardiotónicos/síntesis química , Cardiotónicos/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , ADN-Topoisomerasas de Tipo II/metabolismo , Daunorrubicina/toxicidad , Dicetopiperazinas , Humanos , Simulación del Acoplamiento Molecular , Simulación de Dinámica Molecular , Estructura Molecular , Miocitos Cardíacos/efectos de los fármacos , Piperazinas/síntesis química , Piperazinas/metabolismo , Unión Proteica , Ratas Wistar , Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/metabolismo , Relación Estructura-Actividad , Inhibidores de Topoisomerasa II/síntesis química , Inhibidores de Topoisomerasa II/metabolismoRESUMEN
The bisdioxopiperazine topoisomerase IIß inhibitor ICRF-193 has been previously identified as a more potent analog of dexrazoxane (ICRF-187), a drug used in clinical practice against anthracycline cardiotoxicity. However, the poor aqueous solubility of ICRF-193 has precluded its further in vivo development as a cardioprotective agent. To overcome this issue, water-soluble prodrugs of ICRF-193 were prepared, their abilities to release ICRF-193 were investigated using a novel UHPLC-MS/MS assay, and their cytoprotective effects against anthracycline cardiotoxicity were tested in vitro in neonatal ventricular cardiomyocytes (NVCMs). Based on the obtained results, the bis(2-aminoacetoxymethyl)-type prodrug GK-667 was selected for advanced investigations due to its straightforward synthesis, sufficient solubility, low cytotoxicity and favorable ICRF-193 release. Upon administration of GK-667 to NVCMs, the released ICRF-193 penetrated well into the cells, reached sufficient intracellular concentrations and provided effective cytoprotection against anthracycline toxicity. The pharmacokinetics of the prodrug, ICRF-193 and its rings-opened metabolite was estimated in vivo after administration of GK-667 to rabbits. The plasma concentrations of ICRF-193 reached were found to be adequate to achieve cardioprotective effects in vivo. Hence, GK-667 was demonstrated to be a pharmaceutically acceptable prodrug of ICRF-193 and a promising drug candidate for further evaluation as a potential cardioprotectant against chronic anthracycline toxicity.
Asunto(s)
Antraciclinas/efectos adversos , Cardiotónicos/farmacología , Cardiotoxicidad/tratamiento farmacológico , ADN-Topoisomerasas de Tipo II/metabolismo , Dicetopiperazinas/farmacología , Piperazina/farmacología , Inhibidores de Topoisomerasa II/farmacología , Animales , Cardiotónicos/química , Cardiotoxicidad/metabolismo , Dexrazoxano/química , Dexrazoxano/farmacología , Dicetopiperazinas/química , Masculino , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/metabolismo , Piperazina/química , Profármacos/química , Profármacos/farmacología , Conejos , Razoxano/química , Razoxano/farmacología , Inhibidores de Topoisomerasa II/química , Agua/químicaRESUMEN
Bisdioxopiperazine agent dexrazoxane (ICRF-187) has been the only effective and approved drug for prevention of chronic anthracycline cardiotoxicity. However, the structure-activity relationships (SARs) of its cardioprotective effects remain obscure owing to limited investigation of its derivatives/analogs and uncertainties about its mechanism of action. To fill these knowledge gaps, we tested the hypothesis that dexrazoxane derivatives exert cardioprotection via metal chelation and/or modulation of topoisomerase IIß (Top2B) activity in chronic anthracycline cardiotoxicity. Dexrazoxane was alkylated in positions that should not interfere with the metal-chelating mechanism of cardioprotective action; that is, on dioxopiperazine imides or directly on the dioxopiperazine ring. The protective effects of these agents were assessed in vitro in neonatal cardiomyocytes. All studied modifications of dexrazoxane molecule, including simple methylation, were found to abolish the cardioprotective effects. Because this challenged the prevailing mechanistic concept and previously reported data, the two closest derivatives [(±)-4,4'-(propane-1,2-diyl)bis(1-methylpiperazine-2,6-dione) and 4-(2-(3,5-dioxopiperazin-1-yl)ethyl)-3-methylpiperazine-2,6-dione] were thoroughly scrutinized in vivo using a rabbit model of chronic anthracycline cardiotoxicity. In contrast to dexrazoxane, both compounds failed to protect the heart, as demonstrated by mortality, cardiac dysfunction, and myocardial damage parameters, although the pharmacokinetics and metal-chelating properties of their metabolites were comparable to those of dexrazoxane. The loss of cardiac protection was shown to correlate with their abated potential to inhibit and deplete Top2B both in vitro and in vivo. These findings suggest a very tight SAR between bisdioxopiperazine derivatives and their cardioprotective effects and support Top2B as a pivotal upstream druggable target for effective cardioprotection against anthracycline cardiotoxicity. SIGNIFICANCE STATEMENT: This study has revealed the previously unexpected tight structure-activity relationships of cardioprotective effects in derivatives of dexrazoxane, which is the only drug approved for the prevention of cardiomyopathy and heart failure induced by anthracycline anticancer drugs. The data presented in this study also strongly argue against the importance of metal-chelating mechanisms for the induction of this effect and support the viability of topoisomerase IIß as an upstream druggable target for effective and clinically translatable cardioprotection.
Asunto(s)
Antraciclinas/efectos adversos , Cardiotoxicidad/tratamiento farmacológico , ADN-Topoisomerasas de Tipo II/metabolismo , Dexrazoxano/farmacología , Corazón/efectos de los fármacos , Sustancias Protectoras/farmacología , Inhibidores de Topoisomerasa II/farmacología , Animales , Cardiomiopatías/tratamiento farmacológico , Cardiomiopatías/metabolismo , Línea Celular Tumoral , Células HL-60 , Humanos , Masculino , Modelos Animales , Miocardio/metabolismo , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/metabolismo , Conejos , Ratas , Ratas Wistar , Relación Estructura-ActividadRESUMEN
Although proteasome inhibitors (PIs) are modern targeted anticancer drugs, they have been associated with a certain risk of cardiotoxicity and heart failure (HF). Recently, PIs have been combined with anthracyclines (ANTs) to further boost their anticancer efficacy. However, this raised concerns regarding cardiac safety, which were further supported by several in vitro studies on immature cardiomyocytes. In the present study, we investigated the toxicity of clinically used PIs alone (bortezomib (BTZ), carfilzomib (CFZ)) as well as their combinations with an ANT (daunorubicin (DAU)) in both neonatal and adult ventricular cardiomyocytes (NVCMs and AVCMs) and in a chronic rabbit model of DAU-induced HF. Using NVCMs, we found significant cytotoxicity of both PIs around their maximum plasma concentration (cmax) as well as significant augmentation of DAU cytotoxicity. In AVCMs, BTZ did not induce significant cytotoxicity in therapeutic concentrations, whereas the toxicity of CFZ was significant and more profound. Importantly, neither PI significantly augmented the cardiotoxicity of DAU despite even more profound proteasome-inhibitory activity in AVCMs compared with NVCMs. Furthermore, in young adult rabbits, no significant augmentation of chronic ANT cardiotoxicity was noted with respect to any functional, morphological, biochemical or molecular parameter under study, despite significant inhibition of myocardial proteasome activity. Our experimental data show that combination of PIs with ANTs is not accompanied by an exaggerated risk of cardiotoxicity and HF in young adult animal cardiomyocytes and hearts.
Asunto(s)
Antraciclinas/toxicidad , Antineoplásicos/toxicidad , Cardiotoxicidad/etiología , Inhibidores de Proteasoma/toxicidad , Animales , Antraciclinas/administración & dosificación , Antineoplásicos/administración & dosificación , Protocolos de Quimioterapia Combinada Antineoplásica/toxicidad , Bortezomib/administración & dosificación , Bortezomib/toxicidad , Daunorrubicina/administración & dosificación , Daunorrubicina/toxicidad , Relación Dosis-Respuesta a Droga , Masculino , Miocitos Cardíacos/efectos de los fármacos , Oligopéptidos/administración & dosificación , Oligopéptidos/toxicidad , Complejo de la Endopetidasa Proteasomal/efectos de los fármacos , Complejo de la Endopetidasa Proteasomal/metabolismo , Inhibidores de Proteasoma/administración & dosificación , Conejos , Ratas , Ratas WistarRESUMEN
Sobuzoxane (MST-16) is an approved anticancer agent, a pro-drug of bisdioxopiperazine analog ICRF-154. Due to the structural similarity of ICRF-154 to dexrazoxane (ICRF-187), MST-16 deserves attention as a cardioprotective drug. This study presents for the first time UHPLC-MS/MS assay of MST-16, ICRF-154 and its metabolite (EDTA-diamide) in cell culture medium, buffer, plasma and cardiac cells and provides data on MST-16 bioactivation under conditions relevant to investigation of cardioprotection of this drug. The analysis of these compounds that differ considerably in their lipophilicity was achieved on the Zorbax SB-Aq column using a mixture of aqueous ammonium formate and methanol as a mobile phase. The biological samples were either diluted or precipitated with methanol, which was followed by acidification for the assay of MST-16. The method was validated for determination of all compounds in the biological materials. The application of the method for analysis of samples from in vitro experiments provided important findings, namely, that (1) MST-16 is quickly decomposed in biological environments, (2) the cardiac cells actively metabolize MST-16, and (3) MST-16 readily penetrates into the cardiac cells and is converted into ICRF-154 and EDTA-diamide. These data are useful for the in-depth examination of the cardioprotective potential of this drug.
Asunto(s)
Antineoplásicos/análisis , Ácido Edético/química , Piperazinas/análisis , Razoxano/análogos & derivados , Animales , Antineoplásicos/metabolismo , Células Cultivadas , Cromatografía Líquida de Alta Presión , Miocitos Cardíacos/citología , Miocitos Cardíacos/metabolismo , Ratas , Ratas Wistar , Razoxano/química , Razoxano/metabolismo , Espectrometría de Masas en TándemRESUMEN
Solid-phase microextraction (SPME) is an alternative method to dialysis and ultrafiltration for the determination of plasma protein binding (PPB) of drugs. It is particularly advantageous for complicated analytes where standard methods are not applicable. Di-2-pyridylketone 4-cyclohexyl-4-methyl-3-thiosemicarbazone (DpC) is a lead compound of novel thiosemicarbazone anti-cancer drugs, which entered clinical trials in 2016. However, this agent exhibited non-specific binding on filtration membranes and had intrinsic chelation activity, which precluded standard PPB methods. In this study, using a simple and fast procedure, we prepared novel SPME fibers for extraction of DpC based on a metal-free, silicon string support, covered with C18 sorbent. Reproducibility of the preparation process was demonstrated by the percent relative standard deviation (RSD) of ≤ 9.2% of the amount of DpC extracted from PBS by several independently prepared fibers. The SPME procedure was optimized by evaluating extraction and desorption time profiles. Suitability of the optimized protocol was verified by examining reproducibility, linearity, and recovery of DpC extracted from PBS or plasma. All samples extracted by SPME were analyzed using an optimized and validated UHPLC-MS/MS method. The developed procedure was applied to the in vitro determination of PPB of DpC at two clinically relevant concentrations (500 and 1000 ng/mL). These studies showed that DpC is highly bound to plasma proteins (PPB ≥ 88%) and this did not differ significantly between both concentrations tested. This investigation provides novel data in the applicability of SPME for the determination of PPB of chelators, as well as useful information for the clinical development of DpC. Graphical abstract.
Asunto(s)
Antineoplásicos/metabolismo , Proteínas Sanguíneas/metabolismo , Piridinas/metabolismo , Microextracción en Fase Sólida/instrumentación , Tiosemicarbazonas/metabolismo , Adsorción , Animales , Bovinos , Cromatografía Líquida de Alta Presión/métodos , Diseño de Equipo , Unión Proteica , Ratas , Silicio/química , Microextracción en Fase Sólida/métodos , Espectrometría de Masas en Tándem/métodosRESUMEN
Heart is a particularly sensitive organ to iron overload and cardiomyopathy due to the excessive cardiac iron deposition causes most deaths in disorders such as beta-thalassemia major. Free or loosely bound iron ions readily cycle between ferrous and ferric states and catalyze Haber-Weiss reaction that yields highly reactive and toxic hydroxyl radicals. Treatment with iron chelators (desferrioxamine, deferiprone, and deferasirox) substantially improved cardiovascular morbidity and mortality in iron overloaded patients. Furthermore, iron chelators have been studied in various cardiovascular disorders with known or presumed oxidative stress roles (e.g., ischemia/reperfusion injury) also in patients with normal body iron contents. The pharmacodynamic and pharmacokinetic properties of these chelators are critical for effective therapy. For example, the widely clinically used but hydrophilic chelator desferrioxamine suffers from poor plasma membrane permeability, which means that high and clinically unachievable concentrations/doses must be employed to obtain cardioprotection. Therefore, small-molecular and lipophilic chelators with oral availability are more suitable for this purpose, particularly in states without systemic iron overload. Apart from agents that are already used in clinical practice, aroylhydrazone iron chelators, namely salicylaldehyde isonicotinoyl hydrazone (SIH), have provided promising results. However, the use of classical iron-chelating agents is associated with a risk of toxicity due to indiscriminate iron depletion. Recent studies have therefore focused on "masked" prochelators that have little or no affinity for iron until site-specific activation by reactive oxygen species.
Asunto(s)
Cardiotónicos/uso terapéutico , Quelantes del Hierro/uso terapéutico , Sobrecarga de Hierro/tratamiento farmacológico , Profármacos/uso terapéutico , Animales , Cardiotónicos/farmacología , Línea Celular , Femenino , Humanos , Hierro/química , Quelantes del Hierro/farmacología , Masculino , Profármacos/farmacologíaRESUMEN
Salicylaldehyde isonicotinoyl hydrazone (SIH) is a small molecule and lipophilic chelating agent that firmly binds ferric ions from the cellular labile iron pool and is able to protect various tissues against oxidative damage. Previously, SIH possessed the best ratio of cytoprotective efficiency to toxicity among various iron chelators, including the desferrioxamine, deferiprone, and deferasirox used in clinical practice. Here, we prepared a series of 2,6-dihydroxybenzaldehyde aroylhydrazones as SIH analogues with an additional hydroxyl group that can be involved in the chelation of metal ions. Compound JK-31 (2,6-dihydroxybenzaldehyde 4-chlorobenzohydrazone) showed the best cytoprotective efficiency among the studied compounds including SIH. This compound significantly protected H9c2 cardiomyoblast cells against oxidative stress induced by various pro-oxidants, such as hydrogen peroxide, tert-butyl hydroperoxide, paraquat, epinephrine, N-acetyl- p-benzoquinone imine (a toxic metabolite of paracetamol), and 6-hydroxydopamine. The exceptional cytoprotective activity of JK-31 was confirmed using epifluorescence microscopy, where JK-31-treated H9c2 cells maintained a higher mitochondrial inner membrane potential in the presence of a lethal dose of hydrogen peroxide than was observed with cells treated with SIH. Hence, this study demonstrates the deleterious role of free iron ions in oxidative injury and the potential of 2,6-dihydroxybenzaldehyde aroylhydrazones in the prevention of various types of cardiac injuries, highlighting the need for further investigations into these compounds.
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Aldehídos/química , Benzaldehídos/química , Hidrazonas/química , Quelantes del Hierro/química , Estrés Oxidativo , Aldehídos/farmacología , Animales , Línea Celular , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Estabilidad de Medicamentos , Humanos , Hidrazonas/farmacología , Hidrólisis , Quelantes del Hierro/farmacología , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Miocitos Cardíacos/citología , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/metabolismo , Estrés Oxidativo/efectos de los fármacos , Sustancias Protectoras/química , Sustancias Protectoras/farmacología , RatasRESUMEN
Aroylhydrazone iron chelators such as salicylaldehyde isonicotinoyl hydrazone (SIH) protect various cells against oxidative injury and display antineoplastic activities. Previous studies have shown that a nitro-substituted hydrazone, namely, NHAPI, displayed markedly improved plasma stability, selective antitumor activity, and moderate antioxidant properties. In this study, we prepared four series of novel NHAPI derivatives and explored their iron chelation activities, anti- or pro-oxidant effects, protection against model oxidative injury in the H9c2 cell line derived from rat embryonic cardiac myoblasts, cytotoxicities to the corresponding noncancerous H9c2 cells, and antiproliferative activities against the MCF-7 human breast adenocarcinoma and HL-60 human promyelocytic leukemia cell lines. Nitro substitution had both negative and positive effects on the examined properties, and we identified new structure-activity relationships. Naphthyl and biphenyl derivatives showed selective antiproliferative action, particularly in the breast adenocarcinoma MCF-7 cell line, where they exceeded the selectivity of the parent compound NHAPI. Of particular interest is a compound prepared from 2-hydroxy-5-methyl-3-nitroacetophenone and biphenyl-4-carbohydrazide, which protected cardiomyoblasts against oxidative injury at 1.8 ± 1.2 µM with 24-fold higher selectivity than SIH. These compounds will serve as leads for further structural optimization and mechanistic studies.
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Antineoplásicos/farmacología , Antioxidantes/farmacología , Hidrazonas/farmacología , Quelantes del Hierro/farmacología , Animales , Antineoplásicos/síntesis química , Antineoplásicos/química , Antineoplásicos/toxicidad , Antioxidantes/síntesis química , Antioxidantes/química , Antioxidantes/toxicidad , Línea Celular Tumoral , Estabilidad de Medicamentos , Humanos , Hidrazonas/síntesis química , Hidrazonas/química , Hidrazonas/toxicidad , Quelantes del Hierro/síntesis química , Quelantes del Hierro/química , Quelantes del Hierro/toxicidad , Radioisótopos de Hierro , Estructura Molecular , Estrés Oxidativo/efectos de los fármacos , Ratas , Relación Estructura-ActividadRESUMEN
Liposomes are able to load a range of cargos and have been used for drug delivery applications, including for stimuli-triggered drug release. Here, we describe an approach for imparting near infrared (NIR) light-triggered release to pre-formed liposomes, using a newly-synthesized cationic, amphiphilic phthalocyanine. When simply mixed in aqueous solution with cargo-loaded liposomes, the cationic amphiphilic phthalocyanine, but not a cationic hydrophilic azaphthalocyanine, spontaneously incorporates into the liposome bilayer. This enables subsequent release of loaded cargo (doxorubcin or basic orange) upon irradiation with NIR light. The rate of release could be altered by varying the amount of photosensitizer added to the liposomes. In the absence of NIR light exposure, stable cargo loading of the liposomes was maintained. Introduction.
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Dexrazoxane (DEX), the only cardioprotectant approved against anthracycline cardiotoxicity, has been traditionally deemed to be a prodrug of the iron-chelating metabolite ADR-925. However, pharmacokinetic profile of both agents, particularly with respect to the cells and tissues essential for its action (cardiomyocytes/myocardium), remains poorly understood. The aim of this study is to characterize the conversion and disposition of DEX to ADR-925 in vitro (primary cardiomyocytes) and in vivo (rabbits) under conditions where DEX is clearly cardioprotective against anthracycline cardiotoxicity. Our results show that DEX is hydrolyzed to ADR-925 in cell media independently of the presence of cardiomyocytes or their lysate. Furthermore, ADR-925 directly penetrates into the cells with contribution of active transport, and detectable concentrations occur earlier than after DEX incubation. In rabbits, ADR-925 was detected rapidly in plasma after DEX administration to form sustained concentrations thereafter. ADR-925 was not markedly retained in the myocardium, and its relative exposure was 5.7-fold lower than for DEX. Unlike liver tissue, myocardium homogenates did not accelerate the conversion of DEX to ADR-925 in vitro, suggesting that myocardial concentrations in vivo may originate from its distribution from the central compartment. The pharmacokinetic parameters for both DEX and ADR-925 were determined by both noncompartmental analyses and population pharmacokinetics (including joint parent-metabolite model). Importantly, all determined parameters were closer to human than to rodent data. The present results open venues for the direct assessment of the cardioprotective effects of ADR-925 in vitro and in vivo to establish whether DEX is a drug or prodrug.
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Cardiotónicos/farmacocinética , Dexrazoxano/farmacocinética , Etilenodiaminas/farmacocinética , Glicina/análogos & derivados , Miocitos Cardíacos/metabolismo , Animales , Cardiotónicos/sangre , Cardiotónicos/metabolismo , Cardiotónicos/farmacología , Dexrazoxano/sangre , Dexrazoxano/metabolismo , Dexrazoxano/orina , Etilenodiaminas/metabolismo , Glicina/metabolismo , Glicina/farmacocinética , Conejos , Ratas , Distribución TisularRESUMEN
The PHARMINE ("Pharmacy Education in Europe") project studied the organisation of pharmacy education, practice and legislation in the European Union (EU) with the objectives of evaluating to what degree harmonisation had taken place with the EU, and producing documents on each individual EU member state. Part of this work was in the form of a survey of pharmacy education, practice, and legislation in the various member states. We will publish the individual member state surveys as reference documents. This paper presents the results of the PHARMINE survey on pharmacy education, training, and practice in the Czech Republic. Czech community pharmacies sell and provide advice on Rx and Over-the-counter (OTC) medicines; they also provide diagnostic services (e.g., blood pressure measurement). Pharmacists (lékárník in Czech) study for five years and graduate with a Magister (Mgr., equivalent to M.Pharm.) degree. The Mgr. diploma is the only requirement for registration as a pharmacist. Pharmacists can own and manage community pharmacies, or work as responsible pharmacists in pharmacies. All practising pharmacists must be registered with the Czech Chamber of Pharmacists. The ownership of a community pharmacy is not restricted to members of the pharmacy profession; the majority of pharmacies are organised into various pharmacy chains. There are two universities providing higher education in pharmacy in the Czech Republic: the Faculty of Pharmacy in Hradec Kralove, Charles University, which was established in 1969, and the Faculty of Pharmacy of the University of Veterinary and Pharmaceutical Sciences in Brno, which was established in 1991. The pharmacy curriculum is organized as a seamless, fully integrated, five-year master degree course. There is a six-month traineeship supervised by the university, which usually takes place during the fifth year. Thus, the pharmacy curriculum is organised in accordance with the EU directive on sectoral professions that lays down the imperatives for pharmacy education, training, and practice in the various member states of the EU. Currently, no specialisation courses are available at the university level. Specialisation is organised in the form of postgraduate, continuing professional development by the Czech Chamber of Pharmacists, and delivered by the Institute of Postgraduate Education for Health Professions.
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Novel dexrazoxane derivative JR-311 was prepared to investigate structure-activity relationships and mechanism(s) of protection against anthracycline cardiotoxicity. Its cardioprotective, antiproliferative, iron (Fe) chelation and inhibitory and/or depletory activities on topoisomerase IIbeta (TOP2B) were examined and compared with dexrazoxane. While in standard assay, JR-311 failed in both cardioprotection and depletion of TOP2B, its repeated administration to cell culture media led to depletion of TOP2B and significant protection of isolated rat neonatal ventricular cardiomyocytes from daunorubicin-induced damage. This effect was explained by a focused analytical investigation that revealed rapid JR-311 decomposition, resulting in negligible intracellular concentrations of the parent compound but high exposure of cells to the decomposition products, including Fe-chelating JR-H2. Although chemical instability is an obstacle for the development of JR-311, this study identified a novel dexrazoxane analogue with preserved pharmacodynamic properties, contributed to the investigation of structure-activity relationships and suggested that the cardioprotection of bis-dioxopiperazines is likely attributed to TOP2B activity of the parent compound rather than Fe chelation of their hydrolytic metabolites/degradation products. Moreover, this study highlights the importance of early stability testing during future development of novel dexrazoxane analogues.
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Cardiotónicos/farmacología , ADN-Topoisomerasas de Tipo II/metabolismo , Dexrazoxano/farmacología , Quelantes del Hierro/farmacología , Animales , Animales Recién Nacidos , Antraciclinas/toxicidad , Cardiotoxicidad/tratamiento farmacológico , Cardiotoxicidad/etiología , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Daunorrubicina/toxicidad , Dexrazoxano/análogos & derivados , Dicetopiperazinas/farmacología , Hierro/metabolismo , Miocitos Cardíacos/efectos de los fármacos , Ratas , Ratas Wistar , Relación Estructura-ActividadRESUMEN
OBJECTIVES: Rutin, quercetin-3-O-rutinoside, a natural flavonol glycoside, has shown various in vitro benefits with potential use treating human diseases, especially cardiovascular system disorders. Antioxidant properties are assumed to underlie the majority of these benefits. Yet rutin pro-oxidant properties have been reported as well. Our research group has recently shown aggravating effects on isoprenaline (ISO)-induced cardiotoxicity in Wistar:Han rats after 24â hours. METHODS: This study was designed to examine in more detail the reasons for the negative effects of rutin (11.5 and 46â mg/kg, i.v.) after administration of ISO (100â mg/kg, s.c.) in rats within 2â hours of continuous experiment and in the H9c2 cardiomyoblast-derived cell line. RESULTS: Like our previous findings, rutin did not (11.5 or 46â mg/kg, i.v.) reduce the ISO-induced mortality within 2â hours although the lower dose significantly reduced cardiac troponin T (cTnT) and partly improved the histological findings. In contrast, the higher dose increased the mortality in comparison with solvent (1.26% w/v sodium bicarbonate). This was not caused by any specific haemodynamic disturbances. It appears to be associated with oxidative stress as rutin enhanced intracellular reactive oxygen species formation in vitro and had the tendency to increase it in vivo. CONCLUSIONS: Rutin, likely due to its pro-oxidative effects, can exacerbate catecholamine cardiotoxicity depending on the dose used.