Mitoxantrone Concentration Research Articles

Method for Testing of Drugs Belonging to Substrates and Inhibitors of the Transporter Protein BCRP on Caco-2 Cells

Introduction. Breast cancer resistance protein (BCRP) is an efflux membrane transporter that controls the pharmacokinetics of a large number of drugs. Its activity may change when taking some endo- and exogenous substances, thus making it a link in drug interactions.Aim. The aim of the study was to develop a method for testing of drugs for belonging to BCRP substrates and inhibitors in vitro.Materials and methods. The work was performed on Caco-2 cells overexpressing BCRP, the cultivation was performed in a transwell-system consisting of the apical and basolateral chambers. Cells were seeded at the bottom of the apical chamber, which is a semipermeable membrane. Primarily, the transport of BCRP substrates: methotrexate, mitoxantrone and quercetin was evaluated in the concentration range of 1, 5, 10, and 50 μM in the direction from the basal chamber to the apical one (Papp b-a) and in the opposite direction (Papp a-b). The ratio Papp b-a / Papp a-b more than «2» characterizes the participation of transporter proteins in the transcellular transport of substances. To confirm the participation of BCRP in their transport the experiment was carried out with the addition of a transporter inhibitor, reserpine, to the transport medium at a concentration of 50 μM. The concentration of substrates in the chambers was analyzed by HPLC-MS/MS.Results and their discussion. The addition of methotrexate (1 μM), mitoxantrone (1 μM), and quercetin (1–10 μM) to both the apical or basolateral chambers of the transwell-system, their content in the recipient chamber was not detected. When methotrexate concentration became 5 μM the Papp b-a / Papp a-b ratio was 3.38 ± 0.08, which indicates the involvement of transporters in its transfer. The addition of methotrexate to the donor chamber at concentrations of 10 and 50 μM, Papp b-a / Papp a-b decreased to values below «2». At mitoxantrone concentration of 5 μM Papp b-a / Papp a-b was 2.72 ± 0.16. An increase in the concentration to 10 μM led to an increase in Papp b-a / Papp a-b to 6.18 ± 0.08. With a substance content of 50 μM the indicator decreased but remained above the value «2». In the quercetin concentration of 50 microns, Papp b-a / Papp was below "2". Reserpine reduced Papp b-a / Papp a-b of methotrexate by 3.31 times (p = 0.0002), which indicates the elimination of asymmetry in the transport of the substance. At a mitoxantrone concentration of 10 microns, reserpine reduced its Papp b-a / Papp a-b by 3.36 times (p < 0.0001). The results indicate the participation of BCRP in the control of the transfer of both substances through the cellular monolayer.Conclusion. A method of testing drugs belonging to BCRP substrates and inhibitors using methotrexate (5 μM) and mitoxantrone (10 μM) as marker substrates and reserpine (50 μM) as inhibitor was developed and tested on Caco-2 cells.

In vivo evaluation of Sono-chemo therapy via hollow gold nanoshells conjugated to mitoxantrone on breast cancer.

Conventional methods of cancer treatment include surgery, chemotherapy, radiation therapy, and immunotherapy. Chemotherapy, as one of the main methods of cancer treatment, due to the lack of targeted distribution of the drug in tumor tissues, is not able to destroy cancer cells and also affects healthy tissues and causes serious side effects in patients. Sonodynamic therapy (SDT) is a promising strategy for non-invasive treatment of deep solid cancer tumors. In this study, for the first time, the sono-sensitive activity of mitoxantrone was investigated and then mitoxantrone (MTX) was conjugated to hollow gold nanostructure (HGN) to improve the efficiency of in vivo SDT. Firstly, after the synthesis of hollow gold nanoshells and the PEGylation process, conjugation of MTX was performed. Then, after evaluating the toxicity of the treatment groups in vitro, in order to perform an in vivo study, 56 male Balb/c mice that had been tumorized by subcutaneous injection of 4T1 cells were divided into eight groups of breast tumor model. Ultrasonic irradiation (US) conditions including intensity of 1.5 W/cm2 (with a frequency of 800 kHz, 5 min), MTX concentration of 2 μM, and HGN dose of 2.5 mg/kg (unit of animal weight) were used. The results show that administration of PEG-HGN-MTX caused a slight reduction in tumor size and growth compared with free MTX. Ultrasound also improved the therapeutic effect of the gold nanoshell in treated groups, and the HGN-PEG-MTX-US treated groups were able to significantly reduce and control tumor size and growth. The findings also show that MTX and HGN can be used as sonosensitizers in SDT. Also, HGN-PEG-MTX can act as a sono-chemotherapy agent for the combination of sonodynamic therapy and chemotherapy for in vivo breast tumors.

A cooperative transition from the semi-flexible to the flexible regime of polymer elasticity: Mitoxantrone-induced DNA condensation

We report a high cooperative transition from the semi-flexible to the flexible regime of polymer elasticity during the interaction of the DNA molecule with the chemotherapeutic drug Mitoxantrone (MTX). By using single molecule force spectroscopy, we show that the force-extension curves of the DNA-MTX complexes deviate from the typical worm-like chain behavior as the MTX concentration in the sample increases, becoming straight lines for sufficiently high drug concentrations. The behavior of the radius of gyration of the complexes as a function of the bound MTX concentration was used to quantitatively investigate the cooperativity of the condensation process. The present methodology can be promptly applied to other ligands that condense the DNA molecule upon binding, opening new possibilities in the investigation of this type of process and, more generally, in the investigation of phase transitions in polymer physics.

Thermostability of DNA Complexes with Mitoxantrone at Small Fillings

Thermostability of DNA complexes with anti-tumorous compound — mitoxantrone — at 0.011[Formula: see text]M and 0.11 M NaCl ionic strengths of solution by spectrophotometric and microcalorimetric methods has been studied. It was shown that at small fillings when one molecule of mitoxantrone corresponds to 250 or more base pairs of DNA, the thermostability of complexes strongly depends on the solution’s ionic strength; moreover, at an ionic strength of 0.011 M NaCl the melting temperature dependence on mitoxantrone concentration passes through a minimum. In the mentioned region of mitoxantrone concentration the melting enthalpy of the complexes increases linearly with mitoxantrone concentration enhancement. The observed phenomenon, which is not observed for ethidium bromide complexes with DNA, is qualitatively explained by the increasing of coil-shaped state of DNA–mitoxantrone complex due to additional freedom of rotation of mitoxantrone side groups.

In situ preparation of magnetic Fe3O4 nanoparticles inside nanoporous poly(l-glutamic acid)/chitosan microcapsules for drug delivery

The magnetic polymer microcapsules, as a promising environmental stimuli-responsive delivery vehicle, have been increasingly exploited to tackle the problem of remotely navigated delivery. This study presented a novel design and fabrication of magnetic poly(l-glutamic acid)/chitosan (PGA/CS) microcapsules. Magnetic Fe3O4 nanoparticles were in situ synthesized inside nanoporous PGA/CS microcapsules and resultant magnetic PGA/CS microcapsules were characterized. Mitoxantrone (MTX), an antineoplastic drug, was chosen as a water-soluble model drug to research the loading and release properties of the microcapsules. The results showed the carboxylate groups of PGA within polyelectrolyte walls could be used as binding sites for the absorption of iron ions and reaction sites for the synthesis of magnetic nanoparticles. Magnetic PGA/CS microcapsules were dissected using a dual-beam scanning electron microscope/focused ion beam (SEM/FIB) for morphological and microstructural examination. It was found that Fe3O4 nanoparticles with size of about 10nm were homogeneously dispersed in the polymer matrix and adhered to the pore walls of the microcapsules. Increasing the concentration of iron ions led to an increasing loading content of Fe3O4 nanoparticles and an increase in the resultant magnetization. The magnetic PGA/CS microcapsules could be easily manipulated by an external magnetic field. The MTX loading capacity depended on loading time and MTX concentration. The high loading could be ascribed to spontaneous deposition of MTX induced by electrostatic interaction. The microcapsules exhibited sustained release behavior. The MTX release from microcapsules could be best described using Korsmeyer–Peppas and Baker–Lonsdale models, indicating the diffusion mechanism of drug release from both PGA/CS microcapsules and magnetic PGA/CS microcapsules. Therefore, the novel magnetic PGA/CS microcapsules are expected to find application in drug delivery systems because of the properties of magnetic sensitivity, high drug loading and sustained release.

Mitoxantrone-Iron Oxide Biodistribution in Blood, Tumor, Spleen, and Liver—Magnetic Nanoparticles in Cancer Treatment

Magnetic drug targeting is a new treatment principle for tumors using cytostatics coupled to ferromagnetic nanoparticles and extracorporeal magnets. Higher concentrations in tumor tissue with lower systemic concentrations and without damage of healthy organs should be achieved. n = 42 adult Wag/Rij rats were transfected with rhabdomyosarcoma R(1)H in their right gastrocnemius muscle. In the biodistribution trial (n = 36) concentrations of mitoxantrone-iron oxide with and without an extracorporeal 0.6 tesla magnet and regular mitoxantrone were measured in plasma and tumor tissue for one- and two-dose administration. In the plasma iron trial (n = 6) iron concentrations were measured in plasma before, during, and up to 30 min after drug administration. Seven days after the trial liver, spleen and tumor samples were obtained and histologically assessed. Mitoxantrone iron-oxide concentration in plasma was significantly (P < 0.05) lower when a magnet was placed over the tumor area and as low as uncoupled mitoxantrone. Mitoxantrone concentration in tumor tissue was always significantly higher with magnetic drug targeting when compared with uncoupled mitoxantrone. Two doses resulted in drug accumulation in tumor tissue. Plasma iron concentrations rose when the drug was first administered. Plasma levels fell below the starting level with a magnet applied. A rebound phenomenon with rising iron concentrations was observed after the magnet was removed. Tumors showed fresh necrosis and liver and spleen had detectable iron depositions but no necrosis 7 d after treatment. No allergies or toxic reactions were observed. We showed that magnetic drug targeting achieves higher concentrations of cytostatics in tumor tissue compared with blood. During magnetic drug targeting, iron particles are quickly sliced and kept in the tumor area. Organs of the reticuloendothelial system are not affected by cytostatic damage.

Evaluation of mitoxantrone-loaded albumin microspheres following intraperitoneal administration to rats

Mitoxantrone has demonstrated therapeutic efficacy in the regional treatment of intraperitoneal malignancies. However, severe local toxicity was dose limiting. Consequently, a new injectable sustained delivery formulation of mitoxantrone has been developed: the drug was incorporated (8.2 w/w%) in highly hydrophilic albumin microspheres. In vitro drug release profile was modified by matrix crosslinking extent. The extractable amount of residual crosslinking agent (glutaraldehyde) in the microspheres was lower than 6 ppm. Mitoxantrone concentration in peritoneal fluid and plasma was determined up to 72 h after intraperitoneal administration of 30, 60 and 120 mg mitoxantrone per m 2 body surface area as solution and in the form of a dispersion containing mitoxantrone-loaded microspheres to rats. Data analysis revealed sustained release of mitoxantrone from microspheres into peritoneal fluid in all dosage groups. The initial high drug levels in peritoneal fluid and plasma observed after application of mitoxantrone in the solution form were prevented by administration of the drug incorporated in microspheres. However, tumoricidal drug levels in peritoneal fluid were maintained over a comparable time span. In addition, preliminary toxicity data suggest a superior local tolerability of mitoxantrone-loaded microspheres. The dose of intraperitoneally administered mitoxantrone might be increased from 30 to 60 mg per m 2 body surface area using the slow release formulation. In conclusion, the described microsphere drug delivery system for mitoxantrone might overcome dose-limiting drug toxicity.

The combined effects of IL-3 and PSC 833 on daunorubicin- and mitoxantrone cytotoxicity in two growth factor-dependent leukemic cell lines.

In the present study it was investigated whether and by which mechanisms the co-administration of interleukin-3 (IL-3) and the P-glycoprotein blocker PSC 833 can augment mitoxantrone (MX) and daunorubicin (DAU) cytotoxicity in two human growth factor dependent P-glycoprotein (P-gp) positive myeloid leukemic cell lines, Mo-7 and GF-D8. Cytotoxicity was determined in MTT assay. Increased cytotoxicity occurred in Mo-7 cells preincubated with 24h IL-3 followed by 1 h MX (cell survival: 85% +/- 6 vs 68% +/- 2, at 0.05 microM MX, P < 0.005) or DAU (79% +/- 8 vs 62% +/- 9 at 0.8 microg/ml DAU, P < 0.05). Similar results were obtained for the GF-D8 cell line. In this cell line, at 0.5 microM MX the cell survival decreased from 84% +/- 13 to 61% +/- 19 (P < 0.05) and at 5.0 microg/ml DAU from 102% +/- 8 to 69% +/- 5, (P < 0.002). The IL-3 administration did not affect the P-gp and bcl-2 protein expression, cellular MX concentration or MX efflux but coincided with an increased percentage of cells in S-phase and topoisomerase II (topo II)-alpha mRNA and topo II activity especially in the Mo-7 cell line. PSC 833 enhanced DAU cytotoxicity in both cell lines. The administration of IL-3 plus PSC 833 in the Mo-7 cell line resulted in an additive effect on DAU cytotoxicity. At 0.8 microg/ml DAU and 2 microg/ml PSC 833, the percentage surviving cells decreased from 62% +/- 9 in the absence of IL-3 to 37% +/- 3 in the presence of IL-3 (P < 0.01). The additive effect of combined treatment was most pronounced in GF-D8 cells which also had the highest P-gp expression. In contrast, PSC 833 did not modulate the MX effects, irrespective of the presence of IL-3. In summary, the results demonstrate that the combined administration of IL-3 and PSC 833 can enhance the cytotoxic effects of DAU but not MX in these P-gp positive cell lines whereas the effects of MX could be modulated by factors which influence topo II activity.

Pharmacokinetics of Mitoxantrone in Plasma and Leukemic Cells During Treatment of Patients with Acute Non-Lymphocytic Leukemia

The pharmacokinetics of mitoxantrone was studied in the plasma and peripheral leukemic cells in 14 patients receiving induction treatment for acute non-lymphocytic leukemia. The disappearance of mitoxantrone from plasma was biphasic with a mean distribution half life of 10.4 minutes and a mean elimination half life of 17.1 hours. At the end of the infusion, the mean mitoxantrone concentration in leukemic cells was 9.6 times (range 2.1-22.7) higher than in plasma and remained relatively stable in the cells until the next infusion following 24 hours after the first. The area under the concentration-time curve (AUC0-20h) in leukemic cells was 23-191 times larger than the AUC0-α in plasma. In the individual patient there was no correlation between the AUC in leukemic cells and AUC in plasma. Since mitoxantrone acts intracellularly, concentrations in the leukemic cells might predict the antileukemic effect better than plasma concentrations.

Potentiation of cytotoxicity of mitoxantrone toward CHO-K1 cells in vitro by dipyridamole.

Dipyridamole (DP), a clinically used vasodilator and an antiplatelet compound, augmented the activity of the anticancer drug mitoxantrone (MXN) toward chinese hamster ovary (CHO-K1) cells in culture. Clonogenic assays indicated that DP (1.0, 2.5, and 5.0 microM) decreased the survival of cells treated with MXN (5 to 25 nM) in a dose-dependent manner. Further, DP (1 and 5 microM) decreased the MXN concentration required for 50% inhibition of cell growth from 3.2 to 1.8 and from 3.0 to 0.5 nM, respectively, over a period of 3 days. DP (10 microM) increased the accumulation of MXN by 1.8-fold in exponentially growing cells exposed to MXN. The enhanced levels of MXN in CHO-K1 cells in the presence of the chemosensitizer may account for the potentiation of MXN-cytotoxicity by DP.

Characteristics of mitoxantrone loading on liposomes

The characteristics of mitoxantrone loading on liposomes was investigated. It was found that negatively charged liposomes at the ionic strength 0.154 and pH 5.7 gave a high loading efficiency. It should be noted that the surface charges of the liposomes were subject to change from negative to positive by the influence of the interacted mitoxantrone. The neutral liposomes demonstrated a profile of loading efficiency similar to that of the negatively charged liposomes against the added mitoxantrone concentration; however, they had a lower magnitude of the loading efficiency. In the positively charged liposomes the loading efficiency increased with the added mitoxantrone concentration. At a mitoxantrone concentration of 26 μmol/ml, the loading efficiency was similar to that of the negatively charged liposomes. The -effect of galactocerebroside' on the loading efficiency increased with added mitoxantrone concentration. The liposomes with 1.0 molar ratio galactocerebroside resulted in the highest loading efficiency in high concentrations of mitoxantrone. The loading efficiency had no dependence on the phospholipid concentration used for making liposomal membrane. An almost identical value of the loading efficiency was obtained when ultracentrifuge and dialysis methods were used to separate the mitoxantrone-containing liposomes from free drugs. The effect of vortexed and sonicated liposomes on the loading efficiency gave a similar result although the particle size was different.

Human central nervous system and plasma pharmacology of mitoxantrone.

Mitoxantrone 5-6 mg/m2 was administered IV to 10 consenting patients prior to surgical resection of an intracerebral tumor. Plasma pharmacokinetic parameters were calculated and concentration of mitoxantrone in intracerebral tumors was determined. Concentrations of mitoxantrone were also determined in autopsy tissues of one of the patients who expired 192 days after receiving the drug. The plasma pharmacokinetics were best described by a 3 compartment model, with a tl/2 gamma of 4.74 +/- 5.53 h. Mitoxantrone concentrations in the intracerebral tumors were potentially cytotoxic and ranged from 4 to 322 ng/g. In all but one case, mitoxantrone concentration was higher in tumor than in concurrent plasma samples. There was no obvious relation between tumor mitoxantrone concentration and peak plasma mitoxantrone concentration or time from mitoxantrone administration to tumor removal. Low grade gliomas and viable tumors tended to have lower mitoxantrone concentrations than did other tumor types and necrotic tumors. In the patient undergoing autopsy, highest mitoxantrone concentrations were found in liver, thyroid and heart.

Mitoxantrone sensitivity of human hematopoietic cell lines

A panel of 18 human hematopoietic cell lines was tested for sensitivity to mitoxantrone, an anthracenedione derivative with antitumor activity in vitro and in vivo. Cells in logarithmic growth phase were tested for mitoxantrone at concentrations of 0.1–1000 ng/ml for 1–3 days. Cell growth inhibition was profound for all lines at a mitoxantrone concentration of 10 ng/ml, whereas a variable response was seen at a concentration of 1 ng/ml. Inhibition of cell proliferation was mainly depending on arrest at the G2 phase of the cell cycle (or S + G2 at higher mitoxantrone concentrations), together with a cytotoxic effect. The cell cycle block was found in all lines tested, whereas the cytotoxic activity varied much between different cell lines.

Inhibition of endogenous respiration in rat heart slices as a measure of mitoxantrone cardiac toxicity.

Anthracycline antibiotics are among the more active agents currently available for the treatment of solid tumors. Their use is limited by the development of severe cardiomyopathy. This makes it necessary to subject new anticancer agents to laboratory and clinical tests in order to determine their capacity to damage cardiac cells. Mitoxantrone is an anthracenedione derivative with structural and functional similarities to doxorubicin. Comparative trials on rat suggest that mitoxantrone induced cardiac toxicity might be less marked than that caused by doxorubicin, while some clinical studies report a significant incidence of cardiac failure following mitoxantrone administration. The present study was undertaken to evaluate (in vitro) the effect of increasing concentrations of mitoxantrone on respiratory control, measuring oxygen uptake of rat heart slices in a Warburg manometric apparatus. Over a period of 60 min, cellular endogenous respiration was progressively inhibited by increasing mitoxantrone concentration (from 5 to 20 microM). The date show that the rate of oxygen uptake levels off with time at all concentrations except the lowest (5 microM) and with the control. Oxygen uptake values range from 2.87 to 1.41 microliter/mg dry weight and differ significantly as compared to controls (p less than 0.01). The values of oxygen uptake between 0 and 20 min show that a linear correlation is approached by all the date group and if we consider an exponential relation (i.e., log slope versus dose), the linear correlation coefficient is somewhat improved (r = -0.939). These results indicate that mitoxantrone, in a manner analogous to doxorubicin, inhibits cellular respiration and impairs the cardiac respiratory control. This impairment is probably one of the aspects of cellular damage leading to cardiac failure. Moreover, in a recent study we observed the same pathological lesions induced by doxorubicin in the heart of rats treated with mitoxantrone.

Acute effects of mitoxantrone on the template activity of isolated nuclei from the T-47D human breast tumor cell line

The effect of mitoxantrone on the template activity of nuclei isolated from the T-47D human breast tumor cell line was investigated. The results suggest that mitoxantrone significantly inhibits total RNA synthesis of these nuclei in a concentration-dependent manner. At low drug concentrations (10 −9 M and 10 −7 M) RNA synthesis was inhibited by 21.9% and 41% compared to control values, respectively. Greater inhibition was observed when the mitoxantrone concentration was increased to 10 −5 M or 10 −4 M (56% and 77%, respectively). Experiments utilizing α-amanitin revealed that mitoxantrone inhibits RNA polymerase II activity in a concentration-dependent manner.