Self-Assembled Dual-Targeted Epirubicin-Hybrid Polydopamine Nanoparticles for Combined Chemo-Photothermal Therapy of Triple-Negative Breast Cancer.

PURPOSE: Folic acid and cyclic arginylglycylaspartic acid peptides were introduced to the surface of negatively charged lipid-coated hybrid polydopamine-cysteine cores for the delivery of epirubicin (EPI) (E/PCF-NPs). The combined chemo-photothermal therapy using E/PCF-NPs for triple-negative breast cancer was evaluated. MATERIALS AND METHODS: The temperature elevation and thermal toxicity of nanoparticles were studied. The morphology and properties of E/PCF-NPs were characterized by transmission electron microscopy, scanning electron microscopy, and atomic force microscopy. Physicochemical properties, including particle size, zeta potential, drug loading, entrapment efficiency (EE%), stability and in vitro release, were determined. The cell viability, reactive oxygen species (ROS) levels, ratios of oxidized nicotinamide adenine dinucleotide to its reduced form (NAD+/NADH), apoptosis assays, and cellular uptake of E/PCF-NPs were determined on 4T1 cells. Pharmacokinetic studies and tissue distributions were performed and detected by an ultra-high performance liquid chromatography/mass spectrometry system. The antitumor effects of E/PCF-NPs under near-infrared (NIR) laser irradiation were also evaluated. RESULTS: The sphere-like morphology of E/PCF-NPs showed a high EE%, uniform size of 106.7 nm, remarkable stability, and highly improved cytotoxicity under NIR laser, when compared to that of photothermal treatment alone. In vitro release of EPI from E/PCF-NPs was pH sensitive, and a greater response was achieved under NIR laser irradiation. Compared to chemotherapy or photothermal treatment alone, the combined treatment in vitro significantly inhibited the survival rate of 4T1 cells to 17.7%, induced ROS generation, and reduced NAD+/NADH significantly. Treatment with E/PCF-NPs under irradiation induced 4T1 cell apoptosis in approximately 93.6% cells. In vitro cellular uptake of E/PCF-NPs was time-dependent. The long-circulating and higher tumor accumulation of E/PCF-NPs resulted in complete ablation of breast tumor tissue through the enhanced photothermal effect by NIR laser irradiation-mediated cell apoptosis. CONCLUSION: E/PCF-NPs show enhanced anti-cancer effects due to synergistic effects of chemotherapy with photothermal therapy and may be potential therapeutic agents for cancer treatment.

Use of a Glass Membrane Pumping Emulsification Device Improves Systemic and Tumor Pharmacokinetics in Rabbit VX2 Liver Tumor in Transarterial Chemoembolization.

PURPOSE: To evaluate the phamacokinetics of epirubicin in conventional transarterial chemoembolization using a developed pumping emulsification device with a microporous glass membrane in VX2 rabbits. MATERIALS AND METHODS: Epirubicin solution (10 mg/mL) was mixed with ethiodized oil (1:2 ratio) using the device or 3-way stopcock. Forty-eight rabbits with VX2 liver tumor implanted 2 weeks prior to transarterial chemoembolization were divided into 2 groups: a device group (n = 24) and a 3-way-stopcock group (n = 24). Next, 0.5 mL of emulsion was injected into the hepatic artery, followed by embolization using 100-300-µm microspheres. The serum epirubicin concentrations (immediately after, 5 minutes after, and 10 minutes after) and the tumor epirubicin concentrations (20 minutes after and 48 hours after) were measured after transarterial chemoembolization. Histopathologic evaluation was performed with a fluorescence microscope. RESULTS: The area under the curve and maximum concentrations of epirubicin in plasma were 0.45 ± 0.18 µg min/mL and 0.13 ± 0.06 µg/mL, respectively, in the device group and 0.71 ± 0.45 µg min/mL and 0.22 ± 0.17 µg/mL, respectively, in the 3-way-stopcock group (P = .013 and P = .021, respectively). The mean epirubicin concentrations in VX2 tumors at 48 hours in the device group and the 3-way-stopcock group were 13.7 ± 6.71 and 7.72 ± 3.26 µg/g tissue, respectively (P = .013). The tumor necrosis ratios at 48 hours were 62 ± 11% in the device group and 51 ± 13% in the 3-way-stopcock group (P = .039). CONCLUSIONS: Conventional transarterial chemoembolization using the pumping emulsification device significantly improved the pharmacokinetics of epirubicin compared to the current standard technique using a 3-way stopcock.

Pharmacogenetics of adjuvant breast cancer treatment with cyclophosphamide, epirubicin and 5-fluorouracil.

PURPOSE: Most adjuvant breast cancer treatment regimens include the combination of an anthracycline (epirubicin or doxorubicin) and the alkylating agent cyclophosphamide. This study sought to investigate the influence of pharmacogenetics on the pharmacokinetics and metabolism of these agents. METHODS: Blood samples were taken from patients treated with cyclophosphamide (n = 51) and epirubicin (n = 35), with or without 5-fluorouracil (5-FU). The pharmacokinetics and metabolism of the three drugs were investigated, together with pharmacogenetic investigations for cyclophosphamide and epirubicin. Cyclophosphamide and its metabolites and also epirubicin and epirubicinol were measured in plasma. DNA was extracted from whole blood and genotyping performed using RT-PCR. RESULTS: Patients with at least one variant CYP2C19*17 allele had a longer CP half-life (p = 0.007), as did homozygous variants for the CYP2B6*6 allele. There was no significant effect of GSTP1, CYP2B6*2, CYP2B6*5 or CYP2C19*2 on any pharmacokinetic parameter of CP. An NQO2 exonic SNP was associated with a higher exposure to epirubicinol relative to epirubicin (p = 0.011). Other polymorphic variants of NQO1, carbonyl reductase, UGT enzymes and transporters had no influence on epirubicin or its metabolite. CONCLUSION: Overall, pharmacogenetic factors had only a minor influence on cyclophosphamide or anthracycline-based adjuvant therapy of breast cancer.

Characterization of endogenous G-CSF and the inverse correlation to chemotherapy-induced neutropenia in patients with breast cancer using population modeling.

PURPOSE: Neutropenia is a severe adverse-event of chemotherapeutics. Neutrophils (ANC) are mainly regulated by granulocyte colony stimulating factor (G-CSF). The aim was to characterize the dynamics between endogenous G-CSF and ANC over time following chemotherapy. METHODS: Endogenous G-CSF and ANC were monitored in forty-nine breast cancer patients treated with sequential adjuvant 5-fluorouracil-epirubicin-cyclophosphamide and docetaxel. RESULTS: During treatment courses ANC was transiently decreased and was reflected in an endogenous G-CSF increase, which was well described by a semi-mechanistic model including control mechanisms; when G-CSF concentrations increased the proliferation rate increased and the bone maturation time reduced for ANC. Subsequently, ANC in the circulation increased leading to increased elimination of G-CSF. Additionally, a non-specific elimination for G-CSF was quantified. The ANC-dependent elimination contributed to 97% at baseline and 49% at an ANC of 0.1 · 10(9)/L to the total G-CSF elimination. CONCLUSION: The integrated G-CSF-myelosuppression model captured the initial rise in endogenous G-CSF following chemotherapy-induced neutropenia and the return to baseline of G-CSF and ANC. The model supported the self-regulatory properties of the system and may be a useful tool for further characterization of the biological system and in optimization of chemotherapy treatment.

Comparison of epirubicin-iodized oil suspension and emulsion for transcatheter arterial chemoembolization in VX2 tumor.

To compare the antitumor efficacy and safety of transcatheter arterial chemoembolization (TACE) by epirubicin suspension (epirubicin suspension: epirubicin-iodized oil mixture without solution) to that by epirubicin emulsion (epirubicin emulsion: epirubicin-iodized oil mixture with solution), the efficacy of treatment by administration of either an epirubicin suspension or emulsion was examined in an animal model. Changes in plasma epirubicin concentration were compared over 24 h immediately after treatment, and enhanced ultrasonographic and histopathological analysis subsequently conducted 7 days after treatment to determine the growth ratio and proportion of viable tumor cells. The growth ratio and proportion of viable tumor cells were found to be significantly lower in the suspension group than in the emulsion group while the plasma epirubicin concentration was found to be significantly higher in the suspension group than in the emulsion group. These results indicate that administration of an epirubicin suspension is a superior form of TACE compared to that of administration of an epirubicin emulsion.

C-arm CT for assessing initial failure of iodized oil accumulation in chemoembolization of hepatocellular carcinoma.

OBJECTIVE: The purpose of our study was to assess the feasibility of using C-arm CT to detect incomplete accumulation of iodized oil in hepatocellular carcinoma immediately after transarterial chemoembolization (TACE). MATERIALS AND METHODS: This retrospective study included 80 hepatocellular carcinoma lesions in 55 patients (41 men and 14 women; mean age, 69.2 years; mean tumor size, 18.1 mm [range, 5-55 mm]) who underwent TACE with a flat-detector C-arm angiographic system. C-arm CT images were acquired at the end of each session, and unenhanced MDCT images were obtained 7 days later. Two independent observers scored both sets of images, using a predefined detection scale for incomplete iodized oil accumulation. The accuracy for predicting residual lesions was compared using the area under the receiver-operating characteristic curve (A(z)). Contrast-enhanced CT findings obtained 1 month after TACE served as reference standards. RESULTS: Viable lesions were observed in 18 of the 80 study lesions by contrast-enhanced CT. The accuracy of the C-arm CT (A(z) = 0.816) was not significantly different (p = 0.449) from that of the MDCT (A(z) = 0.841). Sensitivity, specificity, and positive and negative predictive values for C-arm CT (80.5%, 74.2%, 47.5%, and 92.9%, respectively) and MDCT (86.1%, 75.0%, 50.0%, and 94.9%, respectively) did not differ significantly. CONCLUSION: C-arm CT is nearly equivalent to MDCT for detecting incomplete iodized oil accumulation after TACE, suggesting that the immediate assessment of iodized oil accumulation with C-arm CT without the need to perform follow-up unenhanced MDCT is likely feasible.

Phase II trial of dexverapamil and epirubicin in patients with non-responsive metastatic breast cancer.

Agents capable of reversing P-glycoprotein-associated multidrug resistance have usually failed to enhance chemotherapy activity in patients with solid tumours. Based on its toxicity profile and experimental potency, dexverapamil, the R-enantiomer of verapamil, is considered to be promising for clinical use as a chemosensitizer. The purpose of this early phase II trial was to evaluate the effects of dexverapamil on epirubicin toxicity, activity and pharmacokinetics in patients with metastatic breast cancer. A two-stage design was applied. Patients first received epirubicin alone at 120 mg m(-2) i.v. over 15 min, repeated every 21 days. Patients with refractory disease continued to receive epirubicin at the same dose and schedule but supplemented with oral dexverapamil 300 mg every 6 h x 13 doses. The Gehan design was applied to the dexverapamil/epirubicin cohort of patients. Thirty-nine patients were entered on study, 25 proceeded to receive epirubicin plus dexverapamil. Dexverapamil did not increase epirubicin toxicity. The dose intensity of epirubicin was similar when used alone or with dexverapamil. In nine intrapatient comparisons, the area under the plasma concentration-time curve (AUC) of epirubicin was significantly reduced by dexverapamil (mean 2968 vs 1901 microg ml[-1] h[-1], P= 0.02). The mean trough plasma levels of dexverapamil and its major metabolite nor-dexverapamil were 1.2 and 1.5 microM respectively. The addition of dexverapamil to epirubicin induced partial responses in 4 of 23 patients evaluable for tumour response (17%, CI 5-39%, s.e.P 0.079). The remissions lasted 3, 8, 11 and 11+ months. These data suggest that the concept of enhancing chemotherapy activity by adding chemosensitizers may function not only in haematological malignancies but also in selected solid tumours. An increase in the AUC and toxicity of cytotoxic agents does not seem to be a prerequisite for chemosensitizers to enhance anti-tumour activity.

Clinical experiences with magnetic drug targeting: a phase I study with 4'-epidoxorubicin in 14 patients with advanced solid tumors.

Anticancer drugs reversibly bound to magnetic fluids (ferrofluids) could be concentrated in locally advanced tumors by magnetic fields that are arranged at the tumor surface outside of the organism. If certain requirements are met, systemic toxicity might be minimized, and local tumor efficacy might be increased. We have conducted a Phase I clinical trial using this approach in patients with advanced and unsuccessfully pretreated cancers or sarcomas. Nine such patients received two treatment courses, 3 patients received one course, and 2 patients received three courses of magnetic drug targeting consisting of the infusion of epirubicin in increasing doses (from 5 to 100 mg/m2) that had been chemically bound to a magnetic fluid and the application of magnetic fields to the tumors for 60-120 min. In 2 of 14 patients, the same dose of epirubicin not bound to a magnetic fluid was administered systemically 3 weeks after drug targeting for intraindividual comparisons. Magnetic drug targeting with epirubicin was well tolerated. In one case, a planned second treatment was withdrawn, because of an episode of chills 130 min after infusion of the magnetic drug. Two patients received a third treatment because of good responses after the first two therapies. Based on magnetic resonance tomographic techniques, pharmacokinetics, and the histological detection of magnetites, it was shown that the ferrofluid could be successfully directed to the tumors in about one-half of the patients. Organ toxicity did not increase with the treatment, but epirubicin-associated toxicity appeared at doses greater than 50 mg/m2. Although treatment with magnetic drug targeting seems safe, improvements are necessary to make it more effective and independent of patient- or disease-related problems. A study design to compare conventional treatments with the new treatment form within one patient seems crucial to eliminate interindividual differences.

Disposition of epirubicin after intraarterial administration in Lipiodol to patients with hepatocellular carcinoma.

Delivering emulsions of anthracycline drugs in Lipiodol, an iodinated poppy-seed oil, via the hepatic artery for the treatment of hepatocellular carcinoma (HCC) has become increasingly popular. However, investigations to determine the extent to which the Lipiodol sequesters the anthracycline in the liver have been limited. Concern has been expressed that such emulsions are not stable and that the anthracycline is, therefore, released rapidly into the circulation. We studied the pharmacokinetics of epirubicin (50 mg m-2) in five patients with nonresectable primary hepatocellular carcinoma after infusion of an epirubicin/Lipiodol emulsion via the hepatic artery. We used a reliable and specific high-performance liquid chromatography assay that allows quantitation of plasma concentrations of epirubicin, epirubicinol, epirubicin glucuronide, and epirubicin aglycone. Although a large interpatient variability in pharmacokinetics was observed, our results were similar to historical data after epirubicin intravenous therapy. Only the results from one patient provided evidence of significant retention of the drug in the liver. It would appear that more stable formulations of epirubicin/Lipiodol are required to increase the efficacy of this form of treatment. We suggest that pharmacokinetic studies should accompany clinical evaluation of emulsions of epirubicin/Lipiodol for the treatment of HCC.

Influence of lipiodolization on a cirrhotic liver.

Lipiodolization--selective regional cancer chemotherapy using Lipiodol plus an anticancer drug (LPD)--can prolong survival time of patients with an unresectable hepatocellular carcinoma (HCC). To enhance understanding of LPD's influence on the cirrhotic liver, we carried out related studies. Forty-three cirrhotic patients with HCC were treated with LPD (epirubicin in a dose of 15-40 mg/m2 and Lipiodol of 0.02-0.25 ml/kg). Seven cirrhotic patients with HCC were subjected to hepatic angiography alone, and these subjects selected randomly served as controls. Among the 43 treated with LPD, 23 belonged to Child's class A, 15 to class B, and 5 to class C. Blood samples were taken before angiography (pre) and at 24 hours after angiography (post) from each patient. Post/pre ratio of the following parameters were compared between patients of the two groups: sGOT, sGPT, and LDH as a marker for hepatocyte injury; t. bilirubin and hepaplastin test (HPT) as hepatocyte function; alkaline phosphatase and gamma-GTP to examine bile duct injury; and serum hyaluronic acid level to determine an endothelial cell functions. Post/pre ratio of serum GOT, GPT, LDH levels, and HPT in patients treated with vs. without LPD were 1.32 +/- 0.59 vs. 0.92 +/- 0.09 (P < 0.001), 1.18 +/- 0.43 vs. 0.88 +/- 0.09 (P < 0.001), 1.11 +/- 0.20 vs. 1.00 +/- 0.07 (P < 0.05), and 0.95 +/- 0.10 vs. 1.09 +/- 0.12 (P < 0.01), respectively. There were no significant differences in post/pre LPD ratio of other parameters, rates of complications, and hospital stay after LPD for patients with Child's class A, B, and C. Hepatocytes are apparently the primary site of injury in cases of LPD. LPDs, using epirubicin in a dose of 15-40 mg/m2 and Lipiodol in a dose of 0.02-0.25 ml/kg, proved to be safe for cirrhotic patients with HCC.

Coupled-column liquid chromatographic analysis of epirubicin and metabolites in biological material and its application to optimization of liver cancer therapy.

A specific, sensitive and fully automated coupled-column LC method for the determination of the anthracycline cytostatic epirubicin and four metabolites in the biological materials human plasma, liver homogenate and liver tumour homogenate has been developed. System-integrated sample processing was achieved using a new restricted access silica precolumn packing. This porous Alkyl-Diol Silica (ADS) was specially designed for the direct and repetitive injection of proteinaceous samples. It consists of a hydrophilic and electroneutral external particle surface (glyceryl-residues) and a hydrophobic reversed-phase internal surface (butyryl-, octanoyl- or octadecyl-residues). These bimodal chromatographic properties allow retention of low molecular analytes by classical RP-chromatography exclusively at the lipophilic pore surface. Macromolecular constituents of the sample matrix (e.g. proteins) are size-excluded by 5 nm pores and quantitatively eliminated in the interstitial void volume. On-line analysis was performed by coupling a C4-Alkyl-Diol precolumn (20 x 4 mm i.d., particle size 25 microns) and LiChrospher RP Select B analytical column (250 x 4 mm i.d., particle size 5 microns) via an electrically driven six-port valve. Separation of the parent compound and its metabolites was achieved with a mobile phase consisting of water (0.1% triethylamine, v/v, pH 2.0 adjusted with trichloroacetic acid)-acetonitrile (70:30, v/v) at a flow rate of 1 ml min-1. The analytes were detected using their natural fluorescence (excitation 445 nm, emission 560 nm). The method described is used for the determination of pharmacokinetics of epirubicin and its metabolites in order to evaluate and optimize treatment regimen of liver cancer chemoembolization therapy.

[Transplacental passage of epirubicin]. / Passage transplacentaire de l'épirubicine.

We studied the transplacental transfer of epirubicin, an anthracycline used for the treatment of different neoplastic disorders, in particular breast cancers, by in vitro perfusion of term human placenta. Placenta from women with uncomplicated pregnancy were collected immediately after vaginal delivery and put into 37 degrees C thermostated hood. Perfusion of foetal surface of the placenta by modified Earle's solution was started immediately after catheterisation at a flow rate of 6 ml/min and then so was the perfusion of the intervillous space at the rate of 12 ml/min. Samples were collected at different times after the initiation of the perfusion from arterial inflow and venous outflow respective of the maternal and foetal compartment. The transplacental transfer of epirubicin was investigated for two doses: 5 and 9 micrograms/ml. The mean transfer value of epirubicin is low (3.66 +/- 1.07%) for the two tested doses and is only slightly higher than doxorubicin transfer, which drug has provided rare accidents in the clinical reports. These results are in favour of a low placental toxicity of epirubicin. Clinical data have to be collected from pregnant women to confirm the low foetal toxicity of epirubicin.

The pharmacokinetics of high-dose epirubicin and of the cardioprotector ADR-529 given together with cyclophosphamide, 5-fluorouracil, and tamoxifen in metastatic breast-cancer patients.

A high-pressure liquid chromatographic method for determination of the bisdioxopiperazine derivative ADR-529 (ICRF-187), a compound proven effective in protection against anthracycline-induced cardiotoxicity, has been developed. The limit of quantitation was 5 ng/ml using a narrow-bore 5-microns silica column and UV detection. The method was used for determination of pharmacokinetic profiles of ADR-529 after a 3-weekly i.v. administration of different doses of ADR-529 (600-1000 mg/m2) together with different doses of epirubicin (E, 60-100 mg/m2), fixed-dose cyclophosphamide (C, 600 mg/m2), fixed-dose 5-fluorouracil (F, 600 mg/m2), and daily administration of tamoxifen (T, 30 mg; CEF-T) in the treatment of patients with metastatic breast cancer. Pharmacokinetic parameters for epirubicin were also determined. The aim of the study was to determine (1) whether the pharmacokinetics of ADR-529 as part of a combination with CEF-T changes with increasing doses of ADR-529 and increasing doses of epirubicin and (2) whether the pharmacokinetics of epirubicin in the same combinations is altered with the administration of increasing doses of ADR-529. A total of 82 patients were included. A crossover study including 16 of the patients showed no significant difference in epirubicin pharmacokinetic parameters when epirubicin was given with or without concomitant administration of ADR-529. Apart from minor changes in the distributional half-lives, the pharmacokinetic parameters of epirubicin were not altered with increasing doses of ADR-529, nor were the pharmacokinetic parameters of ADR-529 itself. Escalating doses of epirubicin did not significantly alter the pharmacokinetic parameters of ADR-529 with the exception of a 30% increase in the terminal half-life and a decrease in total body clearance when the epirubicin dose was raised from 60 to 100 mg/m2. We conclude that concomitant administration of ADR-529 does not alter the distribution and elimination of epirubicin in doses suitable for preventing the anthracycline-induced cardiotoxicity.

Disposition of epirubicin in an oily contrast medium after intravenous and intrahepato-arterial administration in liver cancer: a preliminary report.

The present study reports findings on the disposition of epirubicin after an intrahepato-arterial administration of the Lipiodol-drug complex, prepared by mixing the drug-aqueous phase with the iodized oil by ultra-sonification, in 14 patients with histologically proven hepatoma or hepatomegaly with serum alpha-fetoprotein level above 500 micrograms.l-1. The volume of Lipiodol used was 5 ml and the epirubicin dose was 50 mg.m-2. Blood samples were obtained at various time intervals up to 72 h post-dose. Serum concentrations of epirubicin were measured by liquid chromatography with fluorometric detection. The area under serum concentration-time curve (AUCinfinity0) was higher in the Lipiodol-epirubicin group (n = 8) while the clearance was faster and elimination t1/2 and mean residence time shorter in the plain epirubicin group (n = 3). However, interindividual variation in metabolism of epirubicin would affect serum level of the drug. In three patients who were given intravenous and intrahepato-arterial injections (90 mg.m-2) of plain epirubicin and Lipiodol-drug complex, the relative bioavailability of Lipiodol-epirubicin complex (F = 0.76 and 0.45) was lower than that of plain epirubicin (F = 0.80 and 0.73) in two patients while it was approximately 100% (F = 1.06 and 1.20) in one patient. It is likely that liver function of the patients might be modified by the disease state over a period of 3 months in the cross-over study. Further studies with larger patient samples are required to confirm if there is a targeting effect of the Lipiodol-drug complex toward hepatoma using a better formulation of the drug in Lipiodol.