Viktorija Dragojevic-Simic

Amifostine protection against doxorubicin cardiotoxicity in rats

Aminothiol amifostine (AMI) protects against toxic effects of both ionizing radiation and numerous anticancer drugs. The aim of this study was to investigate the potential protective effects of AMI against doxorubicin (DOX)-induced cardiotoxicity in rats. Male Wistar rats were treated with AMI (75 mg/kg i.p.) and/or DOX (1.25 mg/kg i.p.), 4 times per week, for 4 weeks. Mortality, general condition and body weight of the animals were observed during the whole treatment, and for a further 4 weeks, until the end of experiment. Evaluation of cardioprotective efficacy of AMI was performed by analyzing the electrocardiographic parameters and response to the pro-arrhythmic agent aconitine, as well as activity registration of the in situ rat heart preparations. Necropsy was also performed at the end of the experiment, and heart excision, weight and macroscopic examination were done before histological evaluation. Doxorubicin caused rat heart disturbances manifested by prominent electrocardiographic changes (S&agr;–T prolongation and T-wave flattening), significantly enhanced response to aconitine, decrease of the heart rate and contractility, as well as histopathologically verified myocardial lesions. The heart changes were accompanied by 40% mortality rate, significant decline in body mass and severe effusion intensity score in 66.6% of the animals. Application of AMI before each dose of DOX significantly reduced or completely prevented its toxic effects. Therefore, since AMI had very good protective effects against a high dose of DOX given as a multiple, low, unitary dose regimen, not only on the heart but on the whole rat as well, it could be recommended for further investigation in this potentially new indication for clinical application.

Radioprotective Efficiency of Fullerenol in Irradiated Mice

In vitro studies have demonstrated that fullerenol, a polyhydroxylated derivative of fullerene (C60(OH)n n = 12-26), has a high antioxidative potential. Since any radiation injury is mainly a consequence of the action of free radical species, the aim of this study was to examine radioprotective efficiency of fullerenol in whole-body irradiated mice. The experiment was performed on male, adult, white mice, whole-body irradiated with Xrays doses of 6 to 8 Gy (X-ray energy of 8 MV). Fullerenol C60(OH)24 was given in doses of 10 and 100 mg/kg i.p. 30 minutes before irradiation. The experimental groups consisted of 25-30 animals each. The survival rate and body mass gain of irradiated animals were monitored for 30 days after irradiation. The mean lethal times (LT50) of irradiated mice and mean lethal dose of X-rays were calculated and compared. The results showed that fullerenol C60(OH)24, in a dose of 100 mg/kg i.p., prolonged LT50 of irradiated mice. This effect was especially pronounced in mice irradiated with 7 and 8 Gy of X-rays. It seems that radioprotective efficiency of fullerenol C60(OH)24 is more marked in mice irradiated by higher doses of X-rays.

Consistency between causality assessments obtained with two scales and their agreement with clinical judgments in hepatotoxicity

Reliability and usefulness of scales for causality assessment in hepatotoxicity have not been fully explored. The goal of this study was to examine consistency between causality assessments obtained with two commonly used scales and their agreement with initial clinical assessments in hepatotoxicity reported in Serbia, and to review usefulness of these scales.

Influence of Fullerenol C60(OH)24 on Doxorubicin Induced Cardiotoxicity in Rats

Earlier investigation of fullerenol, C60(OH)24, features, in vitro, showed that fullerenol have strong antioxidative potential. In this work, we examined the influence of fullerenol as a potential antioxidative protector on doxorubicin induced cardiotoxicity in rats. Experiments were performed on adult Wistar rats, both gender. Animals were divided into six groups, each containing eight individuals. Doxorubicin was administrated i.v. (tail vein) in single dose of 8mg/kg. Fullerenol C60(OH)24 in treated animals was administrated i.p. (in doses 50, 100, 200 mg/kg) for 30 min. before application of doxorubicin. Control group (intact animals) was given saline (1 mL/kg). One group was treated only with fullerenol (100 mg/kg i.p.). Cardiotoxicity of doxorubicin as well as cardioprotective effects of fullerenol were evaluated following the heart function monitored by ECG recording during adrenalin i.v. infusion, and pathomorphological examination of the heart tissue. These evaluations were performed on the day 2 and 7 after doxorubicin administration. Both functional and pathomorphological investigations revealed no heart damage two days after given treatments. However, on the day 7 after doxorubicin injection, changes in cardiovascular reflexes to adrenalin as well as structural damage were manifest. The time for appearance of adrenalin-induced reflex bradicardia in ECG record was significantly longer in doxorubicin treated group in comparison with the control one. Also, pathomorphological examination of the heart tissue showed vacuolization of cardiomyocites. In fullerenol pretreated groups these described changes were ameliorated and corresponded to the control values. These results suggest that fullerenol might be potential cardioprotector in doxorubicin treated individuals.

Accuracy and reproducibility of two scales in causality assessment of unexpected hepatotoxicity

What is known and Objective:  There is almost no published information about reliability of scales for causality assessment in hepatotoxicity at pharmacovigilance centres. The aim of this study was to compare two commonly used scales in cases of unexpected hepatotoxicity, in evaluating their accuracy and reproducibility at pharmacovigilance centres (in signal detection).

Interspecies differences in expression of cytokeratin polypeptides within thymic epithelium: A comparative immunohistochemical study

Cytokeratin (CK) polypeptide expression within the thymic epithelium of several mammalian species (mouse, rat, calf, pig, rabbit, and human) has been analyzed by the streptavidin-biotin immunoperoxidase method. Comparative analysis by a large panel of 17 monoclonal antibodies (mAbs) specific for individual CK polypeptides, pairs, or groups showed considerable heterogeneity of thymic epithelial cells (TEC) in each species. In addition, extreme interspecies difference in CK contents was observed. Four main phenotypic zones: the subcapsule/perivascular area, cortex, medulla, and Hassalls corpuscles (HC) were clearly identified, each characterized by different CK expression. Medullary TEC were more heterogenous and shared common CK polypeptides either with subcapsular/perivascular TEC, cortical TEC, or HC, in most species.

Fullerenol nanoparticles prevents doxorubicin-induced acute hepatotoxicity in rats

Doxorubicin (DOX), commonly used antineoplastic agent, affects bone marrow, intestinal tract and heart, but it also has some hepatotoxic effects. Main mechanism of its toxicity is the production of free reactive oxygen species. Polyhidroxilated C60 fullerene derivatives, fullerenol nanoparticles (FNP), act as free radical scavengers in in vitro systems. The aim of the study was to investigate potential FNP protective role against DOX-induced hepatotoxicity in rats. Experiments were performed on adult male Wistar rats. Animals were divided into five groups: (1) 0.9% NaCl (control), (2) 100mg/kg ip FNP, (3) 10mg/kg DOX iv, (4) 50mg/kg ip FNP 30min before 10mg/kg iv DOX, (5) 100mg/kg ip FNP 30min before 10mg/kg iv DOX. A general health condition, body and liver weight, TBARS level and antioxidative enzyme activity, as well as pathohistological examination of the liver tissue were conducted on days 2 and 14 of the study. FNP, applied alone, did not alter any examinated parameters. However, when used as a pretreatment it significantly increased survival rate, body and liver weight, and decreased TBARS level, antioxidative enzyme activity and hepatic damage score in DOX-treated rats. FNP administered at a dose of 100mg/kg significantly attenuated effects of doxorubicin administered in a single high dose in rats, concerning general condition, body and liver weight, lipid peroxidation level and antioxidative enzyme activity as well as structural alterations of the hepatic tissue.

Five-year survival and costs of care in metastatic colorectal cancer: conventional versus monoclonal antibody-based treatment protocols.

Aim: To evaluate the costs and survival estimates of metastatic colorectal carcinoma patients treated with conventional cytostatic protocols and adjuvant monoclonal antibodies (mAbs). Methods: Retrospective randomized case series and cost-of-illness analysis was used. Metastatic colorectal carcinoma cases (62) were randomly selected from the archive of the largest university military hospital in Southeastern Europe. Results: A 6-month longer survival was attributed to mAbs (p = 0.581). Conventional protocols incurred €5137 (95% CI: €3758–€6517) versus €22,113 (95% CI: €16,201–€28,025) total direct medical costs in mAb-based group. ICER of €32,108 per life year gained attributable to mAbs three-fold exceeded informal willingness to pay threshold of Serbia. Conclusion: mAbs adjuvant protocols had modest positive impact on 5-year survival rates. Costs were driven by targeted biologicals, but significantly higher costs of care were recorded in mAb-treated group in other domains, as well. More selective prescription and reimbursement criteria should be applied to increase cost–effectiveness of targeted oncology agents.

[Simultaneous determination of amoxicillin and clavulanic acid in the human plasma by high performance liquid chromatography-mass spectrometry (UPLC/MS)].

BACKGROUND/AIM Quantitative analysis of amoxicillin and clavulanic acid in biological matrices requires sensitive and specific methods which allow determination of therapeutic concentration in 1 g/mL range. Analytical methods for determination of their concentrations in body fluids described in literature include high performance liquid chromatography coupled to UV detector (HPLC-UV) and liquid chromatography-mass spectrometry (LC-MS). The aim of this study was to develop sensitive and specific ultra performance liquid chromatography/mass spectrometry (UPLC/MS) method which could be used for the spectral identification and quantification of the low concentrations of amoxicillin and clavulanic acid in the human plasma. METHOD A sensitive and specific UPLC/MS method for amoxicillin and clavulanic acid determination was developed in this study. The samples were taken from the adult healthy volunteers receiving per os one tablet of amoxicillin (875 mg) in combination with clavulanic acid (125 mg). RESULTS Plasma samples were pretreated by direct deproteinization with perchloric acid. Quantification limit of 0.01 microg/ml for both amoxicillin and clavulanic acid was achieved. The method was reproducible day by day (RSD < 7%). Analytical recoveries for amoxicillin ranged from 98.82% to 100.9% (for concentrations of 1, 5 and 20 microg/mL), and recoveries for clavulanic acid were 99.89% to 100.1% (for concentrations of 1, 2 and 5 microg/mL). This assay was successfully applied to a pilot pharmacokinetic study in healthy volunteers after a single-oral administration of amoxicillin/clavulanic combination. The determined plasma concentrations of both amoxicillin and clavulanic acid were in the range of the expected values upon the literature data for HPLC-UV and LC-MS methods. CONCLUSION The described method provided a few advantages comparing with LC/MS-MS method. The method is faster using running time of 5 minute, has lower limit of quantification (LOQ) and it could be used in pharmacokinetic studies of both amoxicillin and clavulanic acid.

Impact of clinical pharmacology on health care: Serbian experience

Dear Sirs, During the recent 7th Congress of the European Association for Clinical Pharmacology and Therapeutics, held in Poznan, Poland, serious concerns were expressed by leading invited speakers on the future of clinical pharmacology as a discipline [1]. Their joint conclusion was that clinical pharmacologists in the majority of developed European countries are less and less involved in health care [2]. ‘‘It is the retreat from the bedside into the laboratory that is responsible’’ for the discipline’s decline, one of the speakers said ominously [3]. However, this trend is not taking place in all European countries. In some Eastern European countries, clinical pharmacology is still very much a patientbased discipline, with a significant impact on health care. It is that impact that keeps the discipline alive and prosperous, as can be seen from the Serbian experience. The status and impact of clinical pharmacology on health care in Serbia was investigated in two ways: by field survey and by questionnaire. In the field survey, 38 hospitals and health centers (two tertiary health care hospitals, three mixed tertiary/secondary health care hospitals, and 33 health centers, each consisting of a secondary care hospital plus a primary health care facility) were visited twice during the years 2002 to 2005, and the anonymous questionnaire was sent to 78 available clinical pharmacologists (out of 90 registered) from November 2004 to January 2005. While the field survey confirmed that the tertiary care and the mixed tertiary/secondary care hospitals had three to four clinical pharmacologists each, and four health centers had one clinical pharmacologist each, the questionnaire showed that 50% of the respondents (60 clinical pharmacologists) worked in the health care system, half of them as full-time employees. Both the field survey and the questionnaire indicated that the following services were given by clinical pharmacologists in the Serbian health care system: drug/patient problem consultations, drug information, chairing of drug and therapeutics committees, intensive monitoring of adverse drug effects, therapeutic drug monitoring, formulation of hospital drug policies, auditing of prescribing practice, design of postmarketing studies, continuing medical education, predispensing control of prescribing, and drug utilization analyses. The scope of health care services given by clinical pharmacologists was the largest in the two tertiary care hospitals with independent departments of clinical pharmacology. There is a growing trend in the number of services given, as can be seen from the data from the Center for Clinical and Experimental Pharmacology, Clinical Center ‘‘Kragujevac,’’ for a 10-year period (Fig. 1). Due to good organization of work within the departments, the drug management process in both hospitals was advanced (active drug and therapeutics committees, more than 60 local guidelines adopted), and the availability of all necessary drugs was achieved within the limits of modest drug budgets. Both hospitals had drug formularies completely agreed upon by other clinicians that yet were not far from the World Health Organization’s essential drugs model list (for instance, Clinical Center ‘‘Kragujevac’’ had 338 different drug entities and 584 formulations in total, with 6.08 euros spent for drugs per one bed-day in 2004; the available budget for 2004 was 5.94 euros per one bed-day). With predispensing control of prescribing (in Clinical Center ‘‘Kragujevac,’’ errors were found in about 12% of prescriptions, and drug dispensing was corrected appropriately), continuing medical education, and auditing of prescribing practice, clinical pharmacologists in these S. M. Jankovic (&) AE D. R. Milovanovic University of Kragujevac and Center for Clinical & Experimental Pharmacology, Clinical Center ‘‘Kragujevac’’, Kragujevac, Serbia & Montenegro