Mario Eandi

Determination of Ibuprofen in human plasma by high-performance liquid chromatography: validation and application in pharmacokinetic study.

A specific method for the simultaneous determination of S-(+)Ibuprofen and R-(-)Ibuprofen enantiomers in human plasma is described. Adopting a high-performance liquid chromatographic (HPLC) system with spectrofluorometer detector, the compounds were extracted from plasma in alcohol medium and were separated on C18 column, using a solution of acetonitrile-water-acetic acid-triethylamine as mobile phase. The limit of quantitation was 0.1 microg/mL for both compounds. The method was validated by intra-day assays at three concentration levels and was used in a kinetic study in healthy volunteers. During the study we carried out inter-day assays to confirm the feasibility of the method.

Expression of CYP3A isoforms and P-glycoprotein in human stomach, jejunum and ileum.

1 CYP3A isoforms metabolise a diverse array of clinically important drugs and P‐glycoprotein (P‐gp), a transmembrane efflux pump, can extrude a wide variety of drugs from the cell. It has been suggested that the function of CYP3A4 is complementary to that of P‐gp along the gastrointestinal (GI) tract, together forming a coordinated intestinal barrier against xenobiotics. Therefore, the expression of CYP3A4, CYP3A5, CYP3A7 and ABCB1 (P‐gp) genes were quantified in five normal samples from the human stomach, seven from the jejunum and eight from the ileum by real‐time reverse transcription–polymerase chain reaction and western blot analysis. 2 In the tissues examined, measurable mRNA expression of CYP3A was found in almost all samples from the stomach, jejunum and ileum. The rank order for CYP3A mRNA expression was CYP3A4 > CYP3A5 > CYP3A7 in the GI tract studied, whereas median mRNA CYP3A4 expression was highest in the small intestine and lowest in the stomach. Expression of ABCB1 mRNA was found in almost all samples and the median mRNA expression level was comparable in the jejunum and ileum, but lower in the stomach. Our data also show a significant correlation between all mRNA transcripts studied and a wide interindividual variation. 3 At the protein level, CYP3A4 was detected in all stomach and small intestine samples, the levels being substantially higher in the small intestine than in the stomach. P‐Glycoprotein was detected in all GI samples, but no statistically significant difference was found along the GI tract considered. 4 Collectively, these results demonstrate that CYP3A4 is the main CYP3A expressed in the GI tract investigated, an extensive interindividual variability in the expression of the different CYP3A isoforms in all tissues examined and P‐gp apoprotein levels similar in the stomach, jejunum and ileum.

Static and ELF magnetic fields enhance the in vivo anti-tumor efficacy of cis-platin against lewis lung carcinoma, but not of cyclophosphamide against B16 melanotic melanoma

Previous works showed that exposure to static and extremely low frequency (ELF) magnetic fields (MF) over 3 mT slows down the growth kinetics of human tumors engrafted s.c. in immunodeficient mice, reducing their metastatizing power and prolonging mouse survival. In the experiments reported here, immunocompetent mice bearing murine Lewis Lung carcinomas (LLCs) or B16 melanotic melanomas were exposed to MF and treated respectively with two commonly used anti-cancer drugs: cis-diamminedichloroplatinum (cis-platin) and N,N-bis (2-chloroethyl)tetra-hydro-2H-1,3,2-oxazaphosphorin-2-amine 2-oxide (cyclophosphamide). The experiment endpoint was survival time. The survival time of mice treated with cis-platin (3mg/kg i.p.) and exposed to MF was significantly (P<0.01) longer than that of mice treated only with cis-platin or only exposed to MF, superimposing that of mice treated with 10mg/kg i.p. of the drug, showing that MF act synergically with the pharmacological treatment. On the contrary, when mice treated with cyclophosphamide (50mg/kg i.p.) were exposed to MF no synergic effects were observed, the survival curve being exactly the same as that of mice treated with the drug alone. No clinical signs or toxicity were seen in any of the mice exposed to MF alone or along with cis-platin or cyclophosphamide treatment, compared to mice given only the two known drugs.A possible explanation for the synergic effect of MF being found in mice treated with cis-platin could be that the platinum ion stimulates radical production and that MF enhance active oxygen production bringing about changes in tumor cell membrane permeability, influencing positively the drug uptake. Alternatively, or in addition to this, it has been demonstrated that the rate of conversion of cis-platin to reactive species able to bind to DNA, is increased by localized production of free radicals by MF.

Thiopurine S-methyltransferase pharmacogenetics in a large-scale healthy Italian–Caucasian population: differences in enzyme activity

AIMS To investigate the influence of genotype, age and gender on the thiopurine S-methyltransferase (TPMT) phenotype in healthy Italian-Caucasian subjects. MATERIALS & METHODS The study investigated the TPMT genotype and the TPMT phenotype of 943 healthy Italian-Caucasian subjects of different age and gender (age range: 0.08-68 years; 623 males 320 females). TPMT red blood cell activity was measured in all samples and genotype was determined for the TPMT alleles *2, *3A, *3B and *3C. RESULTS TPMT activity levels in our whole population ranged from 1.6 up to 75.2 U/gHb. Significant TPMT activity differences between wild-type and heterozygous subjects were observed. We divided our TPMT activity into four categories according to our frequency distribution: low (0.1%), intermediate (32.9%), normal (60%) and high (7%), with arbitrary cut-off values of 8.0, 19.4 and 37.0 U/gHb, respectively. The whole population had a total of 94.5% of homozygous wild-type subjects, 5.4% heterozygous variants and one (0.1%) compound heterozygous variant TPMT*3B/*3C. The overall concordance rate between TPMT genotypes and phenotypes was 71.6%. The TPMT activity was significantly higher in wild-type children (0.08-17 years) than in wild-type adults (aged 18-68 years). Moreover, it was noted that wild-type infants from 0.08 to 5 years had a 9% higher average TPMT activity than the other wild-type groups, and only in children from 0.08 to 2 years was the TPMT activity higher in males than in females. CONCLUSION The data obtained in this study show that genetic factors seem to be the major aspect in TPMT phenotype variability in adults, whilst, in children, other physiological factors should be taken into consideration when assessing the TPMT phenotype, such as age and gender.

Preliminary pharmacokinetic study of ibuprofen enantiomers after administration of a new oral formulation (ibuprofen arginine) to healthy male volunteers

The pharmacokinetics of ibuprofen enantiomers were investigated in a crossover study in which seven healthy male volunteers received single oral doses of 800 mg racemic ibuprofen as a soluble granular formulation (sachet) containing L-arginine (designated trade name: Spedifen), 400 mg (-)R-ibuprofen arginine or 400 mg (+)S-ibuprofen arginine. Plasma levels of both enantiomers were monitored up to 480 minutes after drug intake using an enantioselective analytical method (HPLC with ultraviolet detection) with a quantitation limit of 0.25 mg/l. Substantial inter-subject variability in the evaluated pharmacokinetic parameters was observed in the present study. After (+)S-ibuprofen arginine, the following mean pharmacokinetic parameters +/-SD were calculated for (+)S-ibuprofen: tmax 28.6 +/- 28.4 min; Cmax 36.2 +/- 7.7 mg/l; AUC 86.4 +/- 14.9 mg.h/l; t1/2 105.2 +/- 20.4 min. After (-)R-ibuprofen arginine, the following mean pharmacokinetic parameters were calculated for (+)S-ibuprofen and (-)R-ibuprofen, respectively: tmax 90.0 +/- 17.3 and 50.5 +/- 20.5 min; Cmax 9.7 +/- 3.0 and 35.3 +/- 5.0 mg/l; AUC 47.0 +/- 17.2 and 104.7 +/- 27.7 mg.h/l; t1/2 148.1 +/- 63.6 and 97.7 +/- 23.3 min. After racemic ibuprofen arginine, the following mean pharmacokinetic parameters were calculated for (+)S- and (-)R-ibuprofen, respectively: tmax 30.7 +/- 29.1 and 22.9 +/- 29.8 min; Cmax 29.9 +/- 5.6 and 25.6 +/- 4.4 mg/l; AUC 105.1 +/- 23.0 and 65.3 +/- 15.0 mg.h/l; t1/2 136.6 +/- 20.7 and 128.6 +/- 45.0 min. Tmax values of S(+)- and (-)R-ibuprofen after a single dose of 400 mg of each enantiomer did not differ significantly from the corresponding parameters obtained after a single dose of 800 mg of racemic ibuprofen arginine, indicating that the absorption rate of (-)R- and (+)S-ibuprofen is not different when the two enantiomers are administered alone or as a racemic compound. An average of 49.3 +/- 9.0% of a dose of the (-)R-ibuprofen arginine was bioinverted into its antipode during the study period (480 minutes post-dosing). The percent bioinversion during the first 30 minutes after (-)R-ibuprofen arginine intake averaged 8.1 +/- 3.9%. The mean AUC of (+)S-ibuprofen calculated after 800 mg racemic ibuprofen arginine (105.1 +/- 23.0 mg.h/l) was lower than the mean AUC value obtained by summing the AUCs of (+)S-ibuprofen after administration of 400 mg (+)S-ibuprofen arginine and 400 mg (-)R-ibuprofen arginine (133.4 +/- 26.6 mg.h/l). In conclusion, the administration of Spedifen resulted in very rapid absorption of the (+)S-isomer (eutomer) with tmax values much lower than those observed for this isomer when conventional oral solid formulations such as capsules or tablets of racemic ibuprofen are administered. This characteristic is particularly favourable in those conditions in which a very rapid analgesic effect is required.

Modifications of [3H]imipramine binding sites in platelets of chronic pain patients treated with mianserin

&NA; Tritiated imipramine binding to whole platelets was measured in 16 chronic pain patients who were free from major depression, and in a control group. The maximum binding was significantly lower in chronic pain patients than in the control group, whereas the binding affinity was not significantly different. Twelve patients were treated with mianserin for 21 days; this produced a significant improvement in the mean scores for pain (evaluated with the McGill Questionnaire) and depressive symptoms (assessed with the Zung Self‐Rating Scale). The improvement in both types of symptom was accompanied by a significant mean increase in the density of the [3H]imipramine binding sites without modifications in the values of the constant of affinity. All the patients who responded well to treatment (N = 8) had a family history of depressive spectrum disorders (DSD), while none of those who failed to respond had a first degree relative with DSD.

Economic evaluation of cinacalcet in the treatment of secondary hyperparathyroidism in Italy.

Background: Imbalanced levels of parathyroid hormone (PTH), serumcalcium (Ca) and phosphorous (P) are associated with an increased risk of cardiovascular (CV) death and fracture in dialysis patients with secondary hyperparathyroidism (SHPT). The calcimimetic agent cinacalcet can attenuate the mineral and hormonal imbalances characteristic of SHPT and may improve outcomes in such patients. Here we describe a cost-utility analysis of cinacalcet for SHPT in dialysis patients in Italy.Methods: We developed a probabilistic Markov model to simulate the effect of cinacalcet on Ca, P and PTH levels in dialysis patients with SHPT, based on data from a European, multicentre, open-label study. The model then correlated these levels with mortality and morbidity (CV events, fractures and parathyroidectomies) using data from the literature, and incorporated Italian data for dialysis, drugs and management of events according to the national cost structure. The simulation horizon was patient lifetime; simulated treatment alternatives were standard treatment (mainly vitamin D sterols and phosphate binders) and cinacalcet + standard treatment.A 3.5% discount rate was applied to life expectancy (LE), quality-adjusted life-expectancy (QALE), costs and times below the upper ranges (time in range [TiR]) recommended by the National Kidney Foundation–Kidney Disease Outcomes Quality initiative for PTH, Ca, P and Ca × P. Utilities were derived from the published literature and took into account dialysis and the impairment of quality of life due to the occurrence of CV events and fractures. Costs were evaluated in year 2009 values from the perspective of the Italian National Healthcare System.Results: Baseline results were calculated with 10 000 iterations. Compared with standard treatment alone, addition of cinacalcet was associated with a mean (SD) increase in TiR of 5.26 (6.59), 3.63 (6.87), 1.70 (6.66) and 2.68 (5.55) discounted patient-years for PTH, Ca and P, respectively, and combined PTH, Ca, P and Ca × P. Cinacalcet increased LE by 1.20 (3.75) life-years (LYs) and QALE by 0.89 (2.59) QALYs. When including the cost for dialysis, the incremental cost-effectiveness ratio (ICER) was €50 012 per LY and €67 361 per QALY, while, if dialysis costs were not included, the ICER was €23 473 per LY and €31 616 per QALY.Conclusions: The results suggest that cinacalcet treatment could be considered cost effective for treatment of SHPT in the Italian healthcare setting, but further investigations are needed to confirm these findings.

Effectiveness and cost-effectiveness of supplemental glutamine dipeptide in total parenteral nutrition therapy for critically ill patients: a discrete event simulation model based on Italian data.

INTRODUCTION The supplementation of alanyl-glutamine dipeptide in critically ill patients necessitating total parenteral nutrition (TPN) improves clinical outcomes, reducing mortality, infection rate, and shortening intensive care unit (ICU) hospital lengths of stay (LOSs), as compared to standard TPN regimens. METHODS A Discrete Event Simulation model that incorporates outcomes rates from 200 Italian ICUs for over 60,000 patients, alanyl-glutamine dipeptide efficacy data synthesized by means of a Bayesian random effects meta-analysis, and national cost data has been developed to evaluate the alternatives from the cost perspective of the hospital. Simulated clinical outcomes are death and infection rates in ICU, death rate in general ward, and hospital LOSs. Sensitivity analyses are performed by varying all uncertain parameter values in a plausible range. RESULTS The internal validation process confirmed the accuracy of the model in replicating observed clinical data. Alanyl-glutamine dipeptide on average results more effective and less costly than standard TPN: reduced mortality rate (24.6% ± 1.6% vs. 34.5% ± 2.1%), infection rate (13.8% ± 2.9% vs. 18.8% ± 3.9%), and hospital LOS (24.9 ± 0.3 vs. 26.0 ± 0.3 days) come at a lower total cost per patient (23,409 ± 3,345 vs. 24,161 ± 3,523 Euro).Treatment cost is completely offset by savings on ICU and antibiotic costs. Sensitivity analyses confirmed the robustness of these results. CONCLUSIONS Alanyl-glutamine dipeptide is expected to improve clinical outcomes and to do so with a concurrent saving for the Italian hospital.

Cost-effectiveness of omega-3 fatty acid supplements in parenteral nutrition therapy in hospitals: a discrete event simulation model.

BACKGROUND & AIMS A recent meta-analysis showed that supplementation of omega-3 fatty acids in parenteral nutrition (PN) regimens is associated with a statistically and clinically significant reduction in infection rate, and length of hospital stay (LOS) in medical and surgical patients admitted to the ICU and in surgical patients not admitted to the ICU. The objective of this present study was to evaluate the cost-effectiveness of the addition of omega-3 fatty acids to standard PN regimens in four European countries (Italy, France, Germany and the UK) from the healthcare provider perspective. METHODS Using a discrete event simulation scheme, a patient-level simulation model was developed, based on outcomes from the Italian ICU patient population and published literature. Comparative efficacy data for PN regimens containing omega-3 fatty acids versus standard PN regimens was taken from the meta-analysis of published randomised clinical trials (n = 23 studies with a total of 1502 patients), and hospital LOS reduction was further processed in order to split the reduction in ICU stay from that in-ward stays for patients admitted to the ICU. Country-specific cost data was obtained for Italian, French, German and UK healthcare systems. Clinical outcomes included in the model were death rates, nosocomial infection rates, and ICU/hospital LOS. Probabilistic and deterministic sensitivity analyses were undertaken to test the reliability of results. RESULTS PN regimens containing omega-3 fatty acids were more effective on average than standard PN both in ICU and in non-ICU patients in the four countries considered, reducing infection rates and overall LOS, and resulting in a lower total cost per patient. Overall costs for patients receiving PN regimens containing omega-3 fatty acids were between €14 144 to €19 825 per ICU patient and €5484 to €14 232 per non-ICU patient, translating into savings of between €3972 and €4897 per ICU patient and savings of between €561 and €1762 per non-ICU patient. Treatment costs were completely offset by the reduction in hospital stay costs and antibiotic costs. Sensitivity analyses confirmed the robustness of these findings. CONCLUSIONS These results suggest that the supplementation of PN regimens with omega-3 fatty acids would be cost effective in Italian, French, German and UK hospitals.

Blood kinetics of sulfamonomethoxine and oxytetracycline following intrauterine spray injection in dairy cows.

Intrauterine administration was performed in six Friesan cows with a disposable spray preparation containing 3 g of sulfamonomethoxine and 3 g of oxytetracycline, in order to investigate their serum kinetics. Sulfamonomethoxine levels were determined by a reversed-phase HPLC method, whilst oxytetracycline quantities were detected by a microbiological method. The sulphonamide had a peak 1.17 h after the administration, the tetracycline reached its highest concentration after 8 h. The bioavailability of both drugs was low and detectable drug amounts were no longer recovered after 24 h.