A phase-III prevention trial of low-dose tamoxifen in postmenopausal hormone replacement therapy users: the HOT study.

BACKGROUND: Postmenopausal hormone replacement therapy (HRT) relieves menopausal symptoms and may decrease mortality in recently postmenopausal women, but increases breast cancer risk. Low-dose tamoxifen has shown retained activity in phase-II studies. METHODS: We conducted a phase-III trial in 1884 recently postmenopausal women on HRT who were randomly assigned to either tamoxifen, 5 mg/day, or placebo for 5 years. The primary end point was breast cancer incidence. RESULTS: After 6.2 ± 1.9 years mean follow-up, there were 24 breast cancers on placebo and 19 on tamoxifen (risk ratio, RR, 0.80; 95% CI 0.44-1.46). Tamoxifen showed favorable trends in luminal-A tumors (RR, 0.32; 95% CI 0.12-0.86), in HRT users <5 years (RR, 0.35; 95% CI 0.15-0.82) and in women completing at least 12 months of treatment (RR, 0.49; 95% CI 0.23-1.02). Serious adverse events did not differ between placebo and tamoxifen, including, respectively, coronary heart syndrome (6 versus 4), cerebrovascular events (2 versus 5), VTE (2 versus 5) and uterine cancers (3 versus 1). Vasomotor symptoms were 50% more frequent on tamoxifen. CONCLUSIONS: The addition of low-dose tamoxifen to HRT did not significantly reduce breast cancer risk and increased climacteric symptoms in recently postmenopausal women. However, we noted beneficial trends in some subgroups which may deserve a larger study.

Does Alpha-lipoic acid improve effects on polycystic ovary syndrome?

Alpha-lipoic acid (ALA) plays a key role in many physiological processes, exerting anti-inflammatory, immunomodulatory, antioxidant, detoxifying, and insulin sensitizing activities. Since ALA improves insulin resistance (IR), it has been suggested that ALA could be beneficial it in the treatment of PCOS. The natural polyol myo-Inositol (myo-Ins) and its isomers (D-Chiro-Inositol, D-Chiro-Ins) has proven to improve PCOS features and clinical outcome, according to a compelling body of available studies. Few studies have proposed to strengthen the inositol effect by associating ALA. ALA does not seem to influence significantly reproductive hormones, while its beneficial effects are presumably restricted to the metabolic features of insulin resistant PCOS women. Therefore, ALA usefulness in improving inositol activity still awaits convincingly confirmation.  Experimental studies as well as proper randomized clinical trials, should be specifically tailored to assess this hypothesis. In absence of reliable evidence, ALA should not be recommended in the routinary clinical management of PCOS, even if associated to myo-Ins.

Pharmacokinetics of thymoxamine in rabbits after ophthalmic and intravenous administration.

The pharmacokinetics of thymoxamine hydrochloride were studied in rabbits by the assessment of its ocular and systemic absorption after instillation of thymoxamine hydrochloride 0.5% eye drops. Plasma levels were compared with those observed after i.v. bolus administration of thymoxamine hydrochloride at 2.5 mg/kg. Deacetylthymoxamine is the main metabolite of thymoxamine, generated by esterase hydrolysis. It was evaluated, as an indication of the parent drug, in aqueous humor and plasma by an HPLC method with fluorescence detection (detection limit = 5 ng/ml). Thymoxamine was found to permeate the cornea and to be hydrolysed very quickly, showing very good absorption with a maximum aqueous humor concentration of deacetylthymoxamine of 2329 ng/ml 15 min after eye drop instillation. The study of the systemic absorption of thymoxamine allowed the exclusion of the possibility of systemic side effects following ocular treatment. In fact, considering the detection limit of the method, the plasma levels of deacetylthymoxamine are certainly more than 100-times lower than those observed with intravenous treatment.

[What model for pharmacokinetic analysis of tissue distribution and bioequivalence?]. / Quale modello nell'analisi farmacocinetica di studi di biodisponibilità e bioequivalenza?

Pharmacokinetic analysis of experimental data is usually carried out through two different approaches: non compartmental model or standard compartmental models. Benefits and restrictions of these two approaches are here discussed, with attention to the studies of clinical pharmacokinetics and, particularly, to those of bioavailability and bioequivalence. In fact, different opinions about the most appropriate model to be used with these studies were encountered. Besides, the paper deals with European and American guidelines, data from a check of the literature published during 1995 on two international journals of clinical pharmacology and results of non-compartmental and non-linear fitting analyses on plasma concentrations obtained with an oral administration of a delayed dosage form in humans.

Acceptable and unacceptable procedures in bioavailability and bioequivalence trials.

Both the US FDA and EU guidelines recommend fundamental procedures for bioavailability and bioequivalence studies. The use of unacceptable procedures in conducting these studies can in fact complicate drug development at NDA or ANDA levels. The most common unacceptable procedures are as follows: the use of compartmental analysis in calculating Cmax, tmax and AUC, which must be evaluated with the non-compartmental approach in pivotal studies; the need to use only homogeneous concentrations in pharmacokinetic analysis, and to avoid using the sum of various individual concentrations, e.g. parent drug plus metabolite(s); the need to consider plasma clearance in evaluating absolute bioavailability with dose-dependent kinetics; the need to use the simultaneous fitting procedure when drugs and metabolite(s) are considered; the unacceptable procedure of either disregarding some experimental data or adding simulated data in pharmacokinetic and statistical analysis; and the use of the additive model rather than the multiplicative procedure in assessing bioequivalence with Cmax and AUC. This paper describes in detail acceptable and unacceptable procedures in bioavailability and bioequivalence studies, covering operating guidelines and principles of pharmacokinetics. (c) 1998 The Italian Pharmacological Society.

Application of high-performance liquid chromatography to the analysis of propionyl-L-carnitine by a stereospecific enzyme assay.

Propionyl-L-carnitine was converted by chemical hydrolysis (0.3 M potassium hydroxide, pH 12.8, room temperature) into L-carnitine, which competes with crotonoylbetaine formation and can be quantitatively evaluated by an enzyme assay. Under the conditions selected, hydrolysis of propionyl-L-carnitine to L-carnitine was completed in a few minutes with less than 0.5% of crotonoylbetaine being formed. The method described is enantioselective and possesses the analytical requirements for assaying propionyl-L-carnitine during chemical synthesis procedures and in pharmaceutical formulations.

High-performance liquid chromatographic evaluation of salicyloyl pyridixol and systemic metabolites in biological samples.

An high-performance liquid chromatographic analytical method, which allows quantitative evaluation of both salicyloyl pyridixol and its metabolite salicylic acid, is reported. This method has demonstrated to possess the required specifications in terms of linearity, sensitivity, extraction recovery, reproducibility and specificity for pharmacokinetic investigations in both human subjects and experimental animals. The results obtained from an investigation on the rat are briefly discussed.

Experimental, extrapolated and truncated areas under the concentration-time curve in bioequivalence trials.

OBJECTIVE: The extrapolated area under the concentration-time curve (AUC0-infinity) for any drug is considered by operating guidelines as the primary parameter related to the extent of absorption in single-dose bioavailability and bioequivalence trials. Not more than 20% should be added to the experimental AUC (AUC0-t) in the extrapolating procedure. However, in certain specific cases, it is problematic and, in other cases, impossible to respect the above requirement. It was intended to demonstrate that truncated AUC or AUC0-t would be used in bioequivalence trials when the AUC cannot be extrapolated. METHODS: AUC0-t and truncated AUC were compared at various time intervals with AUC0-infinity in a series of 19 single-dose bioequivalence trials covering 15 different drugs, each carried out on 12-24 healthy volunteers. Point estimators and 90% confidence intervals in the 0.80-1.25 and 0.70-1.43 ranges were statistically processed using log-transformed parameters. RESULTS: In all the trials considered, overlapping point estimators and 90% confidence intervals were invariably obtained, regardless of the AUC used. CONCLUSION: The use of AUC0-t or truncated AUC may be considered an ancillary procedure in bioequivalence trials when AUC cannot be extrapolated. This occurs with most extended-release formulations, endogenous substances, and poorly absorbed drugs. It also occurs in trials with one or more poor metabolisers, with drugs possessing very long elimination half-lives, and with bioassays having problems of sensitivity that preclude sufficiently precise plasma concentration measurement over the required sampling period.

Activity and pharmacokinetics of a new oral dosage form of soluble ibuprofen.

The pharmacokinetics and the relative bioavailability of a soluble granular form (sachets) of a pharmaceutical formulation containing ibuprofen (CAS 15687-27-1) and 1-arginine were investigated in healthy volunteers. Two granular dosage forms were evaluated, 200 and 400 mg, in comparison with the commercial equivalents (tablets). After the oral administration of both granular dosage forms, a quicker absorption and a significantly higher plasma bioavailability of ibuprofen in the first hour following the treatment than after tablets administration were observed. The mean values of peak plasma concentration (microgram/ml) were 26.1 and 56.4 after treatment with 200 and 400 mg sachets respectively vs. 16.3 and 43.0 after treatment with 200 and 400 mg tablets. The mean values of peak time were 16.9 and 24.4 min after treatment with 200 and 400 mg sachets respectively vs. 90.0 and 63.7 min after treatment with 200 and 400 mg tablets. The shortening in the absorption time and the increase in the plasma concentrations did not involve a quicker drug elimination nor cause any changes in the bioavailability (mean values of the relative bioavailability indexes of 0.98 for 200 mg dosage form and 1.02 for the 400 mg one). The analgesic activity of soluble ibuprofen 400 mg was compared with that of ibuprofen 400 mg tablets in patients with osteo-articular pain, according to a single dose, double-blind cross-over balanced design. The results showed that the soluble granular form is able to determine an analgesic effect significantly quicker and higher than that of tablets.

High-performance liquid chromatographic evaluation of PCF 39, a new immunomodulator agent.

An high-performance liquid chromatographic analysis of PCF 39, N2-[5-(hypoxanthin-9-yl)pentyloxycarbonyl]-L-arginine, with ultraviolet detection, has been devised and validated. The main pharmacokinetic results encountered for rats treated intravenously with PCF 39 at a dose of 100 mg/kg are described.

Evaluation of ibuprofen dimethyl aminoethanol octyl bromide and related active metabolites in biological samples.

AF 150 is ibuprofen dimethyl aminoethanol octyl bromide, endowed with antibacterial and antiinflammatory activities. An indirect HPLC analytical method for AF 150 with UV detection, to be used for pharmacokinetic studies, has been validated in terms of linearity, reproducibility, specificity and sensitivity. Using this method the bioavailability of AF 150 and its active metabolite ibuprofen was investigated following parenteral administration and topical application.

Chromatographic and non-chromatographic assay of L-carnitine family components.

L-Carnitine and its acyl esters constitute an endogenous pool of the L-carnitine family, involved in the uptake of free fatty acids in the mitochondria by transfer across their membrane of the acyl moieties to fuel the beta-oxidation and the release of the acetyl group from the mitochondria to the cytosol. Therefore acyl-L-carnitine and acyl-L-carnitine transferase are involved in a homeostatic equilibrium with the cells. As most of these substances need to be monitored in foods, chemical and pharmaceutical processes and biological fluids, an overview of the main methods for assaying them is provided here, with specific reference to the intrinsic performance of each analytical procedure and with suggestions on the correct storage and manipulation of analytical samples.

Comparative bioavailability study on naproxen betainate sodium salt monohydrate and naproxen sodium salt in healthy volunteers.

The S-naproxen betainate sodium salt monohydrate (naproxen-betaNa, CAS 104124-26-7, Aprenin, test drug), and the sodium salt of S-naproxen (reference), were administered to twelve healthy volunteers of both sexes according to a crossover design, in a single dose of one 575 mg capsule of test, containing 342 mg of S-naproxen and two 275 mg tablets of reference, containing 502 mg of S-naproxen. Blood samples were drawn off over a 24-h period before (time 0) and after administration at foreseen time intervals. Naproxen was measured in plasma by a validated HPLC assay with UV detection which was able to detect 1 microgram/ml and proved to be linear in the range 1-100 micrograms/ml. The non-compartmental pharmacokinetic parameters obtained were statistically processed according to the EU guidance note on bioavailability and bioequivalence Cmax, AUC0-24h and AUC0-infinity were normalized to the dose of 502 mg of naproxen and log-transformed before statistical analysis to assess bioequivalence. Dose-normalized values of plasma concentrations encountered with the two formulations proved to overlap, with the exception of the first sampling time which showed naproxen concentrations that were higher with test drug than with reference. The specific test for bioequivalence led to 90% confidence intervals within the 80-125% range with target pharmacokinetic parameters, whereas the time to peak (tmax) observed with the test and reference drugs did not differ to any statistically significant degree when analysed with Wilcoxon's non-parametric test. It is concluded that the test drug should be declared bioequivalent with the reference drug in terms of dose-normalized concentrations, despite the more rapid increase in plasma concentrations of naproxen observed at the first sampling time with test drug.

Amlodipine bioequivalence achieved with a very sensitive liquid chromatography tandem mass spectrometric bioassay.

Amlodipine (CAS 88150-42-9) is a 1,4-dihydropyridine derivative, one of the most widely used drugs for the management of essential hypertension. In developing manidipine (CAS 120092-68-4), a new antihypertensive drug, amlodipine was selected as the reference comparator drug in a Phase III double blind clinical trial. However, manidipine is formulated in hard gelatin capsules, whereas amlodipine is presented as a tablet. In order to respect the double blind design of the study, it was necessary to insert the amlodipine tablet into hard gelatin capsules matching those of the new test product. This called for an amlodipine bioequivalence study on two halves of one tablet inserted into a capsule (test formulation) and two halves of one tablet ingested as such (reference formulation). The bioequivalence trial was carried out on 18 healthy volunteers (9 males and 9 females). Subjects were administered a single 10 mg dose of test and reference products according to a two-treatment, two-period, two-sequence crossover design, with a wash-out period of three weeks. Plasma concentrations of the parent compound were monitored over a period of 6 days, considering the long half-life of amlodipine. The drug was quantified with a very sensitive, robust bioassay, which was set up and validated in our laboratory. Peak concentration and area under the curve of plasma concentrations were log-transformed and analyzed to obtain 90% confidence intervals which proved to be 0.94-1.06, and thus within the acceptable bioequivalence range of 0.80-1.25. Time to peak, analyzed according to a non-parametric test, did not show any statistically significant difference between the test and reference. Both the test and reference products showed a similar and very good safety profile. The conclusion is that one amlodipine tablet broken into two halves and administered as such (reference formulation) is bioequivalent with one amlodipine tablet broken into two halves and encapsulated (test formulation).

Bioequivalence of inhaled formoterol fumarate assessed from pharmacodynamic, safety and urinary pharmacokinetic data.

This paper deals with a crossover trial on healthy volunteers performed to obtain combined pharmacodynamic, safety and pharmacokinetic data in order to assess the bioequivalence of formoterol fumarate (CAS 43229-80-7) delivered by mono-dose dry powder inhalers, as test and reference. The trial was carried out on 24 Caucasian healthy male and female volunteers treated with 12 micrograms formoterol fumarate bihydrate capsules for inhalation route. Pharmacodynamics was evaluated through a challenge test with methacholine on the forced expiratory volume in 1 s (FEV1). Safety was achieved from glucose and potassium serum levels assayed on timed samples over a 12-h period cost-dosing and from blood pressure, heart rate and ECG recording. Pharmacokinetics was obtained from urinary excretion of formoterol, assessed by a highly sensitive analytical method (LC-MS-MS). Pharmacodynamic, safety and pharmacokinetic results evidenced the bioequivalence of the two formulations investigated. This investigation is an interesting approach how to assess bioequivalence when the classical approach based on the similarity of plasma concentrations can not be applied.

Comparative bioavailability of two formulations containing atenolol and chlortalidone associated in a 4:1 fixed combination.

Atenolol (CAS 29122-68-7) and chlortalidone (CAS 77-36-1) are marketed associated in a 4:1 strength ratio (100/25 and 50/12.5 mg) for the treatment of hypertension. According to EU guidelines, the bioequivalence of one dosage strength can also cover additional strengths when the pharmacokinetics of a given drug is linearly related with the dose. The kinetics of atenolol is linearly correlated with the dose and chlortalidone has linear kinetics with doses < or = 100 mg. Thus this trial carried out on the 100/25 mg strength also covers the 50/12.5 mg strength. The trial was carried out on 18 healthy volunteers (9 males and 9 females) according to a single dose, two-period, two-treatment, two-sequence study design with washout. Timed atenolol plasma concentrations and chlortalidone blood concentrations were used to assess primary pharmacokinetic parameters Cmax, tmax and AUC extrapolated to infinity by a non-compartmental model. The bioavailability of the two formulations was compared through the 90% confidence intervals (C.I.) of Cmax and AUC in accordance with operating guidelines. C.I. of chlortalidone were fully comprised in the 0.80-1.25 range. In the case of atenolol, which displayed a higher data dispersion, C.I. were comprised in the enlarged 0.70-1.43 range. Time to peak, tmax, did not show any statistically significant difference between the test and reference product with respect to both analytes. Pharmacodynamic measurements of the decrease in systolic blood pressure led to fully overlapping results with test and reference. The authors conclude that the test formulation should be considered bioequivalent with the reference with chlortalidone and in the borderline of bioequivalence with atenolol. As no safety problems were involved and pharmacodynamics led to overlapping results as between test and reference, the bioequivalence conclusion could be extended also to atenolol.

Ocular pharmacokinetics of thiamphenicol in rabbits.

The ocular pharmacokinetics of thiamphenicol (TAP, CAS 15318-45-3) was studied in rabbits by means of the assessment of its ocular and systemic absorption, and urinary excretion after instillation of 0.5% TAP eye drops. TAP concentrations in aqueous humor, plasma and urine were evaluated by a coupled LC/GC method (detection limit = 0.1 ng/ml), because the necessity to have a technique much more sensitive than the traditional chromatographic ones available in order to quantify the very low drug concentrations in biological fluids produced by the ocular treatment, and generally by a topical administration. The intravenous route was chosen as reference and allowed the absolute bioavailability to be estimated. TAP proved to be well absorbed through the cornea with the peak aqueous humor concentration of 110 ng/ml at 45 min following the instillation. The good ocular absorption of TAP was confirmed by the plasma concentrations observed after instillation of 0.5% eye drops. In any case, these concentrations were more than 1000 times lower than those observed after the intravenous treatment at the dose normally used for infectious diseases, allowing to exclude any systemic toxicity of TAP eye drops. The absolute ocular bioavailability was 16.2% when estimated from the AUC values and 34.0% from the cumulative urinary excretion values.

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.

Pharmacokinetics of diclofenac after oral administration of its potassium salt in sachet and tablet formulations.

This paper reports the results of a pharmacokinetic study involving 24 healthy volunteers and designed to characterise the rate and extent of diclofenac absorption after the administration of a single dose of diclofenac (CAS 15307-86-5) potassium salt 50 mg in sachet (Voltfast) and tablet (Cataflam) formulations. Timed plasma concentrations of diclofenac during a 12-h-period after dosing were measured by means of HPLC with UV detection at 275 nm and a quantification limit of 10 ng/ml; the method was fully validated for pharmacokinetic purposes. These plasma concentrations were used to calculate Cmax, tmax, trapezoidal AUC0-t and AUC0-infinity and t1/2 by means of noncompartmental analysis. Cmax and tmax are the parameters expressing the rate of absorption, whereas the AUCs reflect the extent of absorption. The rate of absorption with the sachets proved to be very fast, reaching peak values at 10 min in seven subjects and at 15 min in the remaining subjects: mean time was 13.68 min, with concentrations at 5 min being 38% of Cmax. The average time to peak concentration with the tablets was 53.10 min. The extent of absorption of the sachets and tablets was similar, with AUC0-infinity values of respectively 1362 and 1214 ng.ml-1.h, and a 90% confidence interval 1.05-1.20. The highly soluble potassium salt of diclofenac was rapidly absorbed, especially in its sachet formulation, and thus appears to be an invaluable analgesic agent that is particularly useful for quick pain relief.