Population pharmacokinetics and dynamics in phase II studies of the novel bioreductive alkylating cytotoxic indoloquinone EO9.

Population pharmacokinetic-dynamic analysis was prospectively integrated in the clinical phase II programme of EO9 to determine the population pharmacokinetic profile in a larger population of patients, to estimate individual patient pharmacokinetic parameters, and to investigate relationships between drug exposure and clinical outcome. A sparse sampling method was developed, which involved three sampling times, and was implemented during course 1. A Bayesian algorithm was used to estimate individual pharmacokinetic parameters, in particular total plasma clearance (CL) of EO9 and area under the curve (AUC). In total, samples were collected of 85 (65%) of the patients. Pharmacokinetic evaluation was successful in 61 (72%) of the sampled patients. CL of EO9 showed substantial variability (median 5.08 l/min; range 2.67-6.42) and was of the same magnitude as in the phase I study where full pharmacokinetic profiles were used. No significant relationships were noticed between exposure parameters and safety, but overall limited toxicity was observed. No tumor responses were documented. Prospective implementation of large-scale population pharmacokinetic-dynamic analysis is feasible and may generate important findings, in particular when tumor responses and relevant toxicity are observed.

The active site structure of Thalassiosira weissflogii carbonic anhydrase 1.

X-ray absorption spectroscopy at the Zn K-edge indicates that the active site of the marine diatom Thalassiosira weissflogii carbonic anhydrase is strikingly similar to that of mammalian alpha-carbonic anhydrase enzymes. The zinc has three histidine ligands and a single water at 1.98 A. This is quite different from the beta-carbonic anhydrases of higher plants in which zinc is coordinated by two cysteine thiolates, one histidine, and a water molecule. The diatom carbonic anhydrase shows no significant sequence similarity with other carbonic anhydrases and may represent an example of convergent evolution at the molecular level.

Zinc-dependent protein folding.

Studies of classic zinc-finger peptides over the past 15 years have offered insights into the coupled processes of metal binding and protein folding. Within the past two years, this insight has been used to increase our understanding of the importance of first and second shell contributions (i.e. contributions from direct and indirect metal ligands) to metal binding and protein-folding stability, and led to advances in de novo protein design and protein redesign.

The comparative pharmacodynamics of remifentanil and its metabolite, GR90291, in a rat electroencephalographic model.

BACKGROUND: The purpose of this study was to investigate the in vivo pharmacodynamics and the pharmacodynamic interactions of remifentanil and its major metabolite, GR90291, in a rat electroencephalographic model. METHODS: Remifentanil and GR90291 were administered according to a stepwise infusion scheme. The time course of the electroencephalographic effect (0.5-4.5 Hz) was determined in conjunction with concentrations of the parent drug and the metabolite in blood. RESULTS: Administration of remifentanil resulted in concentrations of remifentanil and GR90291 in the ranges 0-120 ng/ml and 0-850 ng/ml, respectively. When the metabolite was administered, concentrations of the metabolite in the range 0-220 microg/ml and no measurable concentrations of remifentanil were observed. The mean +/- SE values of the pharmacokinetic parameters clearance and volume of distribution at steady state were 920+/-110 ml x min(-1) x kg(-1) and 1.00+/-0.93 l/kg for remifentanil and 15+/-2 ml x min(-1) x kg(-1) and 0.56+/-0.08 l/kg for GR90291. The relative free concentrations in the brain, as determined on the basis of the cerebrospinal fluid/total blood concentration ratio at steady state, were 25+/-5% and 0.30+/-0.11% for remifentanil and GR90291, respectively. Concentration-electroencephalographic effect relations were characterized on the basis of the sigmoidal Emax pharmacodynamic model. The mean +/- SE values for the maximal effect (Emax), the concentration at which 50% of the maximal effect is obtained (EC50), and Hill factor for remifentanil were 109+/-12 microV, 9.4+/-0.9 ng/ml, and 2.2+/-0.3, respectively (n = 8). For GR90291, the mean +/- SE values for EC50 and the Hill factor were 103,000+/-9,000 microg/ml and 2.5+/-0.4, respectively (n = 6). CONCLUSIONS: Analysis of the data on the basis of a previously postulated, mechanism-based pharmacokinetic-pharmacodynamic model for synthetic opioids revealed that the low in vivo potency of GR90291 can be explained by a low affinity to the mu-opioid receptor in combination with a poor brain penetration.

A population pharmacokinetic-pharmacodynamic analysis of repeated measures time-to-event pharmacodynamic responses: the antiemetic effect of ondansetron.

This paper presents and illustrates methodology for specifying, estimating, and evaluating a predictive model for repeated measures time-to-event responses. The illustrative example specifies a model of the antiemetic effect vs. concentration relationship for the 5-HT3 antagonist ondansetron in the human ipecac model for emesis. A key part of this model is a time-dependent log hazard function for emesis that is increased by ipecac administration and decreased by ondansetron concentration. The model is fit using an approximate maximum likelihood method. The data consist of the time free of emeses and, for those individuals with emetic episodes, the time(s) of the episode(s). Model evaluation is accomplished using residual plots adapted to time-to-event data and a "posterior predictive check" wherein observed data statistics are compared to those obtained from data simulated from the fitted model. The ondansetron concentration required to obtain a 50% reduction in the hazard of emesis is estimated to be 1.4 +/- 0.2 ng/ml, and the rate constant for elimination of ipecac-induced hazard is 1.5 +/- 0.2 hr-1.

Pharmacokinetic-pharmacodynamic modelling of the EEG effect of alfentanil in rats: assessment of rapid functional adaptation.

1. The purpose of the present investigation was to quantify rapid functional adaptation in the concentration-pharmacological effect relationship of alfentanil in rats using quantitative EEG parameters as a pharmacodynamic endpoint. Three groups of 6-7 rats received in a randomized fashion two consecutive infusions of 2.00, 3.14, or 4.24 mg/kg(-1) of alfentanil in 20, 40 or 60 min, respectively. The EEG was continuously recorded and frequent arterial blood samples were collected for determination of the alfentanil concentration by gas chromatography. 2. The pharmacokinetics of alfentanil were most adequately described by a bi-exponential function. The values (mean+/-s.e., n=20) of clearance, volume of distribution at steady-state and terminal half-life were 45+/-3 ml x min(-1) x kg(-1), 0.91+/-0.09 l/kg(-1) and 23+/-1 min, respectively, and independent of the administered dose. 3. Increase in power in the 0.5-4.5 Hz (delta) frequency band of the EEG was used as the measure of the pharmacological response. By pharmacokinetic-pharmacodynamic modeling the individual concentration-EEG effect relationships of alfentanil were derived which were successfully quantified by the sigmoidal Emax pharmacodynamic model. When the results of the first of the two consecutive infusions were compared, no systematic differences in the pharmacodynamic parameters were observed for the different infusion rates. The averaged values of the pharmacodynamic parameters of alfentanil were (mean+/-s.e., n=20): E0=56+/-3 microV, Emax=93+/-8 microV, EC50=235+/-27 ng x ml(-1) and Hill factor=1.6+/-0.1, respectively. For the second of the two consecutive infusions a significantly higher value of the EC50 of 404+/-56 ng x ml(-1) was observed (P < 0.05), while the values of the other pharmacodynamic parameters were unchanged. Simulations according to a mechanism-based model indicated that the observed change in concentration effect relationship can be explained by a 40% loss of functional mu-opioid receptors. 4. The results of the present study show that upon the administration of a single intravenous dose, acute functional adaptation does not interfere with the assessment of the concentration-EEG effect relationship of alfentanil. Upon repeated administration however functional adaptation may be a complicating factor.

Pharmacokinetic-pharmacodynamic modelling of the analgesic effect of alfentanil in the rat using tooth pulp evoked potentials.

The purpose of the present study was to develop an animal model for the investigation of the concentration-effect relationship of alfentanil using tooth pulp evoked potentials (TPEP). Six chronically instrumented, freely moving rats received a computer-controlled intravenous infusion of alfentanil resulting in seven pseudo-steady-state blood concentration levels, each maintained for 30 min. At each concentration level, the tooth pulp of the rat upper incisor was electrically stimulated in a time-randomized order with different current intensities (400-800 microA, 1 msec duration) and the electroencephalogram (EEG) was recorded concomitantly. Arterial blood samples were collected serially and assayed for alfentanil using RIA. Repetitive evoked EEG responses were averaged per stimulus intensity and per session. The decrease of the area under the negative peak 15 msec after stimulation (% of preadministration value) was used as pharmacological endpoint. The concentration-TPEP effect of alfentanil was investigated by nonlinear mixed effect modeling (NONMEM). When the observed TPEP effect was plotted versus the alfentanil blood concentration no hysteresis or proteresis was observed, and the two could directly be related to each other on the basis of the sigmoidal Emax pharmacodynamic model. The (population) pharmacodynamic estimates were (+/-S.E.): Emax = 108 +/- 10%, EC50 = 24 +/- 17 ng/ml, Hill factor = 0.81 +/- 0.37. A large interindividual variability for EC50 (omegaEC50) of 164 +/- 107% was observed. The residual variability was 14 +/- 10%. It is concluded that the TPEP is a useful tool for the systematic investigation of the concentration-analgesic effect relationship of centrally acting analgesics in the freely moving rat.

Influence of different fat emulsion-based intravenous formulations on the pharmacokinetics and pharmacodynamics of propofol.

PURPOSE: The influence of different intravenous formulations on the pharmacokinetics and pharmacodynamics of propofol was investigated using the effect on the EEG (11.5-30 Hz) as pharmacodynamic endpoint. METHODS: Propofol was administered as an intravenous bolus infusion (30 mg/kg in 5 min) or as a continuous infusion (150 mg/kg in 5 hours) in chronically instrumented male rats. Propofol was formulated as a 1% emulsion in an Intralipid 10%-like fat emulsion (Diprivan-10, D) or as a 1%- or 6% emulsion in Lipofundin MCT/LCT-10% (P1% and P6%, respectively). EEG was recorded continuously and arterial blood samples were collected serially for the determination of propofol concentrations using HPLC. RESULTS: Following bolus infusion, the pharmacokinetics of the various propofol emulsions could adequately be described by a two-compartmental pharmacokinetic model. The average values for clearance (Cl), volume of distribution at steady-state (Vd,ss) and terminal half-life (t1/2, lambda 2) were 107 +/- 4 ml/min/kg, 1.38 +/- 0.06 l/kg and 16 +/- 1 min, respectively (mean +/- S.E. n = 22). No significant differences were observed between the three propofol formulations. After continuous infusion these values were 112 +/- 11 ml/min/kg, 5.19 +/- 0.41 l/kg and 45 +/- 3 min, respectively (mean +/- S.E., n = 20) with again no statistically significant differences between the three propofol formulations. Comparison between the bolus- and the continuous infusion revealed a statistically significant difference for both Vd,ss and t1/2, lambda 2 (p < 0.05), whereas Cl remained unchanged. In all treatment groups infusion of propofol resulted in a burst-suppression type of EEG. A profound hysteresis loop was observed between blood concentrations and EEG effect for all formulations. The hysteresis was minimized by a semi-parametric method and resulted in a biphasic concentration-effect relationship of propofol that was described non-parametrically. For P6% a larger rate constant onset of drug effect (t1/2,keo) was observed compared to the other propofol formulations (p < 0.05). CONCLUSIONS: The pharmacokinetics and pharmacodynamics of propofol are not affected by to a large extent the type of emulsion nor by the concentration of propofol in the intravenous formulation.

Pharmacokinetic-pharmacodynamic modeling of the electroencephalogram effect of synthetic opioids in the rat: correlation with the interaction at the mu-opioid receptor.

The purpose of our investigation was to characterize the relationships between the pharmacodynamics of synthetic opioids in vivo and the interaction at the mu-opioid receptor. The pharmacokinetics and pharmacodynamics were determined in vivo after a single i.v. infusion of 3.14 mg/kg alfentanil (A), 0.15 mg/kg fentanyl (F) or 0.030 mg/kg sufentanil (S) in rats. Amplitudes in the 0.5 to 4.5 Hz frequency band of the electroencephalogram (EEG) was used as pharmacodynamic endpoint. The EEG effect intensity was related to the (free) concentration in blood (A) or in a hypothetical effect compartment (F, S) on basis of the sigmoidal Emax pharmacodynamic model. The interaction at the mu-opioid receptor was determined in vitro on basis of the displacement of [3H]-naloxone binding in washed rat brain membranes. The value of the sodium shift was used as a measure of in vitro intrinsic efficacy. For the EEG effect the in vivo potencies based on free drug concentrations (EC50,u) were 4.62 +/- 0.66 ng/ml (A), 0.69 +/- 0.05 ng/ml (F) and 0.29 +/- 0.06 ng/ml (S). In the receptor binding studies the affinities at the mu-opioid receptor (Kl) were 47.4 +/- 6.6 nM (A), 8.6 +/- 4.1 nM (F) and 2.8 +/- 0.2 nM (S). For each opioid the ratio between EC50,u and Kl was the same with a value of 0.23-0.25, indicating the existence of receptor reserve for the EEG effect. The intrinsic activity (Emax) of the three opioids in vivo was similar with values of 111 +/- 10 microV (A), 89 +/- 11 microV (F) and 104 +/- 4 microV (S). However, the values of the sodium shift varied between 2.8 (S) and 19.1 (A). Further analysis of the in vivo pharmacodynamic data on basis of an operational model of agonism provided evidence for a large receptor reserve, which explains why compounds with different values of the sodium shift all behave as full agonists in vivo.

Pharmacokinetic-pharmacodynamic modelling of the EEG effect of alfentanil in rats.

The purpose of the present investigation was to develop a methodology for quantification of the concentration-pharmacological effect relationship of alfentanil in individual rats by using effect parameters derived from quantitative EEG analysis. In particular, the role of the opioid-induced side effects of proconvulsant activity, hypothermia, and respiratory depression was investigated. Amplitudes in the 0.5- to 4.5-Hz frequency band of the EEG power spectrum were used as a descriptor of the effect. The pharmacokinetics and pharmacodynamics of alfentanil were determined after intravenous administration of 2000 micrograms/kg during 20 min. The administration of alfentanil induced paroxysmal seizure patterns in the EEG which made meaningful analysis of the EEG effect impossible. A constant infusion of midazolam (5.5 mg/kg/hr) prevented alfentanil-induced seizures. When no precautions were taken to control body temperature, analysis of the concentration-EEG effect relationship was complicated by proteresis due to alfentanil-induced hypothermia. This proteresis disappeared when body temperature was stabilized at 37.5 degrees-38.5 degrees C with isothermal pads. Alfentanil-induced respiratory depression was managed successfully by artificial ventilation. Adequateness of artificial ventilation was ascertained by monitoring of arterial pO2, pCO2, and pH. When these opioid side effects were controlled, the pharmacokinetics were most adequately described by a biexponential function. The values of the pharmacokinetic parameters were (mean +/- SE, n = 7); clearance = 53 +/- 6 ml.min/kg, volume of distribution = 1.19 +/- 0.19 l/kg and terminal half-life = 24.5 +/- 2.3 min. By pharmacokinetic-pharmacodynamic modelling, the individual concentration-effect relationships of alfentanil were derived, which were successfully characterized by the sigmoidal Emax pharmacodynamic model. The values of the pharmacodynamic parameters were (mean +/- SE, n = 7): E0 = 57 +/- 4 microV, Emax = 95 +/- 17 microV, EC50 = 202 +/- 55 ng/ml and Hill factor = 1.53 +/- 0.20. No delay was observed between alfentanil concentration and effect. The results of the present study show that when side effects are controlled adequately, the concentration-EEG effect relationship of alfentanil can be characterized in individual rats using amplitudes in the 0.5- to 4.5-Hz frequency band of the EEG as a measure of the pharmacological response. The described methodology can be very useful for detailed studies into the pharmacodynamics of (new) synthetic opioids in vivo.

Platinum(II)-adenosine phosphothiorate complexes: kinetics of formation and phosphorus-31 NMR characterization studies.

Reactions of chloro(diethylenetriamine)platinum(II) chloride with adenosine 5'-O-thiomonophosphate, adenosine 5'-O-(2-thiodiphosphate), and adenosine 5'-O-(3-thiotriphosphate) yielded exclusively (phosphothiorato)platinum(II) complexes. Phosphorus-31 NMR data for the coordinated phosphothiorate phosphorus atom exhibited about 15-20 ppm upfield chemical shift compared to chemical shifts for free nucleotides. Uncoordinated phosphate groups exhibited insignificant changes in the chemical shift upon complexation. Likewise, proton NMR data indicate no significant changes in chemical shift for the purine or ribose protons. Reactions between phosphothiorates and the platinum complex predominately take place through a second-order process, first order with respect to each of the reactants indicating that the aquated pathway contributes insignificantly toward complexation. The second-order rate constants, 1.9 +/- 0.1 M-1 s-1 for the AMP-S, 2.4 +/- 0.2 M-1 s-1 for the ADP-beta-S, and 2.7 +/- 0.2 M-1 s-1 for the ATP-gamma-S reactions were evaluated at pH 6.5 and at 25 degrees C. These rate data were compared with those reactions of adenosine 5'-monophosphate (AMP) and guanosine 5'-monophosphate (GMP) with the same platinum(II) complex. These reactions proceed through the direct interaction between the starting platinum complex and nucleotides as well as through the reaction between the aquaplatinum complex and nucleotides. The rate constant for the aquation process was evaluated to be (2.0 +/- 0.1) x 10(-4) s-1 for both AMP and GMP reactions.(ABSTRACT TRUNCATED AT 250 WORDS)

Direct and correlated responses to selection for weaning weight, post-weaning weight gain and six-week weight in mice.

Four lines of mice were formed from a common base population and selected for 37 generations for either increased 3-week weight (weaning weight), 6-week weight, 3-6 week gain, or maintained as a randomly bred control line. Realised heritability estimates for short-term (long-term) responses were 0.33±0.20 (0.07±0.10), 0.46±0.14 (0.26±0.09), 0.36±0.14 (0.24±0.11) for 3-week weight, 6-week weight and 3-6 week gain, respectively. Realised genetic correlations estimated from short-term (long-term) responses were 0.23±0.08 (0.35±0.10) between 3-week weight and 3-6 week gain; 0.82±0.04 (0.58±0.08) between 3-week weight and 6-week weight; and 0.81±0.04 (0.97±0.04) between 3-6 week gain and 6-week weight. The genetic correlation between 3-week weight and 6-week weight was asymmetric with a greater correlated response for 3-week weight when selecting for 6-week weight (1.06) than vice versa (0.63).