Effect of quinidine on the tissue binding of digoxin in guinea-pigs

The mechanism of quinidine-induced decrease in the tissue distribution of digoxin to heart, liver and skeletal muscle has been examined in guinea-pigs. Quinidine, in the presence of adenosine-5'-triphosphate (ATP), inhibited the specific binding of digoxin in homogenates of heart, liver and muscle, while in the absence of ATP the inhibition was observed only in heart. The decrease in the tissue-to-plasma unbound concentration ratios (Kpu) of heart and muscle determined from in-vitro binding studies was comparable to that in the Kpu values observed in-vivo, while in liver it was not sufficient to account for the fall in Kpuin-vivo values. It is concluded that quinidine-induced decrease in the tissue distribution of digoxin in heart and muscle is due to inhibition of tissue binding of this drug, while that in liver could be partially attributed to the decrease in the tissue binding.     

The kinetics and tissue distribution of protein transduction in mice.

Protein transduction domains (PTDs) offer an exciting therapeutic opportunity for the treatment of many diseases. An 11-amino acid fragment of human immunodeficiency type 1 (HIV-1) TAT-protein can transduce large, biologically active proteins into mammalian cells; recent evidence has shown an in vivo PTD for the 116 kDa beta-galactosidase protein. However, there is little information on the in vivo distribution of the TAT fusion protein to define the viability of PTDs for human studies. In this study we examined the tissue kinetics and tissue distribution of the PTD-transduced TAT fusion protein in mice. Low (100 microg) or high (500 microg) doses of TAT-beta-galactosidase fusion protein were administrated to mice through four routes (portal vein, i.v., i.p., and oral). Tissues were harvested 15 min, 1h, 6h, 10h, and 24h after treatment. Distribution of beta-galactosidase in various tissues was analysed by in situ staining, enzymatic activity assay, and Western blot analysis. Beta-galactosidase enzyme activity was observed in all tissues (liver, kidney, spleen, lung, bowel, and brain). Beta-galactosidase activity peaked at 15 min in most tissues after portal vein, i.v., and i.p. administration and at 1h after oral dosing in all tissues. Beta-galactosidase activity in the liver at 15 min after portal vein injection (67 milliunits [mU]/mg) was higher than after i.v. (9.8 mU/mg), i.p. (4.4 mU/mg), and oral (0.3 mU/mg) dosing. In situ staining and Western blot results correlated closely with beta-galactosidase enzyme activity assay. The median initial half-life for activity was 2.2h, ranging from 1.2h to 3.4h (coefficient of variation=28.9%). The bioavailability of beta-galactosidase activity after an orally administered PTD was 24%. This study details the kinetics and tissue distribution of delivering of a model TAT fusion protein into the mouse via PTD. These data allow rational selection of delivery route and schedules for therapeutic PTD and will aid the use of TAT fusion protein transduction in the development of protein therapies.     

A strategy for residual error modeling incorporating scedasticity of variance and distribution shape

Nonlinear mixed effects models parameters are commonly estimated using maximum likelihood. The properties of these estimators depend on the assumption that residual errors are independent and normally distributed with mean zero and correctly defined variance. Violations of this assumption can cause bias in parameter estimates, invalidate the likelihood ratio test and preclude simulation of real-life like data. The choice of error model is mostly done on a case-by-case basis from a limited set of commonly used models. In this work, two strategies are proposed to extend and unify residual error modeling: a dynamic transform-both-sides approach combined with a power error model (dTBS) capable of handling skewed and/or heteroscedastic residuals, and a t-distributed residual error model allowing for symmetric heavy tails. Ten published pharmacokinetic and pharmacodynamic models as well as stochastic simulation and estimation were used to evaluate the two approaches. dTBS always led to significant improvements in objective function value, with most examples displaying some degree of right-skewness and variances proportional to predictions raised to powers between 0 and 1. The t-distribution led to significant improvement for 5 out of 10 models with degrees of freedom between 3 and 9. Six models were most improved by the t-distribution while four models benefited more from dTBS. Changes in other model parameter estimates were observed. In conclusion, the use of dTBS and/or t-distribution models provides a flexible and easy-to-use framework capable of characterizing all commonly encountered residual error distributions.     

Contribution of aranidipine metabolites with slow binding kinetics to the vasodilating activity of aranidipine

Aranidipine, a novel dihydropyridine derivative, gives rise to two active metabolites, M-1(α) and M-1(β), which exhibit hypotensive activity comparable to that of nifedipine. The aim of this study was to examine the recovery phase of the vasodilating effect of M-1(α) and of M-1(β), to determine their binding characteristics and to compare the results with those for aranidipine and other dihydropyridine derivatives. During intra-arterial infusion into the femoral vascular beds of anesthetized dogs, M-1(α), M-1(β), and nifedipine, produced increases in femoral blood flow at doses three times higher than the dose of aranidipine required to produce a comparable effect. The onset and recovery of the effects of the metabolites on femoral blood flow were significantly slower than the onset and recovery of the effect of nifedipine. The inhibitory activities of M-1(α) and M-1(β) towards stimulated ⁴⁵Ca uptake in isolated guinea pig aorta were less than that of aranidipine. In binding studies, using porcine heart membrane preparations, [³H]M-1(α) and [³H]M-1(β) had larger K_d values than [³H]aranidipine and [³H]nitrendipine, but the maximal binding number for each of them was almost the same. The association and dissociation rate constants for [³H]M-1(α) and [³H]M-1(β) binding, as well as those for [³H]aranidipine binding, were significantly smaller than those for [³H]nitrendipine, corresponding to the recovery of the in vivo vasodilating effects of the metabolites. The dissociation rate constants of these radiolabeled ligands were highly positively correlated with the elimination rate constants of their in vivo vasodilating effects. From these results, we conclude that M-1(α) and M-1(β), although their binding affinities and Ca²⁺ antagonistic actions are less potent, possess slower kinetic binding properties than many other dihydropyridines and that the slow kinetic interaction of these metabolites with the dihydropyridine receptor may contribute to the long-lasting in vivo vasodilating effect of aranidipine. Received: 2 January 1996 / Accepted: 30 August 1996     

Skeletal muscle digoxin binding in patients with renal failure.

The effect of inhibition of prostaglandin synthesis on the systemic clearance of indocyanine green and antipyrine was studied in seven subjects. Antipyrine clearance was not altered by indomethacin suggesting that oxidative metabolism was not affected. Both aspirin and indomethacin decreased the clearance of indocyanine green presumably by reducing liver blood flow. These results suggest that an effect of inhibitors of prostaglandin synthesis on hepatic drug clearance is likely to be confined to high clearance drugs when given systemically.     

Hepatic macromolecular binding and tissue distribution of ortho- and para-toluidine in rats.

The in vivo covalent binding of ortho- and para-toluidine (OT and PT) to rat hepatic macromolecules was investigated to determine if a relationship exists between the degree of binding for each isomer and its carcinogenic potency. The ortho-isomer has been shown to be a more potent hepatocarcinogen than the para-isomer. In addition to the macromolecular binding, the tissue distribution of each isomer was also measured. The degree of binding to hepatic macromolecules appeared to be at maximum for both at 24 28 h following dosing. At 24 h following dosing, the level of DNA binding of OT was approximately 1.2-fold lower than that of PT. The binding to RNA and protein was also lower for OT than PT, although the differences were not as great as that observed for DNA binding. There were subtle differences in tissue distribution for each isomer. However, in contrast to the macromolecular binding data, the area under the plasma concentration curve for OT was approximately 1.8-fold greater than that for PT. Based on the results of these studies, there was no direct correlation between the degree of macromolecular binding and carcinogenic potency.     

Protein binding of digoxin in human serum

Summary The protein binding of digoxin in human serum was investigated using equilibrium dialysis and tritium-labelled digoxin. A constant protein bound fraction of about 30% was found over a wide range of concentrations of digoxin including therapeutic levels. Interpretation of the findings according to the law of mass-action showed an association constantK = 0.68·10^–5l/mol; and, the number of binding sites,n , indicating an almost infinite apparent maximum binding capacity and a very small affinity of human serum proteins for digoxin.Supported by the Schweizer Nationalfond zur Förderung der wissenschaftlichen Forschung.     

The plasma kinetics of digoxin-specific Fab fragments and digoxin in the rabbit.

The plasma kinetics of total and free digoxin, and digoxin-specific antibody fragments (DSFab) in rabbits which had been given [3H]digoxin one hour before DSFab has been studied over a 5 day period. Injection of DSFab caused a 4- to 5-fold rise in total digoxin and reduced elimination half-life (t1/2 beta), apparent volume of distribution at steady-state (Vdss) and systemic clearance (CL) by 40, 90 and 75% respectively. Early in the experimental period, DSFab reduced free digoxin concentration (measured by ultrafiltration) from 4.1 ng mL-1 to a minimum of 1.3 ng mL-1 at 15 min. However, the concentration had rebound to 2.5 ng mL-1 by 60 min. Subsequently, free digoxin fell to 0.63 ng mL-1 and remained relatively constant over a 7 to 90 h period. The distribution half-life, t1/2 beta, Vdss and CL for DSFab (concentrations measured by enzyme-linked immunosorbent assay) were 0.3 h, 3.2 h, 185 mL kg-1 and 57 mL kg-1 h-1, respectively. A considerable molar excess (about 5) of DSFab in the plasma was necessary to maintain minimum free digoxin concentrations. When the DSFab:digoxin molar ratio was less than 4 during the initial treatment period, free (toxicologically active) concentrations increased. With the elevation in total digoxin, however, an opposite situation appeared to apply. By 24 h the relatively short DSFab t1/2 beta meant that the plasma DSFab concentration was less than 0.05 micrograms mL-1 giving a DSFab:digoxin molar ratio of below 0.06, yet the antibody-induced rise in total digoxin concentration was still detectable at 100 h.     

Accumulation kinetics of drugs with nonlinear plasma protein and tissue binding characteristics

The purpose of this investigation was to study, by digital computer stimulation, the accumulation kinetics of drugs which exhibit concentration-dependent binding to tissues and either linear (constant free fraction) or concentration-dependent (increasing free action with increasing drug concentration) binding to plasma proteins. It was assumed that elimination rate is proportional to free drug concentration in plasma and that there occurs instantaneous equilibration of drug between vascular and nonvascular spaces. Nonlinear binding can yield, under certain conditions, apparently biexponential plasma concentration-time curves which may be misinterpreted as being representative of a linear and biexponential system. Such misinterpretation would cause the following errors in the prediction of drug accumulation and elimination kinetics during and after constant-rate infusion: (a) the time required to reach steady state may be overestimated, and (b) the prominence of the apparent distribution phase after cessation of infusion may be underestimated. Drugs with linear and nonlinear plasma protein binding characteristics differ with respect to the relaionship between infusion rate and steady-state concentration. This relationship is linear when plasma protein binding is linear. Steady-state concentration increases less than proportionally with increasing infusion rate if plasma protein binding is drug concentration dependent.     

Accumulation kinetics of drugs with nonlinear plasma protein and tissue binding characteristics

The purpose of this investigation was to study, by digital computer simulation, the accumulation kinetics of drugs which exhibit concentration-dependent binding to tissues and either linear (constant free fraction) or concentration-dependent (increasing free fraction with increasing drug concentration) binding to plasma proteins. It was assumed that elimination rate is proportional to free drug concentration in plasma and that there occurs instantaneous equilibration of drug between vascular and nonvascular spaces. Nonlinear binding can yield, under certain conditions, apparently biexponential plasma concentrationtime curves which may be misinterpreted as being representative of a linear and biexponential-system. Such misinterpretation would cause the following errors in the prediction of drug accumulation and elimination kinetics during and after constantrate infusion: (a) the time required to reach steady state may be overestimated, and (b) the prominence of the apparent distribution phase after cessation of infusion may be underestimated. Drugs with linear and nonlinear plasma protein binding characteristics differ with respect to the relationship between infusion rate and steadystate concentration. This relationship is linear when plasma protein binding is linear. Steadystate concentration increases less than proportionally with increasing infusion rate if plasma protein binding is drug concentration dependent.     

Volumes of distribution and mean residence time of drugs with linear tissue distribution and binding and nonlinear protein binding

Based on a generalized model, equations for calculating the mean residence time in the body at single dose (MRT) and at steady state (MRT _ss), apparent steady-state volume of distribution ( ) and steady-state volume of distribution (V _ss) are derived for a drug exhibiting nonlinear protein binding. Interrelationships between andV _ss as well as betweenMRT andMRT _ss are also discussed and illustrated with simulated data. In addition, a method for estimating the central volume of distribution of the bound drug and the sum of the central volume of distribution of the unbound drug and the area under the first moment curve of distribution function for drugs with nonlinear protein binding is proposed and illustrated with both simulated and published data.     

Determination of volume of distribution at steady state with complete consideration of the kinetics of protein and tissue binding in linear pharmacokinetics

The assumption of an instant equilibrium between bound and unbound drug fractions is commonly applied in pharmacokinetic calculations. The equation for the calculation of the steady-state volume of distribution V(ss) from the time curve of drug concentration in plasma after intravenous bolus dose administration, which does not assume an immediate equilibrium and thus incorporates dissociation and association rates of protein and tissue binding, is presented. The equation obtained V(ss) = (Dose/AUC)*MRT(u) looks like the traditional equation, but instead of mean residence time MRT calculated using the total drug concentration in plasma, it contains mean residence time MRT(u) calculated using the plasma concentration of the unbound drug. The equation connecting MRT(u) and MRT is derived. If an immediate equilibrium between bound and unbound drug fractions occurs, MRT(u) and MRT are the same, but in general, MRT(u) is always smaller than MRT. For drugs with high protein affinity and slow dissociation rate MRT(u) may be of an order of several hours smaller than MRT, so that V(ss) can be considerably overestimated in the traditional calculation.     

Mechanistic models enable the rational use of in vitro drug-target binding kinetics for better drug effects in patients

Introduction: Drug-target binding kinetics are major determinants of the time course of drug action for several drugs, as clearly described for the irreversible binders omeprazole and aspirin. This supports the increasing interest to incorporate newly developed high-throughput assays for drug-target binding kinetics in drug discovery. A meaningful application of in vitro drug-target binding kinetics in drug discovery requires insight into the relation between in vivo drug effect and in vitro measured drug-target binding kinetics.

Areas covered: In this review, the authors discuss both the relation between in vitro and in vivo measured binding kinetics and the relation between in vivo binding kinetics, target occupancy and effect profiles.

Expert opinion: More scientific evidence is required for the rational selection and development of drug-candidates on the basis of in vitro estimates of drug-target binding kinetics. To elucidate the value of in vitro binding kinetics measurements, it is necessary to obtain information on system-specific properties which influence the kinetics of target occupancy and drug effect. Mathematical integration of this information enables the identification of drug-specific properties which lead to optimal target occupancy and drug effect in patients.     

Enantioselective pharmacokinetics of etodolac in the rat: tissue distribution, tissue binding, and in vitro metabolism.

The nonsteroidal anti-inflammatory agent etodolac (ET) exhibits stereoselectivity in its pharmacokinetics following administration to humans and rats. To underline the factors responsible for this stereoselectivity, the tissue distribution, in vitro tissue binding, and microsomal metabolism of ET enantiomers were studied in the rat. Following iv administration of racemic ET, the S:R AUC ratios in tissues were stereoselective, and different from that in plasma. Binding of enantiomers to tissues was stereoselective, although it did not relate well with in vivo tissue distribution. Rather, the tissue distribution of enantiomers appeared to be better explained by the unbound fractions of enantiomers in plasma. With respect to in vitro glucuronidation by liver microsomes, the Vmax of S-ET was 3.4-fold greater than that of R-ET; the enantiomers possessed similar Km. There appeared to be stereoselectivity in the oxidative metabolism of ET enantiomers by liver and kidney microsomes, in favor of the R-enantiomer. The lower AUC in rat plasma of pharmacologically active S-ET as compared with its antipode is due to its relatively greater distribution to tissues, owing to a lesser degree of binding to plasma proteins, and to its higher rate of glucuronidation.     

Comparative kinetics of serum and vitreous humor digoxin concentrations in a guinea pig model. Part I: Intravenous administration of digoxin

The pharmacokinetics of a single intravenous dose of digoxin in the guinea pig was investigated with emphasis on the penetration of digoxin into the vitreous humor. A controlled study was undertaken and data was collected which indicated that digoxin follows an open, two-compartment pharmacokinetic model with a terminal half-life of 318 minutes. The data indicated that the ratio of vitreous concentrations to serum concentrations were determined to be equal following an initial tissue distribution phase.     

Effect of ketanserin on the kinetics of digoxin and digitoxin

The influence of the serotonin antagonist ketanserin on the kinetics of digoxin and digitoxin were evaluated in healthy volunteers. Subjects received a single intravenous dose of digoxin (1.25 mg) or of digitoxin (1.0 mg) on two occasions: once in the control state, and again during treatment with ketanserin, 40 mg twice daily. Ketanserin caused a prolongation of digoxin elimination half-life (50 versus 40 h) and reduction in clearance (2.4 versus 3.7 ml/min/kg), but differences were not significant. For digitoxin, ketanserin caused a small but significant (p less than 0.005) increase in volume of distribution (0.89 versus 0.78 L/kg), but no significant effect on half-life (7.0 versus 6.8 days), or clearance (0.063 versus 0.059 ml/min/kg). Thus, ketanserin coadministration is not likely to alter steady-state concentrations of digoxin or digitoxin during clinical use of these two digitalis glycosides.     

Pharmacokinetics and tissue distribution of human urinary tumor necrosis factor binding protein in mice.

Iodinated natural human urinary tumor necrosis factor binding protein I (125I-uTBP) was iv injected into BALB/c mice, and its pharmacokinetics and tissue distribution were assessed during a short-term (0-1 hr) and for a long-term (0-24 hr) period. The blood 125I-uTBP concentration displayed a biphasic pattern that was adequately described by a biexponential function with estimated half-lives of 0.1 and 3.8 hr. The apparent volume of distribution (Vc) of the central compartment was 3 ml, which approximated the mouse blood volume. The clearance (CL) derived either from a model-dependent or a model-independent method of analysis was 2.5 and 2.9 ml/hr, respectively. One hr after the iv administration of 125I-uTBP, the radioactivity accumulated in the major organs and tissues. The highest concentrations in terms of pg per organ were seen in the skin and in the liver. When expressed as pg 125I-uTBP per mg organ, the distribution was the highest in the gallbladder, bladder, kidneys, and lungs. At 24 hr, the distribution of 125I-uTBP represented about 10% of the amount measured at 1 hr. The rank order of accumulation of the radiolabeled uTBP in the major organs, expressed as pg per organ at 24 hr was skin greater than liver greater than kidneys greater than lungs greater than gut greater than spleen greater than gallbladder.