Search

Zebrafish larvae are increasingly used for pharmacological research, but internal drug exposure is often not measured. Understanding pharmacokinetics is necessary for reliable translation of pharmacological results to higher vertebrates, including humans. Quantification of drug clearance and distribution requires measurements of blood concentrations. Additionally, measuring drug metabolites is of importance to understand clearance in this model organism mechanistically. We therefore mechanistically studied and quantified pharmacokinetics in zebrafish larvae, and compared this to higher vertebrates, using paracetamol (acetaminophen) as a paradigm compound. A method was developed to sample blood from zebrafish larvae 5 days post fertilization. Blood concentrations of paracetamol and its major metabolites, paracetamol-glucuronide and paracetamol-sulfate, were measured. Blood concentration data were combined with measured amounts in larval homogenates and excreted amounts and simultaneously analyzed through nonlinear mixed-effects modeling, quantifying absolute clearance and distribution volume. Blood sampling from zebrafish larvae was most successful from the posterior cardinal vein, with a median volume (interquartile range) of 1.12 nl (0.676–1.66 nl) per blood sample. Samples were pooled (n= 15–35) to reach measurable levels. Paracetamol blood concentrations at steady state were only 10% of the external paracetamol concentration. Paracetamol-sulfate was the major metabolite, and its formation was quantified using a time-dependent metabolic formation rate. Absolute clearance and distribution volume correlated well with reported values in higher vertebrates, including humans. Based on blood concentrations and advanced data analysis, the mechanistic and quantitative understanding of paracetamol pharmacokinetics in zebrafish larvae has been established. This will improve the translational value of this vertebrate model organism in drug discovery and development.

ABSTRACTThe linear pharmacokinetics (PK) of therapeutic monoclonal antibodies (mAbs) can be considered a class property with values that are similar to endogenous IgG. Knowledge of these parameters across species could be used to avoid unnecessary in vivoPK studies and to enable early PK predictions and pharmacokinetic/pharmacodynamic (PK/PD) simulations. In this work, population-pharmacokinetic (popPK) modeling was used to determine a single set of ‘typical’ popPK parameters describing the linear PK of mAbs in human, cynomolgus monkey and transgenic mice expressing the human neonatal Fc receptor (hFcRn Tg32), using a rich dataset of 27 mAbs. Non-linear PK was excluded from the datasets and a 2-compartment model was applied to describe mAb disposition. Typical human popPK estimates compared well with data from comparator mAbs with linear PK in the clinic. Outliers with higher than typical clearance were found to have non-specific interactions in an affinity-capture self-interaction nanoparticle spectroscopy assay, offering a potential tool to screen out these mAbs at an early stage. Translational strategies were investigated for prediction of human linear PK of mAbs, including use of typical human popPK parameters and allometric exponents from cynomolgus monkey and Tg32 mouse. Each method gave good prediction of human PK with parameters predicted within 2-fold. These strategies offer alternative options to the use of cynomolgus monkeys for human PK predictions of linear mAbs, based on in silicomethods (typical human popPK parameters) or using a rodent species (Tg32 mouse), and call into question the value of completing extensive in vivopreclinical PK to inform linear mAb PK.

ABSTRACTIntroduction: Diseases of the Central Nervous System (CNS) affect millions of people worldwide, with the number of people affected quickly growing. Unfortunately, the successful development of CNS-acting drugs is less than 10%, and this is attributed to the complexity of the CNS, unexpected side effects, difficulties in penetrating the blood-brain barrier and lack of biomarkers.Areas covered: Herein, the authors first review how pharmacokinetic/pharmacodynamic (PK/PD) models are designed to predict the dose-dependent time course of effect, and how they are used to translate drug effects from animal to man. Then, the authors discuss how pharmacometabolomics gives insight into system-wide pharmacological effects and why it is a promising method to study interspecies differences. Finally, the authors advocate the application of PK/PD-metabolomics modeling to advance translational CNS drug development by discussing its opportunities and challenges.Expert opinion: It is envisioned that PK/PD-metabolomics will increase our understanding of CNS drug effects and improve translational CNS drug development, thereby increasing success rates. The successful future development of this concept will require multi-level and longitudinal biomarker evaluation over a large dose range, multi-tissue biomarker evaluation, and the generation of a proof of principle by application to multiple CNS drugs in multiple species.

AbstractZebrafish larvae (Danio rerio) are increasingly used to translate findings regarding drug efficacy and safety from in vitro-based assays to vertebrate species, including humans. However, the limited understanding of drug exposure in this species hampers its implementation in translational research. Using paracetamol as a paradigm compound, we present a novel method to characterize pharmacokinetic processes in zebrafish larvae, by combining sensitive bioanalytical methods and nonlinear mixed effects modeling. The developed method allowed quantification of paracetamol and its two major metabolites, paracetamol-sulfate and paracetamol-glucuronide in pooled samples of five lysed zebrafish larvae of 3 days post-fertilization. Paracetamol drug uptake was quantified to be 0.289 pmole/min and paracetamol clearance was quantified to be 1.7% of the total value of the larvae. With an average volume determined to be 0.290 μL, this yields an absolute clearance of 2.96 × 107L/h, which scales reasonably well with clearance rates in higher vertebrates. The developed methodology will improve the success rate of drug screens in zebrafish larvae and the translation potential of findings, by allowing the establishment of accurate exposure profiles and thereby also the establishment of concentration–effect relationships.

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 vitrodrug-target binding kinetics in drug discovery requires insight into the relation between in vivodrug effect and in vitromeasured drug-target binding kinetics.Areas covered:In this review, the authors discuss both the relation between in vitroand in vivomeasured binding kinetics and the relation between in vivobinding 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 vitroestimates of drug-target binding kinetics. To elucidate the value of in vitrobinding 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.

Intranasal (IN) administration could be an attractive mode of delivery for drugs targeting the central nervous system, potentially providing a high bioavailability because of avoidance of a hepatic first-pass effect and rapid onset of action. However, controversy remains whether a direct transport route from the nasal cavity into the brain exists. Pharmacokinetic modeling is proposed to identify the existence of direct nose-to-brain transport in a quantitative manner. The selective dopamine-D2 receptor antagonist remoxipride was administered at different dosages, in freely moving rats, by the IN and intravenous (IV) route. Plasma and brain extracellular fluid (ECF) concentration-time profiles were obtained and simultaneously analyzed using nonlinear mixed-effects modeling. Brain ECF/plasma area under the curve ratios were 0.28 and 0.19 after IN and IV administration, respectively. A multicompartment pharmacokinetic model with two absorption compartments (nose-to-systemic and nose-to-brain) was found to best describe the observed pharmacokinetic data. Absorption was described in terms of bioavailability and rate. Total bioavailability after IN administration was 89%, of which 75% was attributed to direct nose-to brain transport. Direct nose-to-brain absorption rate was slow, explaining prolonged brain ECF exposure after IN compared with IV administration. These studies explicitly provide separation and quantitation of systemic and direct nose-to-brain transport after IN administration of remoxipride in the rat. Describing remoxipride pharmacokinetics at the target site (brain ECF) in a semiphysiology-based manner would allow for better prediction of pharmacodynamic effects.

Intranasal (IN) administration could be an attractive mode of delivery for drugs targeting the central nervous system, potentially providing a high bioavailability because of avoidance of a hepatic first-pass effect and rapid onset of action. However, controversy remains whether a direct transport route from the nasal cavity into the brain exists. Pharmacokinetic modeling is proposed to identify the existence of direct nose-to-brain transport in a quantitative manner. The selective dopamine-D2 receptor antagonist remoxipride was administered at different dosages, in freely moving rats, by the IN and intravenous (IV) route. Plasma and brain extracellular fluid (ECF) concentration-time profiles were obtained and simultaneously analyzed using nonlinear mixed-effects modeling. Brain ECF/plasma area under the curve ratios were 0.28 and 0.19 after IN and IV administration, respectively. A multicompartment pharmacokinetic model with two absorption compartments (nose-to-systemic and nose-to-brain) was found to best describe the observed pharmacokinetic data. Absorption was described in terms of bioavailability and rate. Total bioavailability after IN administration was 89%, of which 75% was attributed to direct nose-to brain transport. Direct nose-to-brain absorption rate was slow, explaining prolonged brain ECF exposure after IN compared with IV administration. These studies explicitly provide separation and quantitation of systemic and direct nose-to-brain transport after IN administration of remoxipride in the rat. Describing remoxipride pharmacokinetics at the target site (brain ECF) in a semiphysiology-based manner would allow for better prediction of pharmacodynamic effects.

ABSTRACTRecombinant alpha interferon (IFN-α) is used in the treatment of hepatitis C virus (HCV)-infected patients but is not optimal in terms of efficacy or tolerability. Toll-like 7 receptor (TLR-7) agonists stimulate the innate immune system to produce, among other cytokines, IFN-α and are being evaluated as alternative drugs to treat HCV infection. This paper describes the application of pharmacokinetic-pharmacodynamic (PK-PD) modeling to understanding the behavior of a TLR-7 agonist [9-benzyl-8-hydroxy-2-(2-methoxyethoxy) adenine (BHMA)] in mice, using IFN-α as a biomarker. This is the first report of such a PK-PD model, and the conclusions may be of utility in the clinical development of TLR-7 agonists for HCV infection.

Pharmacokinetic-Pharmacodynamic (PK-PD) modeling helps to better understand drug efficacy and safety and has, therefore, become a powerful tool in the learning-confirming cycles of drug-development. In translational drug research, mechanism-based PK-PD modeling has been recognized as a tool for bringing forward early insights in drug efficacy and safety into the clinical development. These models differ from descriptive PK-PD models in that they quantitatively characterize specific processes in the causal chain between drug administration and effect. This includes target site distribution, binding and activation, pharmacodynamic interactions, transduction and homeostatic feedback mechanisms. Compared to descriptive models mechanism-based PK-PD models that utilize receptor theory concepts for characterization of target binding and target activation processes have improved properties for extrapolation and prediction. In this respect, receptor theory constitutes the basis for 1) prediction of in vivodrug concentration-effect relationships and 2) characterization of target association-dissociation kinetics as determinants of hysteresis in the time course of the drug effect. This approach intrinsically distinguishes drug- and system specific parameters explicitly, allowing accurate extrapolation from in vitroto in vivoand across species. This review provides an overview of recent developments in incorporating receptor theory in PK-PD modeling with a specific focus on the identifiability of these models.

ABSTRACT

The objective of this investigation was to compare the in vivo potency and intrinsic activity of buspirone and its metabolite 1-(2-pyrimidinyl)-piperazine (1-PP) in rats by pharmacokinetic-pharmacodynamic modeling. Following intravenous administration of buspirone (5 or 15 mg/kg in 15 min) or 1-PP (10 mg/kg in 15 min), the time course of the concentrations in blood were determined in conjunction with the effect on body temperature. The pharmacokinetics of buspirone and 1-PP were analyzed based on a two-compartment model with metabolite formation. Differences in the pharmacokinetics of buspirone and 1-PP were observed with values for clearance of 13.1 and 8.2 ml/min and for terminal elimination half-life of 25 and 79 min, respectively. At least 26% of the administered dose of buspirone was converted into 1-PP. Complex hypothermic effects versus time profiles were observed, which were successfully analyzed on the basis of a physiological indirect response model with set-point control. Both buspirone and 1-PP behaved as partial agonists relative to R-(+)-8-hydroxy-2-(di-n-propylamino)tetralin (R-8-OH-DPAT) with values of the intrinsic activity of 0.465 and 0.312, respectively. Differences in the potency were observed with values of 17.6 and 304 ng/ml for buspirone and 1-PP, respectively. The results of this analysis show that buspirone and 1-PP behave as partial 5-hydroxytryptamine1Aagonists in vivo and that following intravenous administration the amount of 1-PP formed is too small to contribute to the hypothermic effect.

Background and Aims:Replacement FIX therapy (rIX) is an effective treatment for hemophilia B even with undetectable levels in the blood 1. However, the mechanistic reason for hemostasis with low plasma levels is not well understood. There is growing evidence that FIX interactions with one or multiple binding partners (BP), may play a significant role in the exposure and hemostatic efficacy of rIX 2,3. The aim of this study is to explore this hypothesis by comparing the plasma PK, tissue biodistribution, and in vivo endpoints of different rIX variants using a mouse QSP model.

Pestel: CSL Behring Innovation GmbH: Current Employment, Current equity holder in publicly-traded company. Rezvani-Sharif: CSL Behring Ltd: Current Employment, Current equity holder in publicly-traded company. Muir: CSL Behring Ltd: Current Employment, Current holder of stock options in a privately-held company. Krupa: CSL Behring LLC: Current Employment, Current equity holder in publicly-traded company. Brechmann: CSL Behring Innovation GmbH, Ended employment in the past 24 months: Bayer Ag (Bayer Pharmaceuticals),: Current Employment, Ended employment in the past 24 months, Patents & Royalties: Bayer. Verhagen: CSL Behring Ltd: Current Employment, Current equity holder in publicly-traded company. Dower: CSL Behring Ltd: Current Employment, Current equity holder in publicly-traded company. Herzog: CSL Behring GmbH: Current Employment, Current equity holder in publicly-traded company.

The aim of this study was to investigate the feasibility of a quantitative structure–pharmacokinetic relationships (QSPKR) method based on contemporary three-dimensional (3D) molecular characterization and multivariate statistical analysis. For this purpose, the programs SYBYL/CoMFA, GRID, and Pallas, in combination with the multivariate statistical technique principal component analysis were employed to generate a total of 16 descriptor variables for a series of 12 structurally related adenosine A1receptor agonists. Subsequently, the multivariate regression method, partial least squares, was used to predict clearance (CL), volume of distribution (VdSS) and protein binding (fraction unbound, fU). The QSPKR models obtained could account for most of the variation in CL, VdSS, and fU(R2= 0.82, 0.61 and 0.78, respectively). Cross-validation confirmed the predictive ability of the models (Q2= 0.59, 0.41 and 0.62 for CL, VdSS, and fU, respectively). In conclusion, we have developed a multivariate 3D QSPKR model that could adequately predict overall pharmacokinetic behavior of adenosine A1receptor agonists in rat. This methodology can also be used for other classes of compounds and may facilitate the further integration of QSPKR in drug discovery and preclinical development.

To illuminate the controversy on α1A‐ or α1L‐adrenoceptor involvement in noradrenaline‐mediated contractions of rat small mesenteric artery (SMA), we have studied the effects of subtype‐selective α1‐adrenoceptor agonists and antagonists under different experimental conditions.The agonist potency order in rat SMA was: A61603 >> SKF89748‐A > cirazoline > noradrenaline > ST‐587 > methoxamine. Prazosin antagonized all agonists with a low potency (pA2: 8.29–8.80) indicating the involvement of α1L‐ rather than α1A‐adrenoceptors.The putative α1L‐adrenoceptor antagonist JTH‐601, but not the α1B‐adrenoceptor antagonist chloroethylclonidine (10 μM) antagonized noradrenaline‐induced contractions of SMA. The potency of the selective α1D‐adrenoceptor antagonist BMY 7378 against noradrenaline (pA2=6.16±0.13) and of the selective α1A‐adrenoceptor antagonist RS‐17053 against noradrenaline (pKB=8.35±0.10) and against the selective α1A‐adrenoceptor agonist A‐61603 (pKB=8.40±0.09) were too low to account for α1D‐ and α1A‐adrenoceptor involvement.The potency of RS‐17053 (pKB/pA2's=7.72–8.46) was not affected by lowering temperature, changing experimental protocol or inducing myogenic tone viaKCl or U46619.Selective protection of a putative α1A‐adrenoceptor population against the irreversible action of phenoxybenzamine also failed to increase the potency of RS‐17053 (pA2=8.25±0.06 against A61603).Combined concentration‐ratio analysis demonstrated that tamsulosin, which does not discriminate between α1A‐ and α1L‐adrenoceptors, and RS‐17053 competed for binding at the same site in the SMA.In summary, data obtained in our experiments in rat SMA indicate that the α1‐adrenoceptor mediating noradrenaline‐induced contraction displays a distinct α1L‐adrenoceptor pharmacology. This study does not provide evidence for the hypothesis that α1L‐adrenoceptors represent an affinity state of the α1A‐adrenoceptor in functional assays. Furthermore, there is no co‐existing α1A‐adrenoceptor in the SMA.

The purpose of our investigation was to characterize the relationships between the pharmacodynamics of synthetic opioids in vivoand the interaction at the mu-opioid receptor. The pharmacokinetics and pharmacodynamics were determined in vivoafter 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 Emaxpharmacodynamic model. The interaction at the mu-opioid receptor was determined in vitroon 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 vitrointrinsic efficacy. For the EEG effect the in vivopotencies 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 (KI) 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,uand KIwas 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 vivowas similar with values of 111 ± 10 μV (A), 89 ± 11 μV (F) and 104 ± 4 μV (S). However, the values of the sodium shift varied between 2.8 (S) and 19.1 (A). Further analysis of the in vivopharmacodynamic 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.

The nerve growth factor (NGF) pathway is of great interest as a potential source of drug targets, for example in the management of certain types of pain. However, selecting targets from this pathway either by intuition or by non-contextual measures is likely to be challenging. An alternative approach is to construct a mathematical model of the system and via sensitivity analysis rank order the targets in the known pathway, with respect to an endpoint such as the diphosphorylated extracellular signal-regulated kinase concentration in the nucleus. Using the published literature, a model was created and, via sensitivity analysis, it was concluded that, after NGF itself, tropomyosin receptor kinase A (TrkA) was one of the most sensitive druggable targets. This initial model was subsequently used to develop a further model incorporating physiological and pharmacological parameters. This allowed the exploration of the characteristics required for a successful hypothetical TrkA inhibitor. Using these systems models, we were able to identify candidates for the optimal drug targets in the known pathway. These conclusions were consistent with clinical and human genetic data. We also found that incorporating appropriate physiological context was essential to drawing accurate conclusions about important parameters such as the drug dose required to give pathway inhibition. Furthermore, the importance of the concentration of key reactants such as TrkA kinase means that appropriate contextual data are required before clear conclusions can be drawn. Such models could be of great utility in selecting optimal targets and in the clinical evaluation of novel drugs.

ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.