Application of a new MDCKII-MDR1 cell model to measure the extent of drug distribution in vitro at equilibrium for prediction of in vivo unbound brain-to-plasma drug distribution

Plasma pharmacokinetic (PK) profiles often do not resemble the PK within the central nervous system (CNS) because of blood-brain-border (BBB) processes, like active efflux by P-glycoprotein (P-gp). Methods to predict CNS-PK are therefore desired. Here we investigate whether in vitro apparent permeability (Papp) and corrected efflux ratio (ERc) extracted from literature can be repurposed as input for the LeiCNS-PK3.4 physiologically-based PK model to confidently predict rat brain extracellular fluid (ECF) PK of P-gp substrates. Literature values of in vitro Caco-2, LLC-PK1-mdr1a/MDR1, and MDCKII-MDR1 cell line transport data were used to calculate P-gp efflux clearance (CLPgp). Subsequently, CLPgp was scaled from in vitro to in vivo through a relative expression factor (REF) based on P-gp expression differences. BrainECF PK was predicted well (within 2-fold error of the observed data) for 2 out of 4 P-gp substrates after short infusions and 3 out of 4 P-gp substrates after continuous infusions. Prediction accuracy was influenced by variability in reported ERc and in vitro P-gp expression. Notably, use of Papp and in vitro P-gp expression obtained from a single study did not guarantee an accurate prediction; it often resulted in worse predictions than when using in vitro expression values reported by other labs. Using Papp and P-gp expression as input, LeiCNS-PK3.4 shows promise in predicting brainECF PK but this study highlights that the in vitro to in vivo translation is not yet robust. We conclude that more information is needed about context and drug dependency of in vitro data for robust brainECF PK predictions.

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In vitro, in vivo, and in silico approaches for evaluating the preclinical DMPK profiles of ammoxetine, a novel chiral serotonin and norepinephrine reuptake inhibitor

Zhu,

Li,

Luo

et al. 2024

Front. Pharmacol.

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Background and AimAmmoxetine, a novel chiral serotonin and norepinephrine reuptake inhibitor, holds promise for major depressive disorder treatment. This study aimed to thoroughly investigate its preclinical drug metabolism and pharmacokinetics (DMPK) profiles.MethodsThe preclinical DMPK profiles of ammoxetine were examined through in vitro, in vivo, and in silico methods.ResultsAssessment of blood-brain barrier penetration via MDCK-MDR1 cells revealed strong brain permeation by ammoxetine, despite being a probable P-glycoprotein (P-gp) substrate. Molecular docking indicated a robust binding interaction between ammoxetine and P-gp. Ammoxetine was well absorbed orally, with Tmax ranging from 0.75 to 3.83 h in rats and 0.75–1.40 h in beagle dogs. At a 2 mg/kg dose in beagle dogs, ammoxetine exhibited an absolute bioavailability of approximately 42%. Plasma protein binding rates were around 50%–60% in beagle dogs, rats, and humans, suggesting moderate binding. Tissue distribution studies displayed rapid and extensive ammoxetine spread in major rat tissues post-gavage, with notable brain exposure and no tissue accumulation. Cumulative excretion rates in rats’ urine, feces, and bile accounted for only 1.11% of the total administered drug, indicating extensive transformation into metabolites. Chiral inversion of ammoxetine was absent in vivo. Metabolic stability varied across species using liver microsomes, but beagle dogs showed clearance rates more akin to humans. Metabolic pathways unveiled two key metabolites, M1 and M2. M1, likely generated through methylenedioxyphenyl ring oxidation, involves CYP2C19 and CYP3A4, crucial human cytochrome P450 (CYP) enzymes for liver metabolism, while M2 is M1’s glucuronide conjugate. Ammoxetine may exhibit saturation elimination trends with increasing doses in rats and beagle dogs. A high-throughput assay using the cocktail-substrate method indicated weak CYP inhibition by ammoxetine on CYP2D6 and CYP1A2, with minimal effects on other CYP enzymes, suggesting a low likelihood of CYP inhibition-related drug-drug interactions.ConclusionThis study presents encouraging DMPK profiles of ammoxetine, backing its potential as a candidate compound for future clinical assessments.

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Application of a new MDCKII-MDR1 cell model to measure the extent of drug distribution in vitro at equilibrium for prediction of in vivo unbound brain-to-plasma drug distribution

Cited by 3 publications

References 36 publications

Use of Unbound Exposure Data to Establish In Vitro–In Vivo Correlations for CNS Drug Candidates

Use of Unbound Exposure Data to Establish In Vitro–In Vivo Correlations for CNS Drug Candidates

Reliability of in vitro data for the mechanistic prediction of brain extracellular fluid pharmacokinetics of P-glycoprotein substrates in vivo; are we scaling correctly?

In vitro, in vivo, and in silico approaches for evaluating the preclinical DMPK profiles of ammoxetine, a novel chiral serotonin and norepinephrine reuptake inhibitor

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