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Research ArticleNeuropharmacology

Calculation of an Apical Efflux Ratio from P-Glycoprotein (P-gp) In Vitro Transport Experiments Shows an Improved Correlation with In Vivo Cerebrospinal Fluid Measurements in Rats: Impact on P-gp Screening and Compound Optimization

Holger Fischer, Claudia Senn, Mohammed Ullah, Carina Cantrill, Franz Schuler and Li Yu
Journal of Pharmacology and Experimental Therapeutics March 2021, 376 (3) 322-329; DOI: https://doi.org/10.1124/jpet.120.000158
Holger Fischer
Roche Pharmaceutical Research and Early Development, DMPK/PD project leader (H.F.), Comparative Pharmacology (C.S.), Investigative Safety, Pharmaceutical Sciences (M.U., C.C.), and Immunology, Infectious Disease and Ophthalmology (F.S.), Roche Innovation Center, Basel, Switzerland; and Roche Pharmaceutical Research and Early Development, Pharmaceutical Sciences, Roche Translational and Clinical Research Center, Inc., and LIYU Pharmaceutical Consulting LCC, New Jersey, USA (L.Y.)
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Claudia Senn
Roche Pharmaceutical Research and Early Development, DMPK/PD project leader (H.F.), Comparative Pharmacology (C.S.), Investigative Safety, Pharmaceutical Sciences (M.U., C.C.), and Immunology, Infectious Disease and Ophthalmology (F.S.), Roche Innovation Center, Basel, Switzerland; and Roche Pharmaceutical Research and Early Development, Pharmaceutical Sciences, Roche Translational and Clinical Research Center, Inc., and LIYU Pharmaceutical Consulting LCC, New Jersey, USA (L.Y.)
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Mohammed Ullah
Roche Pharmaceutical Research and Early Development, DMPK/PD project leader (H.F.), Comparative Pharmacology (C.S.), Investigative Safety, Pharmaceutical Sciences (M.U., C.C.), and Immunology, Infectious Disease and Ophthalmology (F.S.), Roche Innovation Center, Basel, Switzerland; and Roche Pharmaceutical Research and Early Development, Pharmaceutical Sciences, Roche Translational and Clinical Research Center, Inc., and LIYU Pharmaceutical Consulting LCC, New Jersey, USA (L.Y.)
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Carina Cantrill
Roche Pharmaceutical Research and Early Development, DMPK/PD project leader (H.F.), Comparative Pharmacology (C.S.), Investigative Safety, Pharmaceutical Sciences (M.U., C.C.), and Immunology, Infectious Disease and Ophthalmology (F.S.), Roche Innovation Center, Basel, Switzerland; and Roche Pharmaceutical Research and Early Development, Pharmaceutical Sciences, Roche Translational and Clinical Research Center, Inc., and LIYU Pharmaceutical Consulting LCC, New Jersey, USA (L.Y.)
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Franz Schuler
Roche Pharmaceutical Research and Early Development, DMPK/PD project leader (H.F.), Comparative Pharmacology (C.S.), Investigative Safety, Pharmaceutical Sciences (M.U., C.C.), and Immunology, Infectious Disease and Ophthalmology (F.S.), Roche Innovation Center, Basel, Switzerland; and Roche Pharmaceutical Research and Early Development, Pharmaceutical Sciences, Roche Translational and Clinical Research Center, Inc., and LIYU Pharmaceutical Consulting LCC, New Jersey, USA (L.Y.)
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Li Yu
Roche Pharmaceutical Research and Early Development, DMPK/PD project leader (H.F.), Comparative Pharmacology (C.S.), Investigative Safety, Pharmaceutical Sciences (M.U., C.C.), and Immunology, Infectious Disease and Ophthalmology (F.S.), Roche Innovation Center, Basel, Switzerland; and Roche Pharmaceutical Research and Early Development, Pharmaceutical Sciences, Roche Translational and Clinical Research Center, Inc., and LIYU Pharmaceutical Consulting LCC, New Jersey, USA (L.Y.)
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Abstract

P-glycoprotein (P-gp) is a major blood-brain barrier (BBB) efflux transporter. In vitro approaches, including bidirectional efflux ratio (ER), are used to measure P-gp–mediated transport, but findings can be inconsistent across models. We propose a novel, more physiologically relevant, in vitro model: unidirectional apical efflux ratio (AP-ER)—a ratio of permeability rates at the apical side of the BBB with and without P-gp inhibitor. To test our approach, ER and AP-ER were calculated for 3227 structurally diverse compounds in porcine kidney epithelial cells (LLC-PK1) overexpressing human or mouse P-gp and classified based on their passive transcellular P-gp permeability or charged properties. In vivo rat infusion studies were performed for selected compounds with high ER but low AP-ER. One-third of the 3227 compounds had bidirectional ER that was much higher than AP-ER; very few had AP-ER higher than ER. Compounds with a large difference between AP-ER and ER were typically basic compounds with low-to-medium passive permeability and high lipophilicity and/or amphiphilicity, leading to strong membrane binding. Outcomes in the human model were similar to those in mice, suggesting AP-ER/ER ratios may be conserved for at least two species. AP-ER predicted measured cerebrospinal fluid (CSF) concentration better than ER for the five compounds tested in our in vivo rat infusion studies. We report superior estimations of the CSF concentrations of the compounds when based on less resource-intensive AP-ER versus classic ER. Better understanding of the properties leading to high P-gp–mediated efflux in vivo could support more efficient brain-penetrant compound screening and optimization.

SIGNIFICANCE STATEMENT To address inconsistencies associated with the historical, bidirectional efflux ratio (ER) calculation of P-glycoprotein–mediated transport, we propose to use the novel, more physiologically relevant, unidirectional apical efflux ratio (AP-ER) model. In vitro experiments suggested that compounds with strong membrane binding showed the largest difference between AP-ER and ER, and in vivo infusion studies showed that AP-ER predicted cerebrospinal fluid concentrations of compounds better than ER; outcomes in the human model were similar to those in mice.

Footnotes

    • Received June 8, 2020.
    • Accepted December 2, 2020.
  • This study was supported by F. Hoffmann-La Roche Ltd./Genentech Inc.

  • Primary laboratory of origin: Roche Pharmaceutical Research and Early Development, Roche Innovation Center (Basel, Switzerland).

  • https://doi.org/10.1124/jpet.120.000158.

  • ↵Embedded ImageThis article has supplemental material available at jpet.aspetjournals.org.

  • Copyright © 2021 by The American Society for Pharmacology and Experimental Therapeutics
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Journal of Pharmacology and Experimental Therapeutics: 376 (3)
Journal of Pharmacology and Experimental Therapeutics
Vol. 376, Issue 3
1 Mar 2021
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Research ArticleNeuropharmacology

P-Glycoprotein Apical Efflux Ratio for Compound Optimization

Holger Fischer, Claudia Senn, Mohammed Ullah, Carina Cantrill, Franz Schuler and Li Yu
Journal of Pharmacology and Experimental Therapeutics March 1, 2021, 376 (3) 322-329; DOI: https://doi.org/10.1124/jpet.120.000158

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Research ArticleNeuropharmacology

P-Glycoprotein Apical Efflux Ratio for Compound Optimization

Holger Fischer, Claudia Senn, Mohammed Ullah, Carina Cantrill, Franz Schuler and Li Yu
Journal of Pharmacology and Experimental Therapeutics March 1, 2021, 376 (3) 322-329; DOI: https://doi.org/10.1124/jpet.120.000158
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