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A CRISPR-Cas9 Generated MDCK Cell Line Expressing Human MDR1 Without Endogenous Canine MDR1 (cABCB1): An Improved Tool for Drug Efflux Studies.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmacy. Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University Drug Optimization and Pharmaceutical Profiling Platform, Science for Life Laboratory.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmacy.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmacy. Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University Drug Optimization and Pharmaceutical Profiling Platform, Science for Life Laboratory.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmacy. AstraZeneca.
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2017 (English)In: Journal of Pharmaceutical Sciences, ISSN 0022-3549, E-ISSN 1520-6017, Vol. 106, no 9, p. 2909-2913Article in journal (Refereed) Published
Abstract [en]

Madin-Darby canine kidney (MDCK) II cells stably transfected with transport proteins are commonly used models for drug transport studies. However, endogenous expression of especially canine MDR1 (cMDR1) confounds the interpretation of such studies. Here we have established an MDCK cell line stably overexpressing the human MDR1 transporter (hMDR1; P-glycoprotein), and used CRISPR-Cas9 gene editing to knockout the endogenous cMDR1. Genomic screening revealed the generation of a clonal cell line homozygous for a 4-nucleotide deletion in the canine ABCB1 gene leading to a frameshift and a premature stop codon. Knockout of cMDR1 expression was verified by quantitative protein analysis and functional studies showing retained activity of the human MDR1 transporter. Application of this cell line allowed unbiased reclassification of drugs previously defined as both substrates and non-substrates in different studies using commonly used MDCK-MDR1 clones. Our new MDCK-hMDR1 cell line, together with a previously developed control cell line, both with identical deletions in the canine ABCB1 gene and lack of cMDR1 expression represent excellent in vitro tools for use in drug discovery.

Place, publisher, year, edition, pages
2017. Vol. 106, no 9, p. 2909-2913
Keyword [en]
ABC transporters, MDCK cells, P-glycoprotein, drug transport, efflux pumps, genomics, membrane transporter, multidrug resistance transporters, permeability, proteomics
National Category
Medical and Health Sciences
Identifiers
URN: urn:nbn:se:uu:diva-343598DOI: 10.1016/j.xphs.2017.04.018ISI: 000417339900089PubMedID: 28450237OAI: oai:DiVA.org:uu-343598DiVA, id: diva2:1186366
Funder
Science for Life Laboratory - a national resource center for high-throughput molecular bioscience
Available from: 2018-02-28 Created: 2018-02-28 Last updated: 2018-03-07Bibliographically approved

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Karlgren, MariaSimoff, IvailoWegler, ChristineHandin, NiklasLundquist, PatrikArtursson, Per

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Karlgren, MariaSimoff, IvailoWegler, ChristineHandin, NiklasLundquist, PatrikArtursson, Per
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Department of PharmacyScience for Life Laboratory, SciLifeLabDepartment of Immunology, Genetics and Pathology
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