CellPort Technologies®

Transporters

We selected a human cell line (Caco-2, C2BBE1 clone) expressing the efflux transporters P-gp, BCRP, and MRP2, and have knocked down the expression of one transporter at a time. Knockdown cell lines can be evaluated in different ways. For example, as a standalone model the P-gp knockdown is a cleaner BCRP assay system. When used in combination with control cells, it can identify P-gp substrates. Our knockdown was achieved using short hairpin RNA (shRNA) in a lentiviral vector, an approach that is capable of producing stable phenotypes.

Our cell lines for evaluation of hepatic and renal uptake transporters were created by stable transfection of transporter genes into HEK293 cells. For initial set-up, stable transfections are more laborious and challening than transient transfections, but they result in permanent insertion into the genome. This allows us to fully characterize the phenotypes of our cells.

All Absorption Systems cell lines have been extensively characterized and validated:

Molecular Biology for Phenotypic Expression
     • Quantitative PCR for mRNA expression
     • Western blot for protein expression
     • Immunofluorescence for protein localization
Assay Window and Phenotype Stability
    • Growth rate
    • Passage number
    • Days in culture
    • Assay window
Appropriate and Robust Functional Expression
    • Uptake and/or bidirectional transport  
    • Polarization and barrier properties (cell lines in bidirectional transport format)

Our CellPort Analytics™ database captures up to ten years of data for our cell lines. This tool helps us ensure our cell lines are behaving optimally, according to stringent criteria we’ve established based on extensive knowledge and experience.

Transporter Reference Guide Ebook
Do you have questions about recently issued regulatory guidance from the FDA and EMA? Do you need to know about potential drug interactions to watch for your candidate? Discover clarity in transporters with the exclusive Transporter Reference Guide from Absorption Systems.
View ebook

Transporters Overview
Researchers studying transporters continue to grapple with a lack of definitive tools. Absorption Systems has over a decade of experience with assessing transporter interactions and has extensive experience with regulatory agencies across the world.

Regulatory Perspective
The new FDA Draft Guidance (2012), EMA Guidelines (2010), and ITC White Paper (2010) all provide various recommendations for the evaluation of transporters.

Dose Selection
Concentrations selected for substrate assessment should be based on the total plasma Cmax of the test compound to ensure that results are clinically relevant.

Waiving Clinical Studies
Based on drug-specific properties (solubility, permeability, metabolism), it may be possible to waive clinical DDI studies, even if your compound is a transporter substrate.

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