期刊
MOLECULAR PHARMACEUTICS
卷 10, 期 6, 页码 2467-2478出版社
AMER CHEMICAL SOC
DOI: 10.1021/mp4000822
关键词
intracellular unbound concentrations; drug binding; drug transport; drug accumulation; membrane partitioning
资金
- Science for Life Laboratory (SciLifeLab)
- Swedish Fund for Research Without Animal Experiments
- Magn. Bergvalls Stiftelse
- Swedish Research Council [9478, 21386]
- Fundacao para a Ciencia e Tecnologia [SFRH/BD/68304/2010]
- Fundação para a Ciência e a Tecnologia [SFRH/BD/68304/2010] Funding Source: FCT
Intracellular unbound drug concentrations determine affinity to targets in the cell interior. However, due to difficulties in measuring them, they are often overlooked in pharmacology. Here we present a simple experimental technique for the determination of unbound intracellular drug concentrations in cultured cells that is based on parallel measurements of cellular drug binding and steady-state intracellular drug concentrations. Binding in HEK293 cells was highly correlated with binding in liver-derived systems, whereas binding in plasma did not compare well with cellular binding. Compound lipophilicity increased drug binding, while negative charge and aromatic functional groups decreased binding. Intracellular accumulation of unbound drug was consistent with pH dependent subcellular sequestration, as confirmed by modeling and by inhibition of subcellular pH gradients. The approach developed here can be used to measure intracellular unbound drug concentrations in more complex systems, for example, cell lines with controlled expression of transporters and enzymes or primary cells.
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