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Human Plasma In‐Cell Western Assays—An In vitro Predictor for In vivo Pharmacology in Oncology Drug Discovery
Author(s) -
Zhang Yang W.,
Gallagher Kirsten,
Angelis Dimitrios,
Rominger Dave,
Scherle Peggy,
Vaddi Kris
Publication year - 2021
Publication title -
current protocols
Language(s) - English
Resource type - Journals
ISSN - 2691-1299
DOI - 10.1002/cpz1.51
Subject(s) - in vivo , potency , drug discovery , pharmacology , pharmacodynamics , in vitro , pharmacokinetics , ic50 , drug , plasma protein binding , computational biology , in vitro toxicology , blood proteins , human plasma , drug development , biological activity , chemistry , biology , biochemistry , microbiology and biotechnology , chromatography
Evaluation of in vivo potencies plays an important role in drug discovery. Traditionally, the cellular activity and percent of plasma protein binding of a test agent are evaluated separately, with the plasma protein binding−adjusted cellular potency computation used to estimate in vivo potency. This process is costly, takes weeks to complete, and is increasingly unreliable for compounds that bind extensively to plasma proteins. Described in this article is a simple, high‐throughput human plasma in‐cell Western (ICW) assay that directly incorporates plasma protein binding into a cellular pharmacodynamic assay to provide a rapid and accurate estimate of in vivo potencies. The assay is versatile and can be readily employed for various targets that require short treatment periods for displaying maximal biological responses. © 2021 Wiley Periodicals LLC. This article was corrected on 19 July 2022. See the end of the full text for details. Basic Protocol : Concentration‐dependent human plasma ICW assay to determine test compound IC 50 against the target of interest