Article abstract
Nature Chemical Biology 4, 132 - 142 (2007)
Published online: 23 December 2007 | doi:10.1038/nchembio.2007.59
High-content single-cell drug screening with phosphospecific flow cytometry
Peter O Krutzik1, Janelle M Crane1, Matthew R Clutter1 & Garry P Nolan1
Abstract
Drug screening is often limited to cell-free assays involving purified enzymes, but it is arguably best applied against systems that represent disease states or complex physiological cellular networks. Here, we describe a high-content, cell-based drug discovery platform based on phosphospecific flow cytometry, or phosphoflow, that enabled screening for inhibitors against multiple endogenous kinase signaling pathways in heterogeneous primary cell populations at the single-cell level. From a library of small-molecule natural products, we identified pathway-selective inhibitors of Jak-Stat and MAP kinase signaling. Dose-response experiments in primary cells confirmed pathway selectivity, but importantly also revealed differential inhibition of cell types and new druggability trends across multiple compounds. Lead compound selectivity was confirmed in vivo in mice. Phosphoflow therefore provides a unique platform that can be applied throughout the drug discovery process, from early compound screening to in vivo testing and clinical monitoring of drug efficacy.
- Department of Microbiology and Immunology, Baxter Laboratory in Genetic Pharmacology, Stanford University, 269 Campus Drive, Stanford, California 94305, USA.
Correspondence to: Garry P Nolan1 e-mail: gnolan@stanford.edu
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