Identification and validation of protein targets of bioactive small molecules

Department of Pharmacology, Johns Hopkins University School of Medicine, MD, USA.
Bioorganic & medicinal chemistry (Impact Factor: 2.79). 03/2012; 20(6):1902-9. DOI: 10.1016/j.bmc.2011.11.070
Source: PubMed


Identification and validation of protein targets of bioactive small molecules is an important problem in chemical biology and drug discovery. Currently, no single method is satisfactory for this task. Here, we provide an overview of common methods for target identification and validation that historically were most successful. We have classified for the first time the existing methods into two distinct and complementary types, the 'top-down' and 'bottom-up' approaches. In a typical top-down approach, the cellular phenotype is used as a starting point and the molecular target is approached through systematic narrowing down of possibilities by taking advantage of the detailed existing knowledge of cellular pathways and processes. In contrast, the bottom-up approach entails the direct detection and identification of the molecular targets using affinity-based or genetic methods. A special emphasis is placed on target validation, including correlation analysis and genetic methods, as this area is often ignored despite its importance.

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    • "Consequently, this approach has the potential to work in model systems that do not permit forward genetics and, unlike reverse genetics, does not require any starting information about likely players in the process being studied. The macromolecular target of any small molecule identified from a library can be identified by biochemical approaches [e.g., Lomenick et al., 2011; Titov and Liu, 2012]. Even prior to such identification , specific inhibitors can be extremely useful with respect to ordering hierarchies of cell biological events. "
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    ABSTRACT: Screening of small molecule libraries offers the potential to identify compounds that inhibit specific biological processes and, ultimately, to identify macromolecules that are important players in such processes. To date, however, most screens of small molecule libraries have focused on identification of compounds that inhibit known proteins or particular steps in a given process, and have emphasized automated primary screens. Here we have used "low tech" in vivo primary screens to identify small molecules that inhibit both cytokinesis and single cell wound repair, two complex cellular processes that possess many common features. The "diversity set", an ordered array of 1990 compounds available from the National Cancer Institute, was screened in parallel to identify compounds that inhibit cytokinesis in Dendraster excentricus (sand dollar) embryos and single cell wound repair in Xenopus laevis (frog) oocytes. Two small molecules were thus identified: Sph1 and Sph2. Sph1 reduces Rho activation in wound repair and suppresses formation of the spindle midzone during cytokinesis. Sph2 also reduces Rho activation in wound repair and may inhibit cytokinesis by blocking membrane fusion. The results identify two small molecules of interest for analysis of wound repair and cytokinesis, reveal that these processes are more similar than often realized and reveal the potential power of low tech screens of small molecule libraries for analysis of complex cellular processe. © 2012 Wiley Periodicals, Inc.
    Full-text · Article · Nov 2012 · Cytoskeleton
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