Small-Molecule Fluorophores To Detect Cell-State Switching in the Context of High-Throughput Screening

• Published in: Journal of the American Chemical Society Vol. 130; no. 13; pp. 4208 - 4209
• Main Authors: Wagner, Bridget K, Carrinski, Hyman A, Ahn, Young-Hoon, Kim, Yun Kyung, Gilbert, Tamara J, Fomina, Dina A, Schreiber, Stuart L, Chang, Young-Tae, Clemons, Paul A
• Format: Journal Article
• Language: English
• Published: United States American Chemical Society 02-04-2008
• Subjects: Animals; Cell Differentiation - physiology; Cell Line; Drug Evaluation, Preclinical; Enzyme Activation - drug effects; Enzyme Inhibitors - chemistry; Enzyme Inhibitors - metabolism; Enzyme Inhibitors - pharmacokinetics; Fluorescent Dyes - chemistry; Fluorescent Dyes - metabolism; Fluorescent Dyes - pharmacokinetics; Mice; Microscopy, Fluorescence; Molecular Probes - chemistry; Molecular Probes - metabolism; Molecular Probes - pharmacokinetics; Molecular Weight; Muscle Fibers, Skeletal - cytology; Muscle Fibers, Skeletal - drug effects; Muscle Fibers, Skeletal - metabolism; Myoblasts - cytology; Myoblasts - drug effects; Myoblasts - metabolism; Phosphotransferases - antagonists & inhibitors; Predictive Value of Tests; Time Factors
• ISSN: 0002-7863; 1520-5126
• DOI: 10.1021/ja077656d

A small molecule capable of distinguishing the distinct states resulting from cellular differentiation would be of enormous value, for example, in efforts aimed at regenerative medicine. We screened a collection of fluorescent small molecules for the ability to distinguish the differentiated state of a mouse skeletal muscle cell line. High-throughput fluorescence-based screening of C2C12 myoblasts and myotubes resulted in the identification of six compounds with the desired selectivity, which was confirmed by high-content screening in the same cell states. The compound that resulted in the greatest fluorescence intensity difference between the cell states was used as the screening agent in a pilot screen of 84 kinase inhibitors, each present in four doses, for inhibition of myogenesis. Of the kinase inhibitors, 17 resulted in reduction of fluorescence at one or more concentrations; among the “hits” included known inhibitors of myogenesis, confirming that this compound is capable of detecting the differentiated myotube state. We suggest that the strategy of screening for screening agents reported here may be extended more broadly in the future.