title: A fluorescent screening platform for the rapid evaluation of chemicals in cellular reprogramming.
abstract: Current strategies to monitor reprogramming into induced pluripotent stem cells (iPSCs) are limited in that they rely on the recognition of advanced stage biomarkers or they involve the transduction of genetically-modified cells. These limitations are particularly problematic in high-throughput screenings where cell availability, low cost and a rapid experimental protocol are critical issues. Herein we report the application of a pluripotent stem cell fluorescent probe (i.e. CDy1) as a reporter for the rapid screening of chemicals in reprogramming iPSCs. CDy1 stains early-stage iPSCs at 7dpi as well as matured iPSCs; hence it can partially overcome the slow kinetics of the reprogramming process. As a proof of concept, we employed a CDy1-based screening in 384 well-plates to examine the effect of newly synthesized hydroxamic acid derivatives in reprogramming mouse fibroblasts transduced with Oct4, Sox2 and Klf-4 without c-Myc. One compound (1-26) was identified as a reprogramming enhancer by 2.5-fold and we confirmed that 1-26 behaves as a histone deacetylase (HDAC) inhibitor. The successful identification of novel small molecules enhancing the generation of iPSCs by means of a rapid and simple protocol demonstrates the suitability of this CDy1-based screening platform for the large scale and high-throughput evaluation of iPSC modulators.
authors.affiliation: Laboratory of Bioimaging Probe Development, Singapore Bioimaging Consortium, Agency for Science, Technology and Research (A*STAR), 11 Biopolis Way, 138667 Singapore, Singapore.
authors.forename: Marc
authors.initials: M
authors.lastname: Vendrell
authors.forename: Sung-Jin
authors.initials: SJ
authors.lastname: Park
authors.forename: Yogeswari
authors.initials: Y
authors.lastname: Chandran
authors.forename: Chi-Lik Ken
authors.initials: CL
authors.lastname: Lee
authors.forename: Hyung-Ho
authors.initials: HH
authors.lastname: Ha
authors.forename: Nam-Young
authors.initials: NY
authors.lastname: Kang
authors.forename: Seong-Wook
authors.initials: SW
authors.lastname: Yun
authors.forename: Young-Tae
authors.initials: YT
authors.lastname: Chang
mesh_terms: D000818:Animals; D000873:Anthracenes / Q000032:analysis / Q000378:metabolism; D002454:Cell Differentiation; D002478:Cells, Cultured; D005347:Fibroblasts / Q000737:chemistry / Q000166:cytology / Q000378:metabolism; D005456:Fluorescent Dyes / Q000032:analysis / Q000378:metabolism; D057166:High-Throughput Screening Assays / Q000379:methods*; D057026:Induced Pluripotent Stem Cells / Q000737:chemistry* / Q000166:cytology* / Q000378:metabolism; D051379:Mice; D016513:Mice, SCID; D008856:Microscopy, Fluorescence / Q000379:methods*; D009025:Morpholines / Q000032:analysis / Q000378:metabolism; D050814:Octamer Transcription Factor-3 / Q000235:genetics / Q000378:metabolism; D055748:SOXB1 Transcription Factors / Q000235:genetics / Q000378:metabolism
chemical_list: D000873:Anthracenes; C568650:CDy1 compound; D005456:Fluorescent Dyes; D009025:Morpholines; D050814:Octamer Transcription Factor-3; C494174:Pou5f1 protein, mouse; D055748:SOXB1 Transcription Factors; C527783:Sox2 protein, mouse