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Journal of Biomedicine and Biotechnology
Volume 2011 (2011), Article ID 868095, 11 pages
doi:10.1155/2011/868095
Identification of Four Potential Epigenetic Modulators from the NCI Structural Diversity Library Using a Cell-Based Assay
1Department of Pharmacology and Hamon Center for Therapeutic Oncology Research, UT Southwestern Medical Center, 6000 Harry Hines Boulevard, Dallas, TX 75390, USA
2Molecular Targets Laboratory, Center for SAIC-Frederick, National Cancer Institute, Frederick, MD 21702, USA
3Molecular Targets Laboratory, Center for Cancer Research, National Cancer Institute, Boulevard, Dallas, TX 75390, USA
Received 29 July 2010; Accepted 7 October 2010
Academic Editor: Minoru Yoshida
Copyright © 2011 Anne M. Best et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Supplementary Material
To confirm that the fluorescent signal derived from assay hits was from bona fide GFP protein expression, we queried LDR cells with assay hits in the presence of estradiol. The estrogen receptor ligand binding domain is expressed downstream of GFP in our system (see main text Figure 1), such that with addition of estradiol, the entire fusion protein translocates to the nucleus. For all assay hits described, co-treatment with estradiol caused GFP nuclear translocation as shown in Supplementary Figure 1, demonstrating that the fluorescence signal was indeed originating from the GFP protein in response to treatment with our candidate drugs. Estradiol alone does not induce GFP production.
To evaluate the effects of assay hits on protein modifications, in addition to histone acetylation, we also examined changes in two other marks known to be affected by epigenetic modulators, global histone 3 trimethylation at lysine 9 and acetylated tubulin. While NSC-22206 and known HDAC inhibitor TSA both increased levels of acetylated tubulin, they did not change histone methylation levels. In contrast, NSC-159631 increased global histone 3 lysine 9 trimethylation levels, raising the possibility that this assay hit may affect histone methylases/demethylases directly or indirectly in cells.