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Journal of Biomedicine and Biotechnology
Volume 2011 (2011), Article ID 868095, 11 pages
Research Article

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.

  1. Supplementary Material