A quantitative high-throughput screen identifies potential epigenetic modulators of gene expression

Ronald L. Johnson, Wenwei Huang, Ajit Jadhav, Christopher P. Austin, James Inglese, Elisabeth D. Martinez

Research output: Contribution to journalArticle

23 Citations (Scopus)

Abstract

Epigenetic regulation of gene expression is essential in embryonic development and contributes to cancer pathology. We used a cell-based imaging assay that measures derepression of a silenced green fluorescent protein (GFP) reporter to identify novel classes of compounds involved in epigenetic regulation. This locus derepression (LDR) assay was screened against a 69,137-member chemical library using quantitative high-throughput screening (qHTS), a titration-response method that assays compounds at multiple concentrations. From structure-activity relationships of the 411 actives recovered from the qHTS, 6 distinct chemical series were chosen for further study. A total of 48 qHTS actives and analogs were counterscreened using the parental line of the LDR cells, which lack the GFP reporter. Three series-8-hydroxy quinoline, quinoline-8-thiol, and 1,3,5-thiadiazinane-2-thione-were not fluorescent and reconfirmed activity in the LDR cells. The three active series did not inhibit histone deacetylase activity in nuclear extracts or reactivate the expression of the densely methylated p16 gene in cancer cells. However, one series induced expression of the methylated CDH13 gene and inhibited the viability of several lung cancer lines at submicromolar concentrations. These results suggest that the identified small molecules act on epigenetic or transcriptional components and validate our approach of using a cell-based imaging assay in conjunction with qHTS.

Original languageEnglish (US)
Pages (from-to)237-248
Number of pages12
JournalAnalytical Biochemistry
Volume375
Issue number2
DOIs
StatePublished - Apr 15 2008

Fingerprint

Epigenomics
Gene expression
Modulators
Assays
Screening
Throughput
Gene Expression
Green Fluorescent Proteins
Genes
Thiones
Imaging techniques
Histone Deacetylases
Small Molecule Libraries
p16 Genes
Pathology
Titration
Sulfhydryl Compounds
Gene Expression Regulation
Structure-Activity Relationship
Cells

Keywords

  • Cancer
  • Cell assay
  • Epigenetic
  • GFP
  • HDAC
  • HTS
  • Small molecule

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology

Cite this

A quantitative high-throughput screen identifies potential epigenetic modulators of gene expression. / Johnson, Ronald L.; Huang, Wenwei; Jadhav, Ajit; Austin, Christopher P.; Inglese, James; Martinez, Elisabeth D.

In: Analytical Biochemistry, Vol. 375, No. 2, 15.04.2008, p. 237-248.

Research output: Contribution to journalArticle

Johnson, Ronald L. ; Huang, Wenwei ; Jadhav, Ajit ; Austin, Christopher P. ; Inglese, James ; Martinez, Elisabeth D. / A quantitative high-throughput screen identifies potential epigenetic modulators of gene expression. In: Analytical Biochemistry. 2008 ; Vol. 375, No. 2. pp. 237-248.
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