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1.  A Set of Time-Resolved Fluorescence Resonance Energy Transfer Assays for the Discovery of Inhibitors of Estrogen Receptor-Coactivator Binding 
Journal of biomolecular screening  2009;14(2):181-193.
Therapeutic block of estrogen action is typically achieved with conventional antagonists (CAs), compounds that displace estradiol from the estrogen receptor (ER) and induce formation of an ER conformation that cannot bind to coactivator proteins, such as the steroid receptor coactivators (SRCs). As an alternative mode for blocking estrogen action, we are seeking small molecules that act as coactivator binding inhibitors (CBIs), i.e., they compete directly with SRC3 for interaction with estradiol-bound ER. CBIs would be interesting mechanistic probes of estrogen action and might also provide an alternative, more durable endocrine therapy for hormone-responsive breast cancer, where cellular adaptations lead to resistance to CAs. We have designed and optimized a set of time-resolved fluorescence resonance energy transfer (TR-FRET) assays to monitor the interaction of ER with SRC3 and ligands, and we have used them in high-throughput screens to discover small molecule CBIs that are able to disrupt this interaction. These assays also distinguish CBIs from CAs. These robust and sensitive “mix and measure” assays use low concentrations of ER labeled with a europium chelate as FRET donor and a Cy5-labeled SRC as acceptor. This multiplexed protocol produces excellent signal to noise ratios (> 100) and Z' values (> 0.8).
PMCID: PMC2731238  PMID: 19196699
2.  High-Throughput Screening Based Identification of Paramyxovirus Inhibitors 
Journal of biomolecular screening  2008;13(7):591-608.
Paramyxoviruses are negative strand non-segmented RNA viruses. Several members of this family constitute major human pathogens that, collectively, are responsible for major morbidity and mortality worldwide. In an effort to ultimately develop novel therapeutics against measles virus (MV), a prominent member of the paramyxovirus family, we report a high-throughput screening protocol that allows hit identification using non-recombinant primary MV strains as targets. Implementation of the assay has yielded 60 hit candidates from a 137,500-entry library. Counterscreening and generation of dose-response curves narrows this pool to 35 compounds with active concentrations ≤15.3 μM against the MV-Alaska strain and specificity indices ranging from 36 to >500. Library mining for structural analogs of several confirmed hits combined with re-testing of identified candidates reveals a low false-negative rate and, thus, a high accuracy of primary hit identification. Eleven of the confirmed hits were found to interfere with the viral entry machinery, while the remaining 24 compounds target post-entry steps of the viral life cycle. Activity testing against selected members of the paramyxovirus family reveals three patterns of activity: 1) exclusively MV-specific blockers; 2) inhibitors of MV and related viruses of the same genus; 3) broader-range inhibitors with activity against a different paramyxovirinae genus. Representatives of the last class may open avenues for the development of broad-range paramyxovirus inhibitors through hit-to-lead chemistry.
PMCID: PMC2556223  PMID: 18626114
high-throughput screening; drug discovery; paramyxovirus; anti-infective drugs

Results 1-2 (2)