Jeff Reese
Last active: 1/7/2021

High-Throughput Screening of Myometrial Calcium-Mobilization to Identify Modulators of Uterine Contractility.

Herington JL, Swale DR, Brown N, Shelton EL, Choi H, Williams CH, Hong CC, Paria BC, Denton JS, Reese J
PLoS One. 2015 10 (11): e0143243

PMID: 26600013 · PMCID: PMC4658040 · DOI:10.1371/journal.pone.0143243

The uterine myometrium (UT-myo) is a therapeutic target for preterm labor, labor induction, and postpartum hemorrhage. Stimulation of intracellular Ca2+-release in UT-myo cells by oxytocin is a final pathway controlling myometrial contractions. The goal of this study was to develop a dual-addition assay for high-throughput screening of small molecular compounds, which could regulate Ca2+-mobilization in UT-myo cells, and hence, myometrial contractions. Primary murine UT-myo cells in 384-well plates were loaded with a Ca2+-sensitive fluorescent probe, and then screened for inducers of Ca2+-mobilization and inhibitors of oxytocin-induced Ca2+-mobilization. The assay exhibited robust screening statistics (Z´ = 0.73), DMSO-tolerance, and was validated for high-throughput screening against 2,727 small molecules from the Spectrum, NIH Clinical I and II collections of well-annotated compounds. The screen revealed a hit-rate of 1.80% for agonist and 1.39% for antagonist compounds. Concentration-dependent responses of hit-compounds demonstrated an EC50 less than 10μM for 21 hit-antagonist compounds, compared to only 7 hit-agonist compounds. Subsequent studies focused on hit-antagonist compounds. Based on the percent inhibition and functional annotation analyses, we selected 4 confirmed hit-antagonist compounds (benzbromarone, dipyridamole, fenoterol hydrobromide and nisoldipine) for further analysis. Using an ex vivo isometric contractility assay, each compound significantly inhibited uterine contractility, at different potencies (IC50). Overall, these results demonstrate for the first time that high-throughput small-molecules screening of myometrial Ca2+-mobilization is an ideal primary approach for discovering modulators of uterine contractility.

MeSH Terms (17)

Animals Calcium Calcium Channel Blockers Cells, Cultured Dose-Response Relationship, Drug Drug Discovery Female High-Throughput Screening Assays Humans Mice Myometrium Oxytocin Pregnancy Primary Cell Culture Reproducibility of Results Uterine Contraction Uterus

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