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Development of Novel Cell Lines for High-Throughput Screening to Detect Estrogen-Related Receptor Alpha Modulators

Christina T. Teng, Jui-Hua Hsieh, Jinghua Zhao, Ruili Huang, Menghang Xia, Negin Martin, Xiaohua Gao, Darlene Dixon, Scott S. Auerbach, Kristine L. Witt, B. Alex Merrick.
SLAS Discovery (2017) DOI: https://doi.org/10.1177/2472555216689772 PMID: 28346099


Publication


Abstract

Estrogen-related receptor alpha (ERRα), the first orphan nuclear receptor discovered, is crucial for the control of cellular energy metabolism. ERRα and its coactivator, peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α), are required for rapid energy production in response to environmental challenges. They have been implicated in the etiology of metabolic disorders such as type 2 diabetes and metabolic syndrome. ERRα also plays a role in the pathogenesis of breast cancer. Identification of compounds that modulate ERRα signaling may elucidate environmental factors associated with these diseases. Therefore, we developed stable cell lines containing an intact ERRα signaling pathway, with and without the coactivator PGC-1α, to use as high-throughput screening tools to detect ERRα modulators. The lentiviral PGC-1α expression constructs and ERRα multiple hormone response element (MHRE) reporters were introduced into HEK293T cells that express endogenous ERRα. A cell line expressing the reporter alone was designated "ERR." A second cell line expressing both reporter and PGC-1α was named "PGC/ERR." Initial screenings of the Library of Pharmacologically Active Compounds (LOPAC) identified 33 ERR and 22 PGC/ERR agonists, and 54 ERR and 15 PGC/ERR antagonists. Several potent ERRα agonists were dietary plant compounds (e.g., genistein). In conclusion, these cell lines are suitable for high-throughput screens to identify environmental chemicals affecting metabolic pathways and breast cancer progression.

Figures


Figure 1. Concentration–response curves for the reference agonist genistein.

Concentration–response curves for the reference agonist genistein and the reference antagonist XCT790 in the ERR and PGC/ERR cell lines. Data are reported as mean ± SEM from three independent experiments.

Figure 2. Venn diagram illustrating the overlap between agonists (A) and antagonists (B).

Venn diagram illustrating the overlap between agonists (A) and antagonists (B) in the ERR and PGC/ERR assays.

Figure 3. Dose–response curves for compounds identified as actives in both ERR and PGC/ERR.

Cell lines, agonists (A, B) and antagonists (C, D); black squares, ERR line; black circles, PGC/ERR line; open squares, cytotoxicity for ERR line; open circles, cytotoxicity for PGC/ERR line; X axis, concentration of compounds; Y axis, response, as a percentage of the positive control. All data points are shown as mean ± SEM from three independent experiments.

Tables


Table 1. Compounds from the LOPAC That Were Identified as Agonists.

Compounds from the LOPAC That Were Identified as Agonists in Either the ERR or PGC/ERR Assays.

Table 2. Compounds from the LOPAC Collection That Were Identified as Antagonists.

Compounds from the LOPAC Collection That Were Identified as Antagonists in Either the ERR or PGC/ERR Assays.

Table 3. Activities (AC50) of ERR, PGC/ERR Agonists and Antagonists.

3A. Activities (AC50) of ERR and PGC/ERR Agonists in Other qHTS Agonist Screens with the LOPAC Collection.
3B. Activities (AC50) of ERR and PGC/ERR Antagonists in Other qHTS Antagonist Screens with the LOPAC Collection.

Supplemental Materials


Supplemental Data

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      • vendor/yajra/laravel-oci8/src/Oci8/Query/OracleBuilder.php:159
      • view::binviews::layouts.manuscript:52
      • vendor/laravel/framework/src/Illuminate/Filesystem/Filesystem.php:124
      • vendor/laravel/framework/src/Illuminate/View/Engines/PhpEngine.php:58
      • vendor/laravel/framework/src/Illuminate/View/Engines/CompilerEngine.php:75
    • OracleBuilder.php#159CEBSBIN5.27msselect * from "BIN_REVIEW" where "BIN_REVIEW"."FILE_ID" = '1de240ff62ed87f901636f1d949a03ea' and "BIN_REVIEW"."FILE_ID" is not null
      Bindings
      • 0: 1de240ff62ed87f901636f1d949a03ea
      Hints
      • Use SELECT * only if you need all columns from table
      Backtrace
      • vendor/yajra/laravel-oci8/src/Oci8/Query/OracleBuilder.php:159
      • modules/BinViews/Models/File.php:63
      • modules/BinViews/Models/Library.php:65
      • modules/BinViews/Models/Library.php:64
      • view::binviews::layouts.manuscript:54
    • OracleBuilder.php#159CEBSBIN2.51msselect "BIN_FILE".*, "BIN_LIBRARY_FILES"."LIBRARY_ID" as "LARAVEL_THROUGH_KEY" from "BIN_FILE" inner join "BIN_LIBRARY_FILES" on "BIN_LIBRARY_FILES"."FILE_ID" = "BIN_FILE"."FILE_ID" where "BIN_LIBRARY_FILES"."LIBRARY_ID" = 9175 and "IS_DELETED" != 'T'
      Bindings
      • 0: 9175
      • 1: T
      Hints
      • The != operator is not standard. Use the <> operator to test for inequality instead.
      Backtrace
      • vendor/yajra/laravel-oci8/src/Oci8/Query/OracleBuilder.php:159
      • view::binviews::layouts.manuscript:52
      • vendor/laravel/framework/src/Illuminate/Filesystem/Filesystem.php:124
      • vendor/laravel/framework/src/Illuminate/View/Engines/PhpEngine.php:58
      • vendor/laravel/framework/src/Illuminate/View/Engines/CompilerEngine.php:75
    • OracleBuilder.php#159CEBSBIN4.89msselect * from "BIN_REVIEW" where "BIN_REVIEW"."FILE_ID" = '1de240ff62ed87f901636f1e27d103eb' and "BIN_REVIEW"."FILE_ID" is not null
      Bindings
      • 0: 1de240ff62ed87f901636f1e27d103eb
      Hints
      • Use SELECT * only if you need all columns from table
      Backtrace
      • vendor/yajra/laravel-oci8/src/Oci8/Query/OracleBuilder.php:159
      • modules/BinViews/Models/File.php:63
      • modules/BinViews/Models/Library.php:65
      • modules/BinViews/Models/Library.php:64
      • view::binviews::layouts.manuscript:54
    • OracleBuilder.php#159CEBSBIN3.55msselect "BIN_FILE".*, "BIN_LIBRARY_FILES"."LIBRARY_ID" as "LARAVEL_THROUGH_KEY" from "BIN_FILE" inner join "BIN_LIBRARY_FILES" on "BIN_LIBRARY_FILES"."FILE_ID" = "BIN_FILE"."FILE_ID" where "BIN_LIBRARY_FILES"."LIBRARY_ID" = 9177 and "IS_DELETED" != 'T'
      Bindings
      • 0: 9177
      • 1: T
      Hints
      • The != operator is not standard. Use the <> operator to test for inequality instead.
      Backtrace
      • vendor/yajra/laravel-oci8/src/Oci8/Query/OracleBuilder.php:159
      • view::binviews::layouts.manuscript:52
      • vendor/laravel/framework/src/Illuminate/Filesystem/Filesystem.php:124
      • vendor/laravel/framework/src/Illuminate/View/Engines/PhpEngine.php:58
      • vendor/laravel/framework/src/Illuminate/View/Engines/CompilerEngine.php:75
    • OracleBuilder.php#159CEBSBIN4.69msselect * from "BIN_REVIEW" where "BIN_REVIEW"."FILE_ID" = '1de240ff62ed87f901636f1e867203ec' and "BIN_REVIEW"."FILE_ID" is not null
      Bindings
      • 0: 1de240ff62ed87f901636f1e867203ec
      Hints
      • Use SELECT * only if you need all columns from table
      Backtrace
      • vendor/yajra/laravel-oci8/src/Oci8/Query/OracleBuilder.php:159
      • modules/BinViews/Models/File.php:63
      • modules/BinViews/Models/Library.php:65
      • modules/BinViews/Models/Library.php:64
      • view::binviews::layouts.manuscript:54
    • OracleBuilder.php#159CEBSBIN4.34msselect * from "BIN_REVIEW" where "BIN_REVIEW"."FILE_ID" = '1de240ff62ed87f901636f1e86b703ed' and "BIN_REVIEW"."FILE_ID" is not null
      Bindings
      • 0: 1de240ff62ed87f901636f1e86b703ed
      Hints
      • Use SELECT * only if you need all columns from table
      Backtrace
      • vendor/yajra/laravel-oci8/src/Oci8/Query/OracleBuilder.php:159
      • modules/BinViews/Models/File.php:63
      • modules/BinViews/Models/Library.php:65
      • modules/BinViews/Models/Library.php:64
      • view::binviews::layouts.manuscript:54
    • OracleBuilder.php#159CEBSBIN2.93msselect "BIN_FILE".*, "BIN_LIBRARY_FILES"."LIBRARY_ID" as "LARAVEL_THROUGH_KEY" from "BIN_FILE" inner join "BIN_LIBRARY_FILES" on "BIN_LIBRARY_FILES"."FILE_ID" = "BIN_FILE"."FILE_ID" where "BIN_LIBRARY_FILES"."LIBRARY_ID" = 9179 and "IS_DELETED" != 'T'
      Bindings
      • 0: 9179
      • 1: T
      Hints
      • The != operator is not standard. Use the <> operator to test for inequality instead.
      Backtrace
      • vendor/yajra/laravel-oci8/src/Oci8/Query/OracleBuilder.php:159
      • view::binviews::layouts.manuscript:52
      • vendor/laravel/framework/src/Illuminate/Filesystem/Filesystem.php:124
      • vendor/laravel/framework/src/Illuminate/View/Engines/PhpEngine.php:58
      • vendor/laravel/framework/src/Illuminate/View/Engines/CompilerEngine.php:75
    • OracleBuilder.php#159CEBSBIN4.53msselect * from "BIN_REVIEW" where "BIN_REVIEW"."FILE_ID" = '1de240ff62ed87f901636f1ea8c003ee' and "BIN_REVIEW"."FILE_ID" is not null
      Bindings
      • 0: 1de240ff62ed87f901636f1ea8c003ee
      Hints
      • Use SELECT * only if you need all columns from table
      Backtrace
      • vendor/yajra/laravel-oci8/src/Oci8/Query/OracleBuilder.php:159
      • modules/BinViews/Models/File.php:63
      • modules/BinViews/Models/Library.php:65
      • modules/BinViews/Models/Library.php:64
      • view::binviews::layouts.manuscript:54
    • OracleBuilder.php#159CEBSBIN1.91msselect * from "BIN" where "BIN_ID" = 4176
      Bindings
      • 0: 4176
      Hints
      • Use SELECT * only if you need all columns from table
      Backtrace
      • vendor/yajra/laravel-oci8/src/Oci8/Query/OracleBuilder.php:159
      • modules/BinViews/Services/BreadcrumbService.php:32
      • modules/BinViews/Routes/breadcrumbs.php:55
      • vendor/diglactic/laravel-breadcrumbs/src/Generator.php:64
      • vendor/diglactic/laravel-breadcrumbs/src/Generator.php:42
    Modules\BinViews\Models\Library
    8Library.php#?
    Modules\BinViews\Models\LibraryGroup
    8LibraryGroup.php#?
    Modules\BinViews\Models\File
    8File.php#?
    Modules\BinViews\Models\Review
    8Review.php#?
    Modules\BinViews\Models\Characteristic
    5Characteristic.php#?
    Modules\BinViews\Models\PublicationManuscript
    3PublicationManuscript.php#?
      web
      null
        _token
        mydml6S4HB0992JJabEHNf4vjTCwxMCYoIoF2Ksi
        ClearShow all
        Date ↕MethodURLData
        #12025-04-07 09:36:45GET/cebs/paper/41761762940