Bayer Healthcare, Berlin, Germany
5th HTRF Symposium, Avignon, France
An overview of ultra high-throughput screening at Bayer with focus on key aspects especially for cell-based assays in GPCR drug discovery is given. General procedures using frozen cells as well as the concept of ready-to-assay plates are outlined and examples of troubleshooting will be highlighted. In more detail, main steps of the development of an uHTS-compatible IP-One assay for GPCR antagonist screening are demonstrated. With this assay a whole library screening campaign in 1536-MTP format was performed including confirmation assay and IC50 determination of hits. As a general procedure, first insights into Mode-of-action of hits were acquired by variation of agonist concentration. To exclude signaling cascade interferers and/or get further hints for the Mode-of-action, Tag-lite® binding assay represented the best option for this project. Therefore, Tag-lite® binding assay was transferred to Bayer and had to be miniaturized to fit to 1536-MTP format. The established and optimized assay was also used for IC50 determination of putative hits from functional screening. Finally, a complete data set for 3000 putative hits in IP-One assay with high and low agonist as well as Tag-lite® assay was accomplished. A good correlation of functional and binding data proved the quality of hits and assay techniques used. Hits from this screening campaign will be further characterized and optimized by using these established assays as well as additional IP-One assays for related GPCRs.