Development of a facile method for high throughput screening with reporter gene assays

Area of Science:

• Biochemistry
• Cell Biology
• Pharmacology

Background:

• High-throughput screening (HTS) is crucial for drug discovery.
• Stable mammalian cell reporter gene assays are valuable tools for HTS.
• Existing methods can be limited by cost, throughput, and compound stability.

Purpose of the Study:

• To develop a facile and efficient methodology for high-throughput screening using stable mammalian cell reporter gene assays.
• To adapt an adherent cell assay to a suspension cell format for improved scalability and cost-effectiveness.
• To validate a new suspension cell assay for screening Galpha(s)- and Galpha(i)-coupled seven-transmembrane receptors.

Main Methods:

• Adapted a 96-well adherent cell assay to a suspension cell format using 96- and 384-well plates.
• Validated a stable Chinese Hamster Ovary (CHO) cell line expressing 6xCRE-luciferase as a reporter gene host.
• Used a derivative CHO cell line stably transfected with the human melanocortin-1 receptor to test the assay's performance.

Main Results:

• The suspension cell assay demonstrated comparable results to the adherent method for receptor response.
• The assay is suitable for screening compounds modulating cyclic adenosine monophosphate (cAMP) levels.
• A single user can screen 100 plates per day in the 384-well format with minimal hands-on time and without expensive automation.

Conclusions:

• The developed suspension cell assay offers significant improvements in cost, throughput, cell culture effort, compound stability, accuracy, and hands-on time.
• This method is highly adaptable for screening various targets and transcriptional response elements.
• The assay provides a high-capacity, cost-effective solution for pharmacological analysis and drug discovery.