High throughput screening for FEN 1 inhibitors using a fluorescence polarization assay
Overexpression of FEN 1 has been observed in multiple cancers and down regulation of FEN 1 sensitizes cells to chemotherapeutic agents. As of now, no inhibitors of FEN 1 that are sufficient to be pursued as drugs have been identified. Therefore, a research group from AstraZeneca sought to design a high throughput screening approach using the PHERAstar microplate reader from BMG LABTECH to identify novel inhibitors of FEN 1. The results of their efforts are published in the current issue of the Journal of Biomolecular Screening.
A FEN 1 oligonucleotide substrate was designed which incorporated a fluorescent label and included key features which have been shown to be important for FEN 1 specificity. The assay then uses fluorescence polarization performed on the PHERAstar to monitor the cleavage of this substrate. The PHERAstar features easy to use assay specific optic modules and a module designed to detect to fluorescence polarization was used in this assay. After assay validation, a screen of 850,000 compounds was performed. A Z' of greater than 0.6 was observed throughout the screening indicating that the PHERAstar is important in delivering a simple and robust assay even in low volumes; like the 1536 well plates used in this assay. In the end 6261 compounds were confirmed as FEN 1 inhibitors.