Fluorescence lifetime assays- an attractive addition to the toolbox of fragment screening technologies (T307)

Hafenbradl D

Fluorescence lifetime (FLT) has recently gained new attraction as a screening technology in drug discovery through the introduction of the high throughput screening (HTS) compatible NanoTaurus FLT plate reader (Edinburgh Instruments) and significant improvements in substrate and assay design using the new fluorophore 9AA as a long lifetime fluorophore reporter (Almac FLEXYTE®). By its nature, FLT has many benefits over other screening technologies; the most important of these are markedly reduced assay interferences. Here we describe novel FLT assays developed at BioFocus. Assay conditions were optimized and activities of reference compounds, and compounds from the BioFocus diverse compound and fragment libraries were determined and compared to orthogonal assay technologies (fluorescence intensity (FI) for metalloprotease and Caliper LC3000 mobility shift assay (MSA) for p38alpha). Fragment-based drug discovery (FBDD) is considered an increasingly important area of drug discovery combining structural biology, binding studies and functional assays which assess the activity of fragments against molecular targets. A subset (42 compounds) of the BioFocus fragment library was tested in the FLT and Caliper LC3000 MSA assays. A good correlation of inhibitory activities observed when both functional assays were compared, which adds the FLT format as a valuable option for FBDD assays. Together with the excellent precision obtained using the combination of the NanoTaurus FLT reader and the Almac FLEXYTE® assay technology, these results render FLT a highly attractive assay technology for fragment based drug discovery.

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