Enzymatic Assays for High‐Throughput Screening

Abstract

AbstractHigh‐throughput screening (HTS) is a multidisciplinary approach to investigate in parallel the pharmacological properties of a large number of small molecules, with the aim of identifying target‐specific lead compounds to be progressed as bioactive probes or therapeutic drugs. The complex integration of chemistry, biology, automation, informatics, and medicinal chemistry has prompted the evolution of HTS as an autonomous discipline integrated in the drug discovery process. Thus, HTS assays are differentiated from traditional laboratory assays by the requirement of extensive assay development and by the strict quality criteria that must be fulfilled to approach a screening campaign. In this perspective, biochemistry and enzymology have provided an essential support for the design and configuration of functional HTS assays for enzymatic targets. In turn, the impressive technological improvements of HTS have made available an unprecedented array of novel methods and instrumentations that have contributed to advance the biochemical and kinetic characterization of enzymes. In this chapter, the paradigms for the configuration of enzymatic assays for HTS are reviewed, emphasizing the tight interdependence between type and quality of the HTS assay and the outcome of a screening campaign. The sequential steps of assay development for HTS are examined, providing examples of alternative approaches to configure the activity of enzyme drug targets as HTS‐compatible assays.