Abstract
It has become generally accepted in the drug discovery community that even the best designed assays are prone to compound interference. However, the consequences of this have only become widely realized through the dedicated efforts of scientists concerned with the misdirected effort and erroneous conclusions arising from pursuit of compounds lacking bona fide useful biological activity. During high-throughput screening, many compounds appear to display a useful biologically relevant target interaction, when in fact they are interfering with the assay components or signal readout. It seems likely that widespread adoption of homogeneous screening technologies has increased the prevalence of compound interference due to the elimination of washing and filtration steps that typically remove excess compound before signal acquisition. However, the important point to emphasize here is that this need not be a problem. The advantages of homogeneous assays far outweigh any disadvantages, providing procedures are in place to triage interfering compounds and focus attention on high-value biologically relevant hits. Thus, it is essential that all scientists involved in compound screening are aware of the prevalence of assay interference and remain skeptical of active compounds until their value is proven.
Enlightened analysis of the output of compound screening is so important that we devote this issue of ASSAY and Drug Development Technologies to “Discriminating High-Value Hits from PAINS.” This Special Issue covers three essential ingredients of a successful drug (or chemical probe) discovery strategy: awareness of the nature of compound interference, procedures to identify interfering compounds and minimize their confounding effects, and workflows with stringent criteria for selection of high-value hits.
Following our popular Literature Search section, the Special Issue begins with an engaging interview with Jonathan Baell, who first coined the term “PAINS” (pan-assay interference compounds) to describe “nuisance compounds” that appear all too often as hits in high-throughput screens. In addition to covering the topic of PAINS, the interview provides interesting perspectives on science in Australia and highlights Dr. Baell's achievements in academic medicinal chemistry. Daylin and Walters continue the “PAINS” theme in a perspective that provides an in-depth analysis of the nature of these compounds, why they persist in the literature, and what can be done to minimize their future impact. These authors also provided humorous cover art that we present to highlight their contribution and the theme of the issue. Hall et al. approach the problem of compound interference from an assay design perspective by presenting a case for using diaphorase-based assays to avoid fluorescence interference. Their perspective is accompanied by a technical brief that provides evidence to support their case. The Special Issue is completed by two exemplary articles describing successful execution of strategies to identify high-value hits from high-throughput screens. Engen et al. identified and rigorously characterized inhibitors of insulin-regulated aminopeptidase, whereas Smith and Kirby validated a parallel screening approach to identify novel single agent antibiotics and synergistic potentiators of existing carbapenem antibiotics in carbapenem-resistant Enterobacteriaceae.
Overall, therefore, this Special Issue not only explores the problem of compound interference in high-throughput screening but also provides practical examples of successful and widely applicable strategies for avoiding the distraction of “nuisance compounds” and focusing instead on high-value hits.