Hit Discovery Methodology

This special issue was conceived with the intent of highlighting the use of existing and progressive technologies in hit discovery and how these technologies may be combined to generate vital hits for drug discovery. These aspects now cross both small-molecule and large-molecule discovery in many organizations. In addition, the issue is intended to remind readers of the heritage of SLAS journals, and how they have supported the use and application of innovative hit identification approaches.

Working with SLAS, we have compiled a list of the most downloaded articles from SLAS journals. All of the articles referenced below have been downloaded more than 4000 times. Many of these very nicely reflect the focus on new technology, its implementation, and the continued interest in how hit matter is identified, characterized, and progressed.

It is very fitting that the article downloaded most frequently (more than 16,000 times) is one that describes the statistical definition of z-factor and how this can be used to define a high-throughput screening (HTS) window aiding the development of HTS assays. ¹ Understanding your HTS assay is further highlighted by high-demand publications describing plate edge effects, ² the relationship between % inhibition and IC₅₀, ³ the time dependency of IC₅₀, ⁴ and the effects of covalent and irreversible molecules on IC₅₀ kinetics. ⁵ In addition to achieving high-quality assays for hit identification, the data show how important it is to conserve the quality of hit matter. Two sample management publications describe the effects of freeze–thaw and managing compound storage in DMSO.^6,7

The list continues to illustrate many approaches that were new or had new implications at the time of the article coming to press. The impact of moving to more complex screening systems is highly represented, with six of the top 20 articles involving some form of screening approach or novel cellular assay system.^8–13 Finally, in this dataset, we can see how large molecules, ¹⁴ complex data analysis in combination screening, ¹⁵ as well as new technology^16,17 can advance hit identification approaches, and how the SLAS journals have constantly represented this section of the drug discovery industry.

In this issue, we have tried to capture stories that exemplify how technologies have been applied successfully to find hits. These often require a combination of both new and existing technologies to identify the best-quality hits.

Emery Smith et al. ¹⁸ open up the special issue with a phenotypic assay approach; on the surface, the assay technology would not be out of place in any of the past 10 years of SLAS journals. This article, however, uses a well-established cellular approach in a membrane potential dye to rethink how this can be used to find new hits, generating a phenotypic assay with a novel approach looking for readthrough rescue of a response. It also nicely describes how, especially in cellular assays, a combination of assays in a hit triage is used and constructed to increase the confidence in hits.

Similarly, Lorena Kallal et al. ¹⁹ use several technologies that have been well used in HTS historically but combines them in a unique way to design both a successful screening assay and a powerful cascade. The article combines six different cellular assay systems to rule compounds in and out of project progression.

Traditional HTS approaches remain extremely important for the mainstay of hit identification. The article from Saman Honarnejad et al. ²⁰ reviewed European Lead Factory collaborative screening efforts that have developed significant experience in hit identification. In their article, they are able to describe a very wide array of screening technologies and successes.

This brings us to newer approaches, as addressed in the technology review ²¹ by David McLaren et al. An example article by Sahasrabuddhe et al. ²² describes how mass spectrometry has established itself in recent years as a screening technology, capable of being applied as both a discrete screening assay and a multiplexed-affinity selection method.

The next two articles reinforce the importance of compound quality and properties when developing small-molecule screening cascades. Wilson Shou et al. ²³ defined how proper high-throughput analysis ensured maintenance of compound integrity and determined the highest-quality molecules to be followed up for HTS. In their article, Gareth Davies et al. ²⁴ shared a high-throughput approach to decipher the mechanism of action of compounds at enzyme targets.

Alternative approaches to traditional HTS-based hit identification are covered in the last four article. Each challenges the way we see the future of hit identification. Gabriel Dreiman et al. ²⁵ described how, with the increase in the capability of in silico approaches, iterative screening has gained favor, especially when reagents or resources are limited, such as with complex cell models. Timothy Foley et al. ²⁶ shared the history of DNA-encoded libraries within Pfizer, again indicating that often a combination of approaches to identify the best starting points is required. The authors also contrasted different hit identification paradigms. We are reminded that both complex cellular screening and large-molecule approaches now feature among the new paradigms of hit discovery. Hui Dou et al. ²⁷ demonstrated this point with a nice example of a high-content imaging assay that combines branched DNA (bDNA) technology with fluorescence in situ hybridization (FISH) to measure gene silencing by small interfering RNAs (siRNAs). And, finally, the possibility of not actually screening a compound collection but using knowledge-driven, phenotypic analysis is a very exciting proposition and a capability that is rapidly being established throughout the pharma and biotech industries. Johanna Nyffeler et al. ²⁸ compared different approaches to determining bioactivity hits from high-dimensional profiling data and framed the concept that compound signatures can be identified and applied in drug discovery.

Finally, we would like to thank everyone who has contributed to this special issue and hope that it serves as a useful, thought-provoking guide to the readers.