Analysis of Current DNA Encoded Library Screening Data Indicates Higher False Negative Rates for Numerically Larger Libraries

Abstract

To optimize future DNA-encoded library design, we have attempted to quantify the library size at which the signal becomes undetectable. To accomplish this we (i) have calculated that percent yields of individual library members following a screen range from 0.002 to 1%, (ii) extrapolated that ∼1 million copies per library member are required at the outset of a screen, and (iii) from this extrapolation predict that false negative rates will begin to outweigh the benefit of increased diversity at library sizes >10. The above analysis is based upon a large internal data set comprising multiple screens, targets, and libraries; we also augmented our internal data with all currently available literature data. In theory, high false negative rates may be overcome by employing larger amounts of library; however, we argue that using more than currently reported amounts of library (≫10 nmoles) is impractical. The above conclusions may be generally applicable to other DNA encoded library platforms, particularly those platforms that do not allow for library amplification.

DNA Encoded Libraries: A Visitor's Guide

“…[27,[80][81][82] Despite notable successes, failures still abound in this area stemming from a lack of structural diversity, poorly designed assays, the inability to convert hits to actual leads, and difficulties in rapidly deconvoluting large numbers of false positives from authentic hits. [82] With regards to larger libraries, improvements in informatic and sequencing technology to rapidly deconvolute hits will have an immediate impact. [82][83][84] Finally, a greater understanding of the dynamics of small molecule binding when attached to DNA could facilitate more effective selections.…”

“…[82] With regards to larger libraries, improvements in informatic and sequencing technology to rapidly deconvolute hits will have an immediate impact. [82][83][84] Finally, a greater understanding of the dynamics of small molecule binding when attached to DNA could facilitate more effective selections. Ultimately the future success of the DEL-based screening paradigm will be reliant on a community-wide collaboration including both industry and academia to advance technology in informatics, sequencing, biochemistry, and synthetic organic chemistry.…”

Design and Construction of a Focused DNA-Encoded Library for Multivalent Chromatin Reader Proteins

“…Large libraries that contain random chemical diversity can increase chances for success for even traditionally difficult targets generally deemed 'undruggable'. However, it has also become apparent that library size does not necessarily correlate to hit rate [18][19][20]. Much like focused screens in traditional HTS, for example kinase-focused sets that retain the 'hinge-binding motif' [21,22], using structure-based information and computational methods for DEL design can improve success rates using smaller libraries (thousands to millions) [18,23].…”

Winner(s)-take-all: nonlinear amplification of DNA-encoded library

“…However, different from the high-fidelity DNA synthesis by DNA polymerase in cell replication cycles, both chemical synthesis and sequencing of DNA are error-prone. For example, in the construction of DNA-encoded chemical library (DEL), mistakes can be caused by side reactions and low reactivity in the combinatorial chemical library synthesis (Figure 1a, i ), as well as in the encoding procedure (Fiure 1a, ii) 5,6 . The affinitybased selection protocols with immobilized target proteins on polymer matrices may yield to decoding artifacts due to promiscuous interaction and protein misfolding (Figure 1a, iii and iv).…”

“…Indeed, the evaluation of the selection is typically accomplished by ranking the NGSsequence counts and it is based on the assumption that the binding affinity of the detected SMC (small molecular compound) to the native target protein, the amount of SMC-DNA conjugate captured on the polymer matrix, the PCR amplification, and the NGS reads are linearly correlated (Figure 1a). Unfortunately, errors and biases accumulate over the steps, and poor correlations between the measured binding affinities of selected compounds to target protein and the counts have frequently been observed 5,8 . DELs with more sophisticated structures have been constructed in recent years, exacerbating the problem by adding more synthetic and DNAmanipulation steps [9][10][11][12][13] .…”