DNA-encoded chemical libraries: foundations and applications in lead discovery

Abstract

DNA-encoded chemical libraries have emerged as a powerful tool for hit identification in the pharmaceutical industry and in academia. Similar to biological display techniques (such as phage display technology), DNA-encoded chemical libraries contain a link between the displayed chemical building block and an amplifiable genetic barcode on DNA. Using routine procedures, libraries containing millions to billions of compounds can be easily produced within a few weeks. The resulting compound libraries are screened in a single test tube against proteins of pharmaceutical interest and hits can be identified by PCR amplification of DNA barcodes and subsequent high-throughput sequencing.

Figure 1

Encoding strategies for the construction of DNA-encoded chemical libraries. Two synthetic strategies are employed: DNA-recorded chemical library synthesis and DNA-templated library synthesis. In DNA-recorded chemical library synthesis encoding may achieved by ligation using a) single, b) double stranded DNA or c) polymerase-catalyzed fill-in reactions.

Figure 2

Different library formats employed for DNA-encoded compound collections. So-called single-pharmacophore libraries (left) employ only one DNA-strand for presenting chemical building blocks. Single pharmacophore libraries are synthesized in a split-and-pool approach resulting in combinatorial libraries of A x B ( in case of a two building block library). Dual pharmacophore libraries (right) are assembled by hybridization of individually synthesized sublibraries A and B (in case of a two building block library).

Figure 3

Dual display, DNA-encoded compound collections employing the “Encoded self-assembling chemical libraries” (ESAC) approach. Sublibrary 1 and 2 are individually synthesized and purified. Sublibrary 1 is generated by coupling a chemical building block to an oligonucleotide which contains an abasic site. Subsequently, enzyme-catalyzed ligation delivers a stable link between the chemical building block and the encoding nucleotide sequence. Sublibrary 2 is synthesized by coupling of chemical building blocks to oligonucleotides bearing a short nucleotide coding sequence. The final combinatorial library is assembled by hybridization and the coding information is transferred to one strand, making the system suitable for decoding by high-throughput sequencing.

Figure 4

Flowchart for hit identification employing DNA-encoded chemical libraries. Libraries are typically incubated with an immobilized target protein of interest and subsequently washed to select binders of the protein. Bound DNA-tagged molecules may then be eluted by heat-denaturation of the target protein. Coding oligonucleotides are then amplified by PCR and submitted for high-throughput sequencing. Identified hits are resynthesized without the DNA tag and validated by biophysical assays such as fluorescence polarization.

Figure 5

Different selection methods for DNA-encoded chemical libraries. Magnetic beads and resins in pipette tips (entries 1 and 2) are routinely used for the selection against target proteins. Electrophoretic separation techniques (entries 3 and 4) have only been used for model selections. Similar model affinity enrichments have been demonstrated using in-solution procedures (entries 5 and 6), whereas novel hits have been identified by directly panning DNA-encoded libraries against receptors on the cell surface (entries 7).