PNA-Encoded Synthesis (PES) and DNA Display of Small-Molecule Libraries

DNA-encoded library technologies have emerged as a powerful platform to rapidly screen for binders to a protein of interest. These technologies are underpinned by the ability to encode a rich diversity of small molecules. While large libraries are accessible through cycles of mix and split synthesis, libraries based on single chemistries tend to be redundant. Furthermore, the quality of libraries generally decreases with the number of synthetic transformations required in their synthesis. An alternative approach is to use hybridization to program the combinatorial assembly of peptide nucleic acid (PNA)-encoded fragment pairs onto a library of DNA templates. A broad molecular diversity is more easily sampled since it arises from the pairing of diverse fragments. Upon identification of productive fragment pairs, a focused library in which the fragments are covalently linked is prepared. This focused library includes linkers of different lengths and geometries and offers the opportunity to enrich the selected fragment set with close neighbors. Herein we describe detailed protocols to covalently link diverse fragments and screen fragment-based libraries using a commercially available microarray platform. Additionally, we present a novel library constructed via a straightforward synthetic route that accesses previously unexplored chemical space.