Lisa A. Marcaurelle

Aldehydes are central synthons for DNA-encoded library (DEL) diversification, yet their limited commercial availability and stability restrict the accessible chemical space. Alcohols, a much more abundant pool of potential building blocks, have far fewer reported avenues to use in DNA-compatible chemistry. We report a DNA-compatible one-pot method that converts primary alcohols into aldehydes off-DNA via Cu(I)/ABNO-mediated Stahl oxidation followed directly by on-DNA reductive alkylation. Systematic optimization of the ligand, reaction time, and solvent composition enabled efficient oxidation under microplate conditions, while a premixed catalyst solution significantly improved reproducibility. A plate screen established broad applicability across substrate classes such as benzylic, heterobenzylic, and aliphatic alcohols. Benchmarking against reactions using isolated aldehydes confirmed that in situ-generated aldehydes perform comparably in reductive alkylation. The workflow also supports additional DNA-compatible transformations, including the Cushman reaction and imidazolidinone formation. qPCR analysis verified excellent DNA amplifiability throughout the sequence. Together, this platform provides a practical route to leverage abundant alcohol building blocks for expanded DEL diversification.

DNA-encoded library (DEL) technology is a powerful platform for hit identification in academia and the pharmaceutical industry. When conducting off-DNA resynthesis hit confirmation after affinity selection, PCR/sequencing, and data analysis, one typically assumes a “one-to-one” relationship between the DNA tag and the chemical structure of the attached small-molecule it encodes. Because library synthesis often yields a mixture, this approximation increases the risk of overlooking positive discoveries and valuable information. To address this issue, we apply a library synthesis “recipe” strategy for on-DNA resynthesis using a cleavable linker, followed by direct affinity selection mass spectrometry (AS-MS) evaluation and identification of binder(s) from the released small-molecule mixture. We validate and showcase this approach employing the receptor-interacting-protein kinase 2 (RIP2) DEL campaign. We also designed and developed two cleavable linkers to enable this method, a photocleavable linker (nitrophenyl-based) and acid-labile linker (tetrahydropyranyl ether). The strategy provides an effective means of hit identification and rapid determination of key active component(s) of the mixture.

We have implemented an interactive and practical sparse matrix design strategy for the synthesis of DOS libraries, which facilitates the selection of diverse library members within a user-defined range of physicochemical properties while still maintaining synthetic efficiency. The utility of this approach is illustrated with the synthesis of an 8000-membered library of stereochemically diverse medium-sized rings accessible via a build/couple/pair DOS strategy. Diverse library members were selected from a virtual library by applying the maximum dissimilarity method, while the selection of similar analogs around each diverse product was ensured by picking near neighbors algorithmically based on fingerprint comparison. Adjustable filters on compound properties, which can be tailored to suit the needs of the target biology, facilitated subset selection from the synthetically accessible compounds.