Picomole‐Scale Synthesis and Screening of Macrocyclic Compound Libraries by Acoustic Liquid Transfer

Macrocyclic compounds are an attractive class of therapeutic ligands against challenging targets, such as protein-protein interactions. However, the development of macrocycles as drugs is hindered by the lack of large combinatorial macrocyclic libraries, which are cumbersome, expensive, and time consuming to make, screen, and deconvolute. Here, we established a strategy for synthesizing and screening combinatorial libraries on a picomolar scale by using acoustic droplet ejection to combine building blocks at nanoliter volumes, which reduced the reaction volumes, reagent consumption, and synthesis time. As a proof-of-concept, we assembled a 2700-member target-focused macrocyclic library that we could subsequently assay in the same microtiter synthesis plates, saving the need for additional transfers and deconvolution schemes. We screened the library against the MDM2-p53 protein-protein interaction and generated micromolar and sub-micromolar inhibitors. Our approach based on acoustic liquid transfer provides a general strategy for the development of macrocycle ligands.