DNA-encoded library (DEL)-enabled discovery of proximity-inducing small molecules

Abstract

Molecular glues and bifunctional compounds that induce protein-protein associations provide a powerful and general mechanism to modulate cell circuitry. We sought to develop a platform for the direct discovery of compounds able to induce association of any two pre-selected proteins, using the first bromodomain of BRD4 and the VHL-elongin C-elongin B (VCB) complex as a test system. Leveraging the screening power of DNA-encoded libraries (DELs), we synthesized ~one million DNA-encoded compounds that possess a VHL-targeting fragment, a variety of connectors, and a diversity element generated by split-and-pool combinatorial chemistry. By screening our DEL against BRD4 in the presence and absence of VCB, we could identify VHL-bound molecules that simultaneously bind BRD4. For highly barcode-enriched library members, ternary complex formation leading to BRD4 degradation was confirmed in cells. Furthermore, a ternary complex crystal structure was obtained for the most enriched library member. Our work provides a foundation for adapting DEL screening to the discovery of proximity-inducing small molecules.

Optimization of PROTAC Ternary Complex Using DNA Encoded Library Approach

“…Importantly, the selection strategy can be applied to molecular glue discovery where positive cooperativity is a key requirement. Interestingly, similar work of BRD4/VHL PROTAC optimization using DEL approach has been recently reported with the DNA attachment point on the VHL ligand as an additional support of this strategy. Trivalent PROTACs targeting dual targets with a shared E3 ligase ligand have been reported and have shown potential pharmacological benefits. − The combination of two strategies may allow the construction of a trivalent PROTAC library, and screening against both POIs and desired E3 ligases would be a very interesting direction for exploration.…”

“…We chose trifunctional linkers (as shown in Figure A) in the design of the PROTAC DEL in order to minimize the potential interference of DNA with the POI and E3 ligase ligands. Several trivalent PROTACs targeting dual proteins using the same E3 ligase have been reported, which suggests that variations of the linker can be tolerated in multiple protein–protein interactions. − If the E3 ligase ligand has multiple known extension points that do not significantly affect its binding, a PROTAC DEL can be designed with the DNA linker attached to one end of the E3 ligase ligand and various linkers and POI binders attaching from the other end . We used this former DEL design as a demonstration of the general strategy for DEL-based PROTAC synthesis and ligand discovery.…”

“…36−38 If the E3 ligase ligand has multiple known extension points that do not significantly affect its binding, a PROTAC DEL can be designed with the DNA linker attached to one end of the E3 ligase ligand and various linkers and POI binders attaching from the other end. 39 We used this former DEL design as a demonstration of the general strategy for DELbased PROTAC synthesis and ligand discovery.…”

Biophysical and Computational Approaches to Study Ternary Complexes: A ‘Cooperative Relationship’ to Rationalize Targeted Protein Degradation

“…Ternary complex formation in intact cells can be studied using technologies such as NanoBRET [21,41,[60][61][62] and NanoBiT, [54,63] as well as by cell imaging techniques (e. g. phase-shift live cell imaging [64][65] ). Furthermore, other approaches, namely coimmunoprecipitation, [66] chemo-proteomics, [31] cellular thermal shift assay (CETSA), [48,53,67] in-cell NMR, [68] and FP [69] can be used to address target engagement in lysates or in cells.…”

PSL Chemical Biology Symposia Third Edition: A Branch of Science in its Explosive Phase

“…[62] According to Schreiber, we are currently only seeing the tip of the iceberg of this concept concerning bifunctional targeting chimeras (TACs) with the latest LYTACs [63] (using the lysosome degradation pathway), AbTACs [64] (using genetically encoded bispecific IgG antibody scaffolds to mediate membrane protein degradation), or (de)phosphorylation agents such as PhoRCs [65] or PHICs. [66] Recent efforts to accelerate the screening for binders uses the combination of diversity-oriented synthesis (DOS) and DNAencoded library (DEL) approaches to generate safe and effective therapeutics (library DOSEDO) [67] with tissue and substrate selectivity, representing new avenues for the development of next-generation therapeutics.…”