Discovery of 2,4-Quinazolinedione Derivatives as LC3B Recruiters in the Facilitation of Protein Complex Degradations

Targeted protein degradation through autophagosome-tethering compounds (ATTECs) that bypasses the ubiquitination process has garnered increasing attention. LC3B, a key protein in autophagosome formation, recruits substrates into the autophagy-lysosome system for degradation. In this study, we systematically optimized 2,4-quinazolinedione derivatives as LC3B-recruiting fragments, utilizing the CDK9 indicator. By attaching the designed LC3B-recruiting fragment to CDK9 inhibitor SNS-032 through a linker, the resulting bifunctional ATTEC molecule simultaneously degraded CDK9 and its associated Cyclin T1. Two-dimensional NMR experiments confirmed the direct interaction between the novel LC3B-recruiting fragments and LC3B. Mechanistic studies elucidated that degradation occurred via an LC3B-dependent autophagy-lysosomal pathway. Additionally, the general applicability of leveraging LC3B-recruiting fragments linked to inhibitors for the targeted degradation of protein complexes was validated with PRC2 and CDK2/4/6 along with their respective Cyclins. This work provides a series of novel LC3B-recruiting fragments that enrich the ATTEC toolbox and can be applied to the degradation of diverse intracellular disease-causing proteins. Based on the reported LC3B binding compound 8F20, we simplified its structure and did systematic structure activity relationship exploration. The obtained fragments 33R and 34R were demonstrated to bind with LC3B directly in 2D NMR titration assay. The novel LC3B-recruiting fragments were developed as a generally applicable degradation platform capable of targeting diverse cellular protein complexes. [Display omitted] •Optimized the structure of the LC3B binding compounds.•The obtained fragments were demonstrated to bind with LC3B directly.•The fragments were developed as a generally applicable degradation platform.•The capability of targeting diverse intracellular protein complexes.