biostudies-literatureUnknown16(1)Steger MMolecular glue degraders (MGDs) are small molecules that co-opt the ubiquitin-proteasome system to induce degradation of target proteins, including those considered undruggable. Their discovery remains challenging due to the lack of rational design strategies and limited throughput of unbiased proteome-wide screening approaches. To address this gap, we develop a high-throughput proteomics platform based on label-free, data-independent acquisition mass spectrometry (DIA-MS), enabling integrated proteomics and ubiquitinomics profiling. Screening a diverse set of 100 cereblon (CRBN)-recruiting ligands on this platform leads to identification of a broad array of novel degraders and neosubstrates. Subsequent hit validation and structure-degradation relationship analyses guided by global proteomNature communications7773https://www.ebi.ac.uk/biostudies/studies/S-EPMC12368047biostudies-literatureUnbiased mapping of cereblon neosubstrate landscape by high-throughput proteomics.PMC12368047Demichev VPrice JGraef TYang JNishiguchi GYang LDaub HWu QBartoschek DShi ZRankovic ZMayasundari AOhmayer UActis MSteger MAggarwal AMcGowan KSchwalb BShashikadze BBednarz AHMachata SfalseUnbiased mapping of cereblon neosubstrate landscape by high-throughput proteomics.Molecular glue degraders (MGDs) are small molecules that co-opt the ubiquitin-proteasome system to induce degradation of target proteins, including those considered undruggable. Their discovery remains challenging due to the lack of rational design strategies and limited throughput of unbiased proteome-wide screening approaches. To address this gap, we develop a high-throughput proteomics platform based on label-free, data-independent acquisition mass spectrometry (DIA-MS), enabling integrated proteomics and ubiquitinomics profiling. Screening a diverse set of 100 cereblon (CRBN)-recruiting ligands on this platform leads to identification of a broad array of novel degraders and neosubstrates. Subsequent hit validation and structure-degradation relationship analyses guided by global proteom2025-01-01T00:00:00Z2025 Aug2026-05-05T19:03:57.68Z2026-04-07T21:51:08.908ZS-EPMC123680474083582510.1038/s41467-025-62829-0