Automated Sample Preparation Coupled to LC-MS Enables Profiling of Molecular Degraders

Targeted Protein Degradation (TPD) is an emerging pharmaceutical modality that enables the modulation of proteins traditionally considered undruggable by conventional small molecules. TPD strategies, such as molecular glue degraders and proteolysis-targeting chimeras (PROTACs), hijack the cellular ubiquitin-proteasome system to selectively degrade pathogenic proteins.

High-content proteomics profiling is a powerful tool to identify targets and confirm degradation by quantifying protein levels across conditions. Here, we present a two-step workflow comprising of automated sample preparation for bottom-up proteomics and subsequent heavy-labeled reference peptide spike-in coupled to LC-MS acquisition. This workflow enables both global proteome profiling and targeted quantification of proteins of interest (POIs), facilitating comprehensive analysis of TPD compound effects. We demonstrate its utility by screening ten preclinical and clinical-stage molecular degraders targeting six disease-relevant proteins.