Chemoproteomics-Enabled De Novo Proteolysis Targeting Chimera Discovery Platform Identifies a Metallothionein Degrader to Probe Its Role in Cancer

作者：Brittney Racioppo, Dany Pechalrieu, Daniel Abegg, Brendan G. Dwyer, Neal Ramseier, Ying Hu, Alexander Adibekian · 发表于：Journal of the American Chemical Society · 年份：2025 · DOI：10.1021/jacs.4c17827 · 被引用次数：14 · 研究领域：Protein Degradation and Inhibitors、Peptidase Inhibition and Analysis、Ubiquitin and proteasome pathways

Proteolysis targeting chimeras (PROTACs) represent powerful tools to modulate the activity of classically “undruggable” proteins, but their application has been limited to known ligands and a few select protein classes. Herein, we present our chemoproteomic strategy for simultaneous de novo discovery of novel degraders and ligands for challenging and previously “undruggable” targets. Using comparative PROTAC versus ligand global proteomics analyses, we rapidly identify proteins selectively downregulated by several “untargeted” PROTACs containing a VHL E3 ligase recruiter and various covalent and noncovalent ligands. We showcase our approach by identifying a first-in-class PROTAC for metallothionein 2A (MT2A), a small, cysteine-rich, metal-binding protein implicated in heavy metal detoxification, zinc homeostasis, and cellular invasion. Notably, isoform-specific MT overexpression has been shown to augment cellular migration and invasion across several cancer cell lines, although the precise mechanisms are unknown due to insufficient tools to study MTs. We show that optimized PROTAC AA-BR-157 covalently binds conserved C44, degrades overexpressed MT2A with nanomolar potency, and reduces the migration and invasion of MDA-MB-231 cells. We further demonstrate a time-dependent increase in intracellular zinc levels following MT2A degradation as well as downregulation of protein diaphanous homolog 3 (DIAPH3), a positive regulator of actin and cell motility. Super-resolution imaging o...