Improving PROTAC-Mediated Degradation in Mouse Models: BRD9 as a Case Study.

Proteolysis-targeting chimeras (PROTACs) have emerged as powerful chemical tools and therapeutics, yet their translation from preclinical models to humans remains inconsistent. Here, we systematically evaluate the activity of BRD9-targeting PROTACs in human and mouse systems to elucidate a key limitation in preclinical studies. Widely used CRBN- and VHL-recruiting degraders, including dBRD9 and VZ185, exhibit markedly reduced potency in mouse cells despite high sequence conservation of BRD9. Mechanistic interrogation reveals that species-specific differences in E3 ligases, particularly the Crbn I391 residue and structural variations in VHL, likely impair productive ternary complex formation. Expression of human CRBN or VHL, or introduction of the CrbnI391V mutation, increases degradation potency in mouse cells. Importantly, in vivo studies demonstrate enhanced BRD9 degradation in CrbnI391V mice using clinically relevant degraders. These findings highlight a critical barrier in PROTAC evaluation and establish humanized E3 ligase models as tools for more accurate assessment of on-target efficacy and toxicity.