Oncogene activation mechanism determines the limits of targeted protein degradation.
Protein degrader drugs such as PROTACs are being advanced as therapeutics targeted against oncogenic proteins. During tumorigenesis, oncogenic proteins can become constitutively activated via mechanisms including gene amplification, which increases protein production, and point mutations, which can extend protein half-life. Few experimental studies have addressed how disease-associated changes in target protein homeostasis influence PROTAC activity. We developed orthogonal methods to increase production or enhance stability of β-catenin, an important oncoprotein and target for degrader therapeutics, and used the dTAG system to evaluate the consequences for PROTAC activity. Stabilizing oncogenic missense mutations increase protein expression up to 5-fold but do not alter the PROTAC-imposed minimal steady-state level. In contrast, transcriptional upregulation increases both pre- and post-treatment target levels, revealing a synthesis-dependent ceiling on achievable depletion. Our results highlight distinct constraints on PROTAC activity arising from different mechanisms of oncogene activation, with potential implications for preclinical modeling, drug resistance and personalized medicine.