Identification of actionable targeted protein degradation effector sites through Site-specific Ligand Incorporation-induced Proximity (SLIP)

ABSTRACT Targeted protein degradation (TPD) is a rapidly emerging and potentially transformative therapeutic modality. However, the large majority of >600 known ubiquitin ligases have yet to be exploited as TPD effectors by proteolysis-targeting chimeras (PROTACs) or molecular glue degraders (MGDs). We report here a chemical–genetic platform, Site-specific Ligand Incorporation-induced Proximity (SLIP), to identify actionable (“PROTACable”) sites on any potential effector protein in intact cells. SLIP uses genetic code expansion (GCE) to encode copper-free “click” ligation at a specific effector site in intact cells, enabling in situ formation of a covalent PROTAC-effector conjugate against a target protein of interest (POI). Modification at actionable effector sites drives degradation of the targeted protein, establishing the potential of these sites for TPD. Using SLIP, we systematically screened dozens of sites across E3 ligases and E2 enzymes from diverse classes, identifying multiple novel potentially PROTACable effector sites which are competent for TPD. SLIP adds a powerful approach to the proximity-induced pharmacology (PIP) toolbox, enabling future effector ligand discovery to fully enable TPD, and other emerging PIP modalities. Competing Interest Statement The authors declare the following competing financial interest(s): EWT is or has been employed as a consultant or scientific advisory board member for Myricx Pharma, Samsara Therapeutics, Roche, Novartis and Fastbase; research in his group has been funded by Pfizer Ltd, Kura Oncology, Daiichi Sankyo, Oxstem, Exscientia, Myricx Pharma, AstraZeneca, Vertex Pharmaceuticals, GSK and ADC Technologies. EWT holds equity in Myricx Pharma, Exactmer and Samsara Therapeutics, and is a named inventor on patents filed by Myricx Pharma, Exactmer, Imperial College London and the Francis Crick Institute. Other authors declare no competing interest.