Graduate Student Sophie Rizzo

"Structure-Guided Design of Targeted Protein Degraders: PROTACs"

Proteolysis-targeting chimeras (PROTACs) represent a major class of molecules that can therapeutically modulate protein levels through targeted protein degradation. PROTACs exploit the protein degradation machinery by bringing together the targeted protein with an E3 ubiquitin ligase. This results in the protein of interest to be tagged with multiple ubiquitin molecules and recognized for degradation by the 26S proteasome. Unlike protein inhibitors and other occupancy-based strategies, PROTAC agents can target multiple equivalents of targeted proteins, allowing it to function catalytically at sub-stoichiometric levels.

The design of PROTAC molecules is predominantly empirical, although data supports that the formation and stabilization of a key intermediate species drives the efficacy of these molecules. I will discuss the first crystal structure of ternary complexed of the MZ1 PROTAC bound to von Hippel-Lindau (VHL) E3 ligase and a member of the bromodomain-containing protein family (Brd4). Interestingly, the PROTAC takes on a sandwiched conformation formed by many stabilizing protein-protein interactions between Brd4 and the E3 ligase. Mutagenesis studies, isothermal titration calorimetry, and proximity assays confirm that these protein-protein interactions dictate how cooperative and stable ternary complex formation is within the bromodomain-containing protein family. These insights guided the structure-based design of a more powerful and selective Brd4 degrader and the use of macrocyclization to conformationally lock the PROTAC in its bound state, thereby lessening the presence of uncooperative ternary complex formation. The resulting macroPROTAC-1 agent exhibit a similar degradation profile and cellular activity as the parent linear agent, despite a 12-fold loss in binary binding affinity. Both of these examples demonstrate the use of structural-based design strategies in order to generate more potent and selective PROTACs.

(1) Gadd, M.S., Testa, A., Lucas, X., Chan, K.H., Chen, W., Lamont, D.J., Zengerle, M. and Ciulli, A. Structural basis of PROTAC cooperative recognition for selective protein degradation Nature chemical biology 2017, 13(5), 514-521.
(2) Testa, A., Hughes, S. J., Lucas, X., Wright, J. E., & Ciulli, A. Structure‐based design of a macrocyclic PROTAC. Angewandte Chemie 2020, 132(4), 1744-1751.
(3) Békés, M., Langley, D. R., & Crews, C. M. PROTAC targeted protein degraders: the past is prologue. Nature Reviews Drug Discovery 2022, 1-20.