1. Structural basis of PROTAC cooperative recognition for selective protein degradation
Kwok-Ho Chan, Michael Zengerle, Xavier Lucas, Alessio Ciulli, Douglas J Lamont, Andrea Testa, Wenzhang Chen, Morgan S Gadd Nat Chem Biol . 2017 May;13(5):514-521. doi: 10.1038/nchembio.2329.
Inducing macromolecular interactions with small molecules to activate cellular signaling is a challenging goal. PROTACs (proteolysis-targeting chimeras) are bifunctional molecules that recruit a target protein in proximity to an E3 ubiquitin ligase to trigger protein degradation. Structural elucidation of the key ternary ligase-PROTAC-target species and its impact on target degradation selectivity remain elusive. We solved the crystal structure of Brd4 degrader MZ1 in complex with human VHL and the Brd4 bromodomain (Brd4BD2). The ligand folds into itself to allow formation of specific intermolecular interactions in the ternary complex. Isothermal titration calorimetry studies, supported by surface mutagenesis and proximity assays, are consistent with pronounced cooperative formation of ternary complexes with Brd4BD2. Structure-based-designed compound AT1 exhibits highly selective depletion of Brd4 in cells. Our results elucidate how PROTAC-induced de novo contacts dictate preferential recruitment of a target protein into a stable and cooperative complex with an E3 ligase for selective degradation.
2. TRIP12 promotes small-molecule-induced degradation through K29/K48-branched ubiquitin chains
Akinori Endo, Takuji Shoda, Yasuko Kawase, Yosuke Demizu, Mikihiko Naito, Yoshino Akizuki, Yasushi Saeki, Keiji Tanaka, Fumiaki Ohtake, Katsuhide Igarashi, Ai Kaiho-Soma Mol Cell . 2021 Apr 1;81(7):1411-1424.e7. doi: 10.1016/j.molcel.2021.01.023.
Targeted protein degradation is an emerging therapeutic paradigm. Small-molecule degraders such as proteolysis-targeting chimeras (PROTACs) induce the degradation of neo-substrates by hijacking E3 ubiquitin ligases. Although ubiquitylation of endogenous substrates has been extensively studied, the mechanism underlying forced degradation of neo-substrates is less well understood. We found that the ubiquitin ligase TRIP12 promotes PROTAC-induced and CRL2VHL-mediated degradation of BRD4 but is dispensable for the degradation of the endogenous CRL2VHLsubstrate HIF-1α. TRIP12 associates with BRD4 via CRL2VHLand specifically assembles K29-linked ubiquitin chains, facilitating the formation of K29/K48-branched ubiquitin chains and accelerating the assembly of K48 linkage by CRL2VHL. Consequently, TRIP12 promotes the PROTAC-induced apoptotic response. TRIP12 also supports the efficiency of other degraders that target CRABP2 or TRIM24 or recruit CRBN. These observations define TRIP12 and K29/K48-branched ubiquitin chains as accelerators of PROTAC-directed targeted protein degradation, revealing a cooperative mechanism of branched ubiquitin chain assembly unique to the degradation of neo-substrates.
3. Phenyl-Glutarimides: Alternative Cereblon Binders for the Design of PROTACs
Anand Mayasundari, Marcus Fischer, Zoran Rankovic, Brandon Young, Fatemeh Keramatnia, Stephen W White, Sergio Chai, Lei Yang, Patrick Ryan Potts, Martine F Roussel, Gisele Nishiguchi, Stanley Nithianantham, Sourav Das, Seung Wook Yang, Taosheng Chen, Zhenmei Li, Yong Li, Jamie Jarusiewicz, Mi-Kyung Yun, Jaeki Min, Anang Shelat, Jake Slavish, Barbara Jonchere, Marisa Actis, Richard E Lee, Shalandus H Garrett, Xiang Fu Angew Chem Int Ed Engl . 2021 Dec 13;60(51):26663-26670. doi: 10.1002/anie.202108848.
Targeting cereblon (CRBN) is currently one of the most frequently reported proteolysis-targeting chimera (PROTAC) approaches, owing to favorable drug-like properties of CRBN ligands, immunomodulatory imide drugs (IMiDs). However, IMiDs are known to be inherently unstable, readily undergoing hydrolysis in body fluids. Here we show that IMiDs and IMiD-based PROTACs rapidly hydrolyze in commonly utilized cell media, which significantly affects their cell efficacy. We designed novel CRBN binders, phenyl glutarimide (PG) analogues, and showed that they retained affinity for CRBN with high ligand efficiency (LE >0.48) and displayed improved chemical stability. Our efforts led to the discovery of PG PROTAC 4 c (SJ995973), a uniquely potent degrader of bromodomain and extra-terminal (BET) proteins that inhibited the viability of human acute myeloid leukemia MV4-11 cells at low picomolar concentrations (IC50=3 pM; BRD4 DC50=0.87 nM). These findings strongly support the utility of PG derivatives in the design of CRBN-directed PROTACs.
