Therapeutic potential of allosteric HECT E3 ligase inhibition.

Journal: Cell
Published Date: Apr 2, 2025

Abstract

Targeting ubiquitin E3 ligases is therapeutically attractive; however, the absence of an active-site pocket impedes computational approaches for identifying inhibitors. In a large, unbiased biochemical screen, we discover inhibitors that bind a cryptic cavity distant from the catalytic cysteine of the homologous to E6-associated protein C terminus domain (HECT) E3 ligase, SMAD ubiquitin regulatory factor 1 (SMURF1). Structural and biochemical analyses and engineered escape mutants revealed that these inhibitors restrict an essential catalytic motion by extending an α helix over a conserved glycine hinge. SMURF1 levels are increased in pulmonary arterial hypertension (PAH), a disease caused by mutation of bone morphogenetic protein receptor-2 (BMPR2). We demonstrated that SMURF1 inhibition prevented BMPR2 ubiquitylation, normalized bone morphogenetic protein (BMP) signaling, restored pulmonary vascular cell homeostasis, and reversed pathology in established experimental PAH. Leveraging this deep mechanistic understanding, we undertook an in silico machine-learning-based screen to identify inhibitors of the prototypic HECT E6AP and confirmed glycine-hinge-dependent allosteric activity in vitro. Inhibiting HECTs and other glycine-hinge proteins opens a new druggable space.

Authors

  • Alexander M K Rothman
    University of Sheffield, Sheffield, UK; Novartis BioMedical Research (NBR), Cambridge, MA, USA. Electronic address: a.rothman@sheffield.ac.uk.
  • Amir Florentin
    School of Neurobiology, Biochemistry and Biophysics, The Life Sciences Faculty, Tel Aviv University, Tel Aviv, Israel.
  • Florence Zink
    Novartis BioMedical Research (NBR), Basel, Switzerland.
  • Catherine Quigley
    Novartis BioMedical Research (NBR), Cambridge, MA, USA.
  • Olivier Bonneau
    Novartis BioMedical Research (NBR), Cambridge, MA, USA.
  • Rene Hemmig
    Novartis BioMedical Research (NBR), Basel, Switzerland.
  • Amanda Hachey
    Novartis BioMedical Research (NBR), Cambridge, MA, USA.
  • Tomas Rejtar
    Novartis BioMedical Research (NBR), Cambridge, MA, USA.
  • Maulik Thaker
    Novartis BioMedical Research (NBR), Cambridge, MA, USA.
  • Rishi Jain
    Novartis BioMedical Research (NBR), Cambridge, MA, USA.
  • Shih-Min Huang
    Novartis BioMedical Research (NBR), Cambridge, MA, USA.
  • Daniel Sutton
    Imaging and AI, Clinical Pharmacology and Safety Sciences, BioPharmaceuticals R&D, AstraZeneca, Cambridge CB4 0WG, U.K.
  • Jan Roger
    Novartis Institutes for BioMedical Research (NIBR), Horsham, UK.
  • Ji-Hu Zhang
    Novartis BioMedical Research (NBR), Cambridge, MA, USA.
  • Sven Weiler
    Novartis BioMedical Research (NBR), Cambridge, MA, USA.
  • Simona Cotesta
    Novartis BioMedical Research (NBR), Basel, Switzerland.
  • Johannes Ottl
    Novartis BioMedical Research (NBR), Basel, Switzerland.
  • Salil Srivastava
    Novartis BioMedical Research (NBR), Cambridge, MA, USA.
  • Alina Kordonsky
    School of Neurobiology, Biochemistry and Biophysics, The Life Sciences Faculty, Tel Aviv University, Tel Aviv, Israel.
  • Reut Avishid
    School of Neurobiology, Biochemistry and Biophysics, The Life Sciences Faculty, Tel Aviv University, Tel Aviv, Israel.
  • Elon Yariv
    School of Neurobiology, Biochemistry and Biophysics, The Life Sciences Faculty, Tel Aviv University, Tel Aviv, Israel.
  • Ritu Rathi
    School of Neurobiology, Biochemistry and Biophysics, The Life Sciences Faculty, Tel Aviv University, Tel Aviv, Israel.
  • Oshrit Khvalevsky
    School of Neurobiology, Biochemistry and Biophysics, The Life Sciences Faculty, Tel Aviv University, Tel Aviv, Israel.
  • Thomas Troxler
    School of Neurobiology, Biochemistry and Biophysics, The Life Sciences Faculty, Tel Aviv University, Tel Aviv, Israel; Novartis BioMedical Research (NBR), Basel, Switzerland.
  • Sarah K Binmahfooz
    University of Sheffield, Sheffield, UK; Department of Clinical Biochemistry, Faculty of Medicine, King Abdulaziz University, Jeddah, Saudi Arabia.
  • Oded Kleifeld
    Faculty of Biology, Technion-Israel Institute of Technology, Haifa 32000, Israel.
  • Nicholas W Morrell
    Department of Medicine, School of Clinical Medicine, University of Cambridge, Cambridge, UK.
  • Marc Humbert
    Université Paris-Saclay, INSERM UMR_S 999 (HPPIT), Service de Pneumologie et Soins Intensifs Respiratoires, Hôpital Bicêtre (Assistance Publique Hôpitaux de Paris), Le Kremlin Bicêtre, France.
  • Matthew J Thomas
    Novartis Institutes for BioMedical Research (NIBR), Horsham, UK.
  • Gabor Jarai
    Novartis Institutes for BioMedical Research (NIBR), Horsham, UK.
  • Rohan E J Beckwith
    Novartis BioMedical Research (NBR), Cambridge, MA, USA.
  • Jennifer S Cobb
    Novartis BioMedical Research (NBR), Cambridge, MA, USA.
  • Nichola Smith
    Novartis BioMedical Research (NBR), Cambridge, MA, USA.
  • Nils Ostermann
    Novartis BioMedical Research (NBR), Basel, Switzerland.
  • John Tallarico
    Novartis BioMedical Research (NBR), Cambridge, MA, USA.
  • Duncan Shaw
    Novartis BioMedical Research (NBR), Cambridge, MA, USA.
  • Sabine Guth-Gundel
    Novartis BioMedical Research (NBR), Basel, Switzerland.
  • Gali Prag
    School of Neurobiology, Biochemistry and Biophysics, The Life Sciences Faculty, Tel Aviv University, Tel Aviv, Israel; Sagol School of Neuroscience, Tel Aviv University, Tel Aviv, Israel. Electronic address: prag@tauex.tau.ac.il.
  • David J Rowlands
    Novartis BioMedical Research (NBR), Cambridge, MA, USA. Electronic address: davidjrowlands8@gmail.com.

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