Related Experiment Video

Updated: Jun 15, 2025

Measuring Enzymatic Activity of Neurodevelopmental Disorder-Associated Deubiquitylating Enzymes via an In Vitro Ubiquitin Chain Cleavage Assay
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Measuring Enzymatic Activity of Neurodevelopmental Disorder-Associated Deubiquitylating Enzymes via an In Vitro Ubiquitin Chain Cleavage Assay

Published on: September 27, 2024

422

Structural and Biochemical Insights into the Mechanism of Action of the Clinical USP1 Inhibitor, KSQ-4279

Martin Luke Rennie1, Mehmet Gundogdu2, Connor Arkinson1

  • 1School of Molecular Biosciences, College of Medical Veterinary and Life Sciences, University of Glasgow, Glasgow G12 8QQ, U.K.

Journal of Medicinal Chemistry
|August 27, 2024
PubMed

Insights

This study reveals how KSQ-4279, a clinical ubiquitin-specific protease 1 (USP1) inhibitor, binds to USP1. The findings offer molecular insights into USP1 inhibitor mechanisms for cancer therapy development.

Area of Science:

  • Biochemistry
  • Structural Biology
  • Cancer Biology

Background:

  • DNA damage response pathways are crucial for cellular integrity.
  • Dysregulation of DNA repair signaling is a hallmark of cancer.
  • Ubiquitin-specific protease 1 (USP1) is a key regulator in DNA repair and a therapeutic target in oncology.

Purpose of the Study:

  • To elucidate the molecular mechanism of action of the clinical USP1 inhibitor KSQ-4279.
  • To compare the binding and structural effects of KSQ-4279 with the known inhibitor ML323.
  • To understand the structural basis for USP1 inhibition by small molecules.

Main Methods:

  • Biochemical assays to assess enzyme activity and inhibitor binding.
  • Cryo-electron microscopy (cryo-EM) for high-resolution structural determination.
  • Thermal stability assays to evaluate protein conformational changes upon inhibitor binding.

Main Results:

  • KSQ-4279 binds to the same cryptic site on USP1 as ML323.
  • Inhibitor binding induces distinct structural perturbations in USP1 compared to ML323.
  • Binding of KSQ-4279 significantly increases USP1's thermal stability, potentially via filling a hydrophobic pocket.

Conclusions:

  • The study provides a detailed molecular understanding of KSQ-4279's interaction with USP1.
  • Subtle differences in inhibitor-induced structural changes may impact USP1 inhibitor efficacy.
  • These findings advance the rational design of USP1-targeting cancer therapeutics.

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