Structural basis for a Polθ helicase small-molecule inhibitor revealed by cryo-EM

  • Nat Commun. 2024 Aug 14;15(1):7003. doi: 10.1038/s41467-024-51351-4.
Fumiaki Ito  #  1 ,  Ziyuan Li  #  1 ,  Leonid Minakhin  2 ,  Gurushankar Chandramouly  2 ,  Mrityunjay Tyagi  2 ,  Robert Betsch  3 ,  John J Krais  3 ,  Bernadette Taberi  2 ,  Umeshkumar Vekariya  4 ,  Marissa Calbert  2 ,  Tomasz Skorski  4 ,  Neil Johnson  3 ,  Xiaojiang S Chen  5 ,  Richard T Pomerantz  6
Affiliations
  • 1. Molecular and Computational Biology, Department of Biological Sciences and Chemistry, University of Southern California, Los Angeles, California, CA, 90089, USA.
  • 2. Department of Biochemistry and Molecular Biology, Sidney Kimmel Cancer Center, Thomas Jefferson University, Philadelphia, PA, 19107, USA.
  • 3. Nuclear Dynamics Program, Fox Chase Cancer Center, Philadelphia, PA, 19111, USA.
  • 4. Fels Cancer Institute for Personalized Medicine, Lewis Katz School of Medicine, Temple University, Philadelphia, PA, 19140, USA.
  • 5. Molecular and Computational Biology, Department of Biological Sciences and Chemistry, University of Southern California, Los Angeles, California, CA, 90089, USA. [email protected].
  • 6. Department of Biochemistry and Molecular Biology, Sidney Kimmel Cancer Center, Thomas Jefferson University, Philadelphia, PA, 19107, USA. [email protected].
  • # Contributed equally.
PMID: 39143110 DOI: 10.1038/s41467-024-51351-4
Abstract

DNA Polymerase theta (Polθ) is a DNA helicase-polymerase protein that facilitates DNA repair and is synthetic lethal with homology-directed repair (HDR) factors. Thus, Polθ is a promising precision oncology drug-target in HDR-deficient cancers. Here, we characterize the binding and mechanism of action of a Polθ helicase (Polθ-hel) small-molecule inhibitor (AB25583) using cryo-EM. AB25583 exhibits 6 nM IC50 against Polθ-hel, selectively kills BRCA1/2-deficient cells, and acts synergistically with olaparib in Cancer cells harboring pathogenic BRCA1/2 mutations. Cryo-EM uncovers predominantly dimeric Polθ-hel:AB25583 complex structures at 3.0-3.2 Å. The structures reveal a binding-pocket deep inside the helicase central-channel, which underscores the high specificity and potency of AB25583. The cryo-EM structures in conjunction with biochemical data indicate that AB25583 inhibits the ATPase activity of Polθ-hel helicase via an allosteric mechanism. These detailed structural data and insights about AB25583 inhibition pave the way for accelerating drug development targeting Polθ-hel in HDR-deficient cancers.

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