Discovery of Tankyrase scaffolding inhibitor specifically targeting the ARC4 peptide binding domain

  1. Chiara Bosetti
  2. Albert Galera-Prat
  3. Sven T. Sowa
  4. Alexandra Gade
  5. Cláudia Braga
  6. Shoshy A. Brinch
  7. Faranak Nami
  8. Johan Pääkkönen
  9. Veeti Pulju
  10. Jo Waaler
  11. Mads H. Clausen
  12. Lari Lehtiö
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Abstract

In the past, development of tankyrase inhibitors has focused on the ADP-ribosyltransferase domain. Targeting tankyrases ability to interact with protein substrates through their ARC domains represents an alternative strategy to be explored as a therapeutic approach against specific protein-protein interactions. In this paper, we employed a FRET-based assay to identify ARC4-binding compounds by screening the EU-OPENSCREEN Pilot and Commercials Diversity libraries. We discovered an effective series of compounds with the same scaffold and through chemical synthesis we obtained the compoundS8(ARCher-142), which binds selectively to ARC4 with potency of 8 µM. NMR analysis and X-ray crystallography allowed us to identify the binding site in ARC4 and to rationalize the observed selectivity. Despite binding exclusively to ARC4, the inhibitor can attenuate the WNT/β-catenin signaling pathway in cells. Our work demonstrates that targeting single ARC domains is possible, offering an inhibition approach tailored to tankyrase ARC4 inhibition.

Significance

Tankyrases impact a variety of cellular processes by binding proteins through their ARC domains and the inhibition of these scaffolding functions represents an alternative therapeutic approach to catalytic inhibitors. With a FRET-based high-throughput screening of the EU-OPENSCREEN Pilot and Commercials Diversity libraries we discovered a pyrrolone-based scaffold that is interestingly selective towards ARC4, despite the high conservation of the ARC binding site. Our synthesized compoundS8(ARCher-142) displays an 8 µM potency for TNKS2 ARC4. With NMR and X-ray crystallography we demonstrate thatS8(ARCher-142) competes with the peptide optimized for binding and extends to a unique hydrophobic sub-pocket of ARC4. The compound attenuates the WNT/β-catenin signaling pathway in cells and interestingly offers the possibility to target specific protein-protein interactions mediated by ARC4, paving the way for the development of a pyrrolone-based class of tankyrase scaffolding inhibitors.

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