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Title: Small molecule microarray identifies inhibitors of tyrosyl-DNA phosphodiesterase 1 that simultaneously access the catalytic pocket and two substrate binding sites

Abstract

Tyrosyl-DNA phosphodiesterase 1 (TDP1) is a member of the phospholipase D family of enzymes, which catalyzes the removal of both 3'- and 5'-DNA phosphodiester adducts. Importantly, it is capable of reducing the anticancer effects of type I topoisomerase (TOP1) inhibitors by repairing the stalled covalent complexes of TOP1 with DNA. It achieves this by promoting the hydrolysis of the phosphodiester bond between the Y723 residue of human TOP1 and the 3'-phosphate of its DNA substrate. Blocking TDP1 function is an attractive means of enhancing the efficacy of TOP1 inhibitors and overcoming drug resistance. Previously, we reported the use of an X-ray crystallographic screen of more than 600 fragments to identify small molecule variations on phthalic acid and hydroxyquinoline motifs that bind within the TDP1 catalytic pocket. Yet, the majority of these compounds showed limited (millimolar) TDP1 inhibitory potencies. We now report examining a 21 000-member library of drug-like Small Molecules in Microarray (SMM) format for their ability to bind Alexa Fluor 647 (AF647)-labeled TDP1. The screen identified structurally similar N,2-diphenylimidazo[1,2-a]pyrazin-3-amines as TDP1 binders and catalytic inhibitors. We then explored the core heterocycle skeleton using one-pot Groebke–Blackburn–Bienayme multicomponent reactions and arrived at analogs having higher inhibitory potencies. Solving TDP1 co-crystal structuresmore » of a subset of compounds showed their binding at the TDP1 catalytic site, while mimicking substrate interactions. Although our original fragment screen differed significantly from the current microarray protocol, both methods identified ligand–protein interactions containing highly similar elements. Importantly inhibitors identified through the SMM approach show competitive inhibition against TDP1 and access the catalytic phosphate-binding pocket, while simultaneously providing extensions into both the substrate DNA and peptide-binding channels. As such, they represent a platform for further elaboration of trivalent ligands, that could serve as a new genre of potent TDP1 inhibitors.« less

Authors:
ORCiD logo [1];  [2]; ORCiD logo [3];  [2];  [1];  [1];  [1]; ORCiD logo [1];  [1];  [2]; ORCiD logo [1]
  1. National Cancer Institute, Frederick, MD (United States)
  2. National Cancer Institute, Bethesda, MD (United States)
  3. Frederick National Lab. for Cancer Research, Frederick, MD (United States)
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Sponsoring Org.:
USDOE Office of Science (SC), Basic Energy Sciences (BES); Frederick National Laboratory for Cancer Research; National Institutes of Health (NIH); National Cancer Institute (NCI)
OSTI Identifier:
1781727
Grant/Contract Number:  
W-31-109-Eng-38; HHSN26120080001E; Z01-BC 006150; Z01-BC 006198
Resource Type:
Journal Article: Accepted Manuscript
Journal Name:
Chemical Science
Additional Journal Information:
Journal Volume: 12; Journal Issue: 11; Journal ID: ISSN 2041-6520
Publisher:
Royal Society of Chemistry
Country of Publication:
United States
Language:
ENGLISH
Subject:
37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY

Citation Formats

Zhao, Xue Zhi, Kiselev, Evgeny, Lountos, George T., Wang, Wenjie, Tropea, Joseph E., Needle, Danielle, Hilimire, Thomas A., Schneekloth, John S., Waugh, David S., Pommier, Yves, and Burke, Terrence R. Small molecule microarray identifies inhibitors of tyrosyl-DNA phosphodiesterase 1 that simultaneously access the catalytic pocket and two substrate binding sites. United States: N. p., 2021. Web. doi:10.1039/d0sc05411a.
Zhao, Xue Zhi, Kiselev, Evgeny, Lountos, George T., Wang, Wenjie, Tropea, Joseph E., Needle, Danielle, Hilimire, Thomas A., Schneekloth, John S., Waugh, David S., Pommier, Yves, & Burke, Terrence R. Small molecule microarray identifies inhibitors of tyrosyl-DNA phosphodiesterase 1 that simultaneously access the catalytic pocket and two substrate binding sites. United States. https://doi.org/10.1039/d0sc05411a
Zhao, Xue Zhi, Kiselev, Evgeny, Lountos, George T., Wang, Wenjie, Tropea, Joseph E., Needle, Danielle, Hilimire, Thomas A., Schneekloth, John S., Waugh, David S., Pommier, Yves, and Burke, Terrence R. 2021. "Small molecule microarray identifies inhibitors of tyrosyl-DNA phosphodiesterase 1 that simultaneously access the catalytic pocket and two substrate binding sites". United States. https://doi.org/10.1039/d0sc05411a. https://www.osti.gov/servlets/purl/1781727.
@article{osti_1781727,
title = {Small molecule microarray identifies inhibitors of tyrosyl-DNA phosphodiesterase 1 that simultaneously access the catalytic pocket and two substrate binding sites},
author = {Zhao, Xue Zhi and Kiselev, Evgeny and Lountos, George T. and Wang, Wenjie and Tropea, Joseph E. and Needle, Danielle and Hilimire, Thomas A. and Schneekloth, John S. and Waugh, David S. and Pommier, Yves and Burke, Terrence R.},
abstractNote = {Tyrosyl-DNA phosphodiesterase 1 (TDP1) is a member of the phospholipase D family of enzymes, which catalyzes the removal of both 3'- and 5'-DNA phosphodiester adducts. Importantly, it is capable of reducing the anticancer effects of type I topoisomerase (TOP1) inhibitors by repairing the stalled covalent complexes of TOP1 with DNA. It achieves this by promoting the hydrolysis of the phosphodiester bond between the Y723 residue of human TOP1 and the 3'-phosphate of its DNA substrate. Blocking TDP1 function is an attractive means of enhancing the efficacy of TOP1 inhibitors and overcoming drug resistance. Previously, we reported the use of an X-ray crystallographic screen of more than 600 fragments to identify small molecule variations on phthalic acid and hydroxyquinoline motifs that bind within the TDP1 catalytic pocket. Yet, the majority of these compounds showed limited (millimolar) TDP1 inhibitory potencies. We now report examining a 21 000-member library of drug-like Small Molecules in Microarray (SMM) format for their ability to bind Alexa Fluor 647 (AF647)-labeled TDP1. The screen identified structurally similar N,2-diphenylimidazo[1,2-a]pyrazin-3-amines as TDP1 binders and catalytic inhibitors. We then explored the core heterocycle skeleton using one-pot Groebke–Blackburn–Bienayme multicomponent reactions and arrived at analogs having higher inhibitory potencies. Solving TDP1 co-crystal structures of a subset of compounds showed their binding at the TDP1 catalytic site, while mimicking substrate interactions. Although our original fragment screen differed significantly from the current microarray protocol, both methods identified ligand–protein interactions containing highly similar elements. Importantly inhibitors identified through the SMM approach show competitive inhibition against TDP1 and access the catalytic phosphate-binding pocket, while simultaneously providing extensions into both the substrate DNA and peptide-binding channels. As such, they represent a platform for further elaboration of trivalent ligands, that could serve as a new genre of potent TDP1 inhibitors.},
doi = {10.1039/d0sc05411a},
url = {https://www.osti.gov/biblio/1781727}, journal = {Chemical Science},
issn = {2041-6520},
number = 11,
volume = 12,
place = {United States},
year = {Thu Jan 28 00:00:00 EST 2021},
month = {Thu Jan 28 00:00:00 EST 2021}
}

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