Cargando…
Lucanthone and Its Derivative Hycanthone Inhibit Apurinic Endonuclease-1 (APE1) by Direct Protein Binding
Lucanthone and hycanthone are thioxanthenone DNA intercalators used in the 1980s as antitumor agents. Lucanthone is in Phase I clinical trial, whereas hycanthone was pulled out of Phase II trials. Their potential mechanism of action includes DNA intercalation, inhibition of nucleic acid biosyntheses...
Autores principales: | , , , , , , , , , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Public Library of Science
2011
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3174134/ https://www.ncbi.nlm.nih.gov/pubmed/21935361 http://dx.doi.org/10.1371/journal.pone.0023679 |
_version_ | 1782212032885424128 |
---|---|
author | Naidu, Mamta D. Agarwal, Rakhi Pena, Louis A. Cunha, Luis Mezei, Mihaly Shen, Min Wilson, David M. Liu, Yuan Sanchez, Zina Chaudhary, Pankaj Wilson, Samuel H. Waring, Michael J. |
author_facet | Naidu, Mamta D. Agarwal, Rakhi Pena, Louis A. Cunha, Luis Mezei, Mihaly Shen, Min Wilson, David M. Liu, Yuan Sanchez, Zina Chaudhary, Pankaj Wilson, Samuel H. Waring, Michael J. |
author_sort | Naidu, Mamta D. |
collection | PubMed |
description | Lucanthone and hycanthone are thioxanthenone DNA intercalators used in the 1980s as antitumor agents. Lucanthone is in Phase I clinical trial, whereas hycanthone was pulled out of Phase II trials. Their potential mechanism of action includes DNA intercalation, inhibition of nucleic acid biosyntheses, and inhibition of enzymes like topoisomerases and the dual function base excision repair enzyme apurinic endonuclease 1 (APE1). Lucanthone inhibits the endonuclease activity of APE1, without affecting its redox activity. Our goal was to decipher the precise mechanism of APE1 inhibition as a prerequisite towards development of improved therapeutics that can counteract higher APE1 activity often seen in tumors. The IC(50) values for inhibition of APE1 incision of depurinated plasmid DNA by lucanthone and hycanthone were 5 µM and 80 nM, respectively. The K(D) values (affinity constants) for APE1, as determined by BIACORE binding studies, were 89 nM for lucanthone/10 nM for hycanthone. APE1 structures reveal a hydrophobic pocket where hydrophobic small molecules like thioxanthenones can bind, and our modeling studies confirmed such docking. Circular dichroism spectra uncovered change in the helical structure of APE1 in the presence of lucanthone/hycanthone, and notably, this effect was decreased (Phe266Ala or Phe266Cys or Trp280Leu) or abolished (Phe266Ala/Trp280Ala) when hydrophobic site mutants were employed. Reduced inhibition by lucanthone of the diminished endonuclease activity of hydrophobic mutant proteins (as compared to wild type APE1) supports that binding of lucanthone to the hydrophobic pocket dictates APE1 inhibition. The DNA binding capacity of APE1 was marginally inhibited by lucanthone, and not at all by hycanthone, supporting our hypothesis that thioxanthenones inhibit APE1, predominantly, by direct interaction. Finally, lucanthone-induced degradation was drastically reduced in the presence of short and long lived free radical scavengers, e.g., TRIS and DMSO, suggesting that the mechanism of APE1 breakdown may involve free radical-induced peptide bond cleavage. |
format | Online Article Text |
id | pubmed-3174134 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2011 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-31741342011-09-20 Lucanthone and Its Derivative Hycanthone Inhibit Apurinic Endonuclease-1 (APE1) by Direct Protein Binding Naidu, Mamta D. Agarwal, Rakhi Pena, Louis A. Cunha, Luis Mezei, Mihaly Shen, Min Wilson, David M. Liu, Yuan Sanchez, Zina Chaudhary, Pankaj Wilson, Samuel H. Waring, Michael J. PLoS One Research Article Lucanthone and hycanthone are thioxanthenone DNA intercalators used in the 1980s as antitumor agents. Lucanthone is in Phase I clinical trial, whereas hycanthone was pulled out of Phase II trials. Their potential mechanism of action includes DNA intercalation, inhibition of nucleic acid biosyntheses, and inhibition of enzymes like topoisomerases and the dual function base excision repair enzyme apurinic endonuclease 1 (APE1). Lucanthone inhibits the endonuclease activity of APE1, without affecting its redox activity. Our goal was to decipher the precise mechanism of APE1 inhibition as a prerequisite towards development of improved therapeutics that can counteract higher APE1 activity often seen in tumors. The IC(50) values for inhibition of APE1 incision of depurinated plasmid DNA by lucanthone and hycanthone were 5 µM and 80 nM, respectively. The K(D) values (affinity constants) for APE1, as determined by BIACORE binding studies, were 89 nM for lucanthone/10 nM for hycanthone. APE1 structures reveal a hydrophobic pocket where hydrophobic small molecules like thioxanthenones can bind, and our modeling studies confirmed such docking. Circular dichroism spectra uncovered change in the helical structure of APE1 in the presence of lucanthone/hycanthone, and notably, this effect was decreased (Phe266Ala or Phe266Cys or Trp280Leu) or abolished (Phe266Ala/Trp280Ala) when hydrophobic site mutants were employed. Reduced inhibition by lucanthone of the diminished endonuclease activity of hydrophobic mutant proteins (as compared to wild type APE1) supports that binding of lucanthone to the hydrophobic pocket dictates APE1 inhibition. The DNA binding capacity of APE1 was marginally inhibited by lucanthone, and not at all by hycanthone, supporting our hypothesis that thioxanthenones inhibit APE1, predominantly, by direct interaction. Finally, lucanthone-induced degradation was drastically reduced in the presence of short and long lived free radical scavengers, e.g., TRIS and DMSO, suggesting that the mechanism of APE1 breakdown may involve free radical-induced peptide bond cleavage. Public Library of Science 2011-09-15 /pmc/articles/PMC3174134/ /pubmed/21935361 http://dx.doi.org/10.1371/journal.pone.0023679 Text en This is an open-access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication. https://creativecommons.org/publicdomain/zero/1.0/ This is an open-access article distributed under the terms of the Creative Commons Public Domain declaration, which stipulates that, once placed in the public domain, this work may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. |
spellingShingle | Research Article Naidu, Mamta D. Agarwal, Rakhi Pena, Louis A. Cunha, Luis Mezei, Mihaly Shen, Min Wilson, David M. Liu, Yuan Sanchez, Zina Chaudhary, Pankaj Wilson, Samuel H. Waring, Michael J. Lucanthone and Its Derivative Hycanthone Inhibit Apurinic Endonuclease-1 (APE1) by Direct Protein Binding |
title | Lucanthone and Its Derivative Hycanthone Inhibit Apurinic Endonuclease-1 (APE1) by Direct Protein Binding |
title_full | Lucanthone and Its Derivative Hycanthone Inhibit Apurinic Endonuclease-1 (APE1) by Direct Protein Binding |
title_fullStr | Lucanthone and Its Derivative Hycanthone Inhibit Apurinic Endonuclease-1 (APE1) by Direct Protein Binding |
title_full_unstemmed | Lucanthone and Its Derivative Hycanthone Inhibit Apurinic Endonuclease-1 (APE1) by Direct Protein Binding |
title_short | Lucanthone and Its Derivative Hycanthone Inhibit Apurinic Endonuclease-1 (APE1) by Direct Protein Binding |
title_sort | lucanthone and its derivative hycanthone inhibit apurinic endonuclease-1 (ape1) by direct protein binding |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3174134/ https://www.ncbi.nlm.nih.gov/pubmed/21935361 http://dx.doi.org/10.1371/journal.pone.0023679 |
work_keys_str_mv | AT naidumamtad lucanthoneanditsderivativehycanthoneinhibitapurinicendonuclease1ape1bydirectproteinbinding AT agarwalrakhi lucanthoneanditsderivativehycanthoneinhibitapurinicendonuclease1ape1bydirectproteinbinding AT penalouisa lucanthoneanditsderivativehycanthoneinhibitapurinicendonuclease1ape1bydirectproteinbinding AT cunhaluis lucanthoneanditsderivativehycanthoneinhibitapurinicendonuclease1ape1bydirectproteinbinding AT mezeimihaly lucanthoneanditsderivativehycanthoneinhibitapurinicendonuclease1ape1bydirectproteinbinding AT shenmin lucanthoneanditsderivativehycanthoneinhibitapurinicendonuclease1ape1bydirectproteinbinding AT wilsondavidm lucanthoneanditsderivativehycanthoneinhibitapurinicendonuclease1ape1bydirectproteinbinding AT liuyuan lucanthoneanditsderivativehycanthoneinhibitapurinicendonuclease1ape1bydirectproteinbinding AT sanchezzina lucanthoneanditsderivativehycanthoneinhibitapurinicendonuclease1ape1bydirectproteinbinding AT chaudharypankaj lucanthoneanditsderivativehycanthoneinhibitapurinicendonuclease1ape1bydirectproteinbinding AT wilsonsamuelh lucanthoneanditsderivativehycanthoneinhibitapurinicendonuclease1ape1bydirectproteinbinding AT waringmichaelj lucanthoneanditsderivativehycanthoneinhibitapurinicendonuclease1ape1bydirectproteinbinding |