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A quantitative LumiFluo assay to test inhibitory compounds blocking p53 degradation induced by human papillomavirus oncoprotein E6 in living cells

Lorenzo Messa, Marta Celegato, Chiara Bertagnin, Beatrice Mercorelli, Giulio Nannetti Orcid Logo, Giorgio Palù, Arianna Loregian

Scientific Reports, Volume: 8, Issue: 1

Swansea University Author: Giulio Nannetti Orcid Logo

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DOI (Published version): 10.1038/s41598-018-24470-4

Abstract

High-risk human papillomaviruses (HR-HPVs) are the causative agents for the onset of several epithelial cancers in humans. The deregulated expression of the viral oncoproteins E6 and E7 is the driving force sustaining the progression of malignant transformation in pre-neoplastic lesions. Targeting t...

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Published in: Scientific Reports
ISSN: 2045-2322
Published: Springer Science and Business Media LLC 2018
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URI: https://cronfa.swan.ac.uk/Record/cronfa57637
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spelling 2021-09-14T15:15:36.6129227 v2 57637 2021-08-18 A quantitative LumiFluo assay to test inhibitory compounds blocking p53 degradation induced by human papillomavirus oncoprotein E6 in living cells 8d6b68ac6d8a7ab60eb04cae67116565 0000-0003-3227-1537 Giulio Nannetti Giulio Nannetti true false 2021-08-18 PHAR High-risk human papillomaviruses (HR-HPVs) are the causative agents for the onset of several epithelial cancers in humans. The deregulated expression of the viral oncoproteins E6 and E7 is the driving force sustaining the progression of malignant transformation in pre-neoplastic lesions. Targeting the viral E6 oncoprotein through inhibitory compounds can counteract the survival of cancer cells due to the reactivation of p53-mediated pathways and represents an intriguing strategy to treat HPV-associated neoplasias. Here, we describe the development of a quantitative and easy-to-perform assay to monitor the E6-mediated degradation of p53 in living cells to be used for small-molecule testing. This assay allows to unbiasedly determine whether a compound can protect p53 from the E6-mediated degradation in cells, through a simple 3-step protocol. We validated the assay by testing two small molecules, SAHA and RITA, reported to impair the E6-mediated p53 degradation. Interestingly, we observed that only SAHA efficiently rescued p53, while RITA could not provide the same degree of protection. The possibility to specifically and quantitatively monitor the ability of a selected compound to rescue p53 in a cellular context through our LumiFluo assay could represent an important step towards the successful development of anti-HPV drugs. Journal Article Scientific Reports 8 1 Springer Science and Business Media LLC 2045-2322 16 4 2018 2018-04-16 10.1038/s41598-018-24470-4 COLLEGE NANME Pharmacy COLLEGE CODE PHAR Swansea University 2021-09-14T15:15:36.6129227 2021-08-18T13:35:18.2709804 Faculty of Medicine, Health and Life Sciences Swansea University Medical School - Medicine Lorenzo Messa 1 Marta Celegato 2 Chiara Bertagnin 3 Beatrice Mercorelli 4 Giulio Nannetti 0000-0003-3227-1537 5 Giorgio Palù 6 Arianna Loregian 7 57637__20845__614cda263bff41f8891e12468bde713c.pdf 57637.pdf 2021-09-14T14:38:57.1549789 Output 2180400 application/pdf Version of Record true © The Author(s) 2018. This article is licensed under a Creative Commons Attribution 4.0 International License true eng http://creativecommons.org/licenses/by/4.0/
title A quantitative LumiFluo assay to test inhibitory compounds blocking p53 degradation induced by human papillomavirus oncoprotein E6 in living cells
spellingShingle A quantitative LumiFluo assay to test inhibitory compounds blocking p53 degradation induced by human papillomavirus oncoprotein E6 in living cells
Giulio Nannetti
title_short A quantitative LumiFluo assay to test inhibitory compounds blocking p53 degradation induced by human papillomavirus oncoprotein E6 in living cells
title_full A quantitative LumiFluo assay to test inhibitory compounds blocking p53 degradation induced by human papillomavirus oncoprotein E6 in living cells
title_fullStr A quantitative LumiFluo assay to test inhibitory compounds blocking p53 degradation induced by human papillomavirus oncoprotein E6 in living cells
title_full_unstemmed A quantitative LumiFluo assay to test inhibitory compounds blocking p53 degradation induced by human papillomavirus oncoprotein E6 in living cells
title_sort A quantitative LumiFluo assay to test inhibitory compounds blocking p53 degradation induced by human papillomavirus oncoprotein E6 in living cells
author_id_str_mv 8d6b68ac6d8a7ab60eb04cae67116565
author_id_fullname_str_mv 8d6b68ac6d8a7ab60eb04cae67116565_***_Giulio Nannetti
author Giulio Nannetti
author2 Lorenzo Messa
Marta Celegato
Chiara Bertagnin
Beatrice Mercorelli
Giulio Nannetti
Giorgio Palù
Arianna Loregian
format Journal article
container_title Scientific Reports
container_volume 8
container_issue 1
publishDate 2018
institution Swansea University
issn 2045-2322
doi_str_mv 10.1038/s41598-018-24470-4
publisher Springer Science and Business Media LLC
college_str Faculty of Medicine, Health and Life Sciences
hierarchytype
hierarchy_top_id facultyofmedicinehealthandlifesciences
hierarchy_top_title Faculty of Medicine, Health and Life Sciences
hierarchy_parent_id facultyofmedicinehealthandlifesciences
hierarchy_parent_title Faculty of Medicine, Health and Life Sciences
department_str Swansea University Medical School - Medicine{{{_:::_}}}Faculty of Medicine, Health and Life Sciences{{{_:::_}}}Swansea University Medical School - Medicine
document_store_str 1
active_str 0
description High-risk human papillomaviruses (HR-HPVs) are the causative agents for the onset of several epithelial cancers in humans. The deregulated expression of the viral oncoproteins E6 and E7 is the driving force sustaining the progression of malignant transformation in pre-neoplastic lesions. Targeting the viral E6 oncoprotein through inhibitory compounds can counteract the survival of cancer cells due to the reactivation of p53-mediated pathways and represents an intriguing strategy to treat HPV-associated neoplasias. Here, we describe the development of a quantitative and easy-to-perform assay to monitor the E6-mediated degradation of p53 in living cells to be used for small-molecule testing. This assay allows to unbiasedly determine whether a compound can protect p53 from the E6-mediated degradation in cells, through a simple 3-step protocol. We validated the assay by testing two small molecules, SAHA and RITA, reported to impair the E6-mediated p53 degradation. Interestingly, we observed that only SAHA efficiently rescued p53, while RITA could not provide the same degree of protection. The possibility to specifically and quantitatively monitor the ability of a selected compound to rescue p53 in a cellular context through our LumiFluo assay could represent an important step towards the successful development of anti-HPV drugs.
published_date 2018-04-16T04:13:32Z
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