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Purification and Characterization of Antibodies in Single-Chain Format against the E6 Oncoprotein of Human Papillomavirus Type 16

In Human Papillomaviruses- (HPV-) associated carcinogenesis, continuous expression of the E6 oncoprotein supports its value as a potential target for the development of diagnostics and therapeutics for HPV cancer. We previously reported that the I7 single-chain antibody fragment (scFv) specific for...

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Autores principales: Verachi, F., Percario, Z., Di Bonito, P., Affabris, E., Amici, C., Accardi, L.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Hindawi 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5985123/
https://www.ncbi.nlm.nih.gov/pubmed/29888271
http://dx.doi.org/10.1155/2018/6583852
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author Verachi, F.
Percario, Z.
Di Bonito, P.
Affabris, E.
Amici, C.
Accardi, L.
author_facet Verachi, F.
Percario, Z.
Di Bonito, P.
Affabris, E.
Amici, C.
Accardi, L.
author_sort Verachi, F.
collection PubMed
description In Human Papillomaviruses- (HPV-) associated carcinogenesis, continuous expression of the E6 oncoprotein supports its value as a potential target for the development of diagnostics and therapeutics for HPV cancer. We previously reported that the I7 single-chain antibody fragment (scFv) specific for HPV16 E6, expressed as an intrabody by retroviral system, could inhibit significantly the growth of cervical cancer cells in vitro and was even able to reduce tumor development in experimental HPV-related cancer models. Nevertheless, for the development of therapeutic tools to be employed in humans, it is important to achieve maximum safety guarantee, which can be provided by the protein format. In the current study, two anti-16E6 scFvs derived from I7 were expressed in E. coli and purified in soluble form by affinity chromatography. Specificity, sensitivity and stability in physiologic environment of the purified scFvs were demonstrated by binding studies using recombinant 16E6 as an antigen. The scFvs functionality was confirmed by immunofluorescence in cervical cancer cells, where the scFvs were able to recognize the nuclear E6. Furthermore, an antiproliferative activity of the scFvI7nuc delivered in protein format to HPV16-positive cell lines was observed. Our results demonstrate that functional anti-16E6 scFvs can be produced in E. coli, suggesting that such purified antibodies could be used in the diagnosis and treatment of HPV-induced malignancies.
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spelling pubmed-59851232018-06-10 Purification and Characterization of Antibodies in Single-Chain Format against the E6 Oncoprotein of Human Papillomavirus Type 16 Verachi, F. Percario, Z. Di Bonito, P. Affabris, E. Amici, C. Accardi, L. Biomed Res Int Research Article In Human Papillomaviruses- (HPV-) associated carcinogenesis, continuous expression of the E6 oncoprotein supports its value as a potential target for the development of diagnostics and therapeutics for HPV cancer. We previously reported that the I7 single-chain antibody fragment (scFv) specific for HPV16 E6, expressed as an intrabody by retroviral system, could inhibit significantly the growth of cervical cancer cells in vitro and was even able to reduce tumor development in experimental HPV-related cancer models. Nevertheless, for the development of therapeutic tools to be employed in humans, it is important to achieve maximum safety guarantee, which can be provided by the protein format. In the current study, two anti-16E6 scFvs derived from I7 were expressed in E. coli and purified in soluble form by affinity chromatography. Specificity, sensitivity and stability in physiologic environment of the purified scFvs were demonstrated by binding studies using recombinant 16E6 as an antigen. The scFvs functionality was confirmed by immunofluorescence in cervical cancer cells, where the scFvs were able to recognize the nuclear E6. Furthermore, an antiproliferative activity of the scFvI7nuc delivered in protein format to HPV16-positive cell lines was observed. Our results demonstrate that functional anti-16E6 scFvs can be produced in E. coli, suggesting that such purified antibodies could be used in the diagnosis and treatment of HPV-induced malignancies. Hindawi 2018-05-20 /pmc/articles/PMC5985123/ /pubmed/29888271 http://dx.doi.org/10.1155/2018/6583852 Text en Copyright © 2018 F. Verachi et al. https://creativecommons.org/licenses/by/4.0/ This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
spellingShingle Research Article
Verachi, F.
Percario, Z.
Di Bonito, P.
Affabris, E.
Amici, C.
Accardi, L.
Purification and Characterization of Antibodies in Single-Chain Format against the E6 Oncoprotein of Human Papillomavirus Type 16
title Purification and Characterization of Antibodies in Single-Chain Format against the E6 Oncoprotein of Human Papillomavirus Type 16
title_full Purification and Characterization of Antibodies in Single-Chain Format against the E6 Oncoprotein of Human Papillomavirus Type 16
title_fullStr Purification and Characterization of Antibodies in Single-Chain Format against the E6 Oncoprotein of Human Papillomavirus Type 16
title_full_unstemmed Purification and Characterization of Antibodies in Single-Chain Format against the E6 Oncoprotein of Human Papillomavirus Type 16
title_short Purification and Characterization of Antibodies in Single-Chain Format against the E6 Oncoprotein of Human Papillomavirus Type 16
title_sort purification and characterization of antibodies in single-chain format against the e6 oncoprotein of human papillomavirus type 16
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5985123/
https://www.ncbi.nlm.nih.gov/pubmed/29888271
http://dx.doi.org/10.1155/2018/6583852
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