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A deimmunised form of the ribotoxin, α-sarcin, lacking CD4(+) T cell epitopes and its use as an immunotoxin warhead
Fungal ribotoxins that block protein synthesis can be useful warheads in the context of a targeted immunotoxin. α-Sarcin is a small (17 kDa) fungal ribonuclease produced by Aspergillus giganteus that functions by catalytically cleaving a single phosphodiester bond in the sarcin–ricin loop of the lar...
Autores principales: | , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Oxford University Press
2016
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5081043/ https://www.ncbi.nlm.nih.gov/pubmed/27578884 http://dx.doi.org/10.1093/protein/gzw045 |
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author | Jones, Tim D. Hearn, Arron R. Holgate, Robert G.E. Kozub, Dorota Fogg, Mark H. Carr, Francis J. Baker, Matthew P. Lacadena, Javier Gehlsen, Kurt R. |
author_facet | Jones, Tim D. Hearn, Arron R. Holgate, Robert G.E. Kozub, Dorota Fogg, Mark H. Carr, Francis J. Baker, Matthew P. Lacadena, Javier Gehlsen, Kurt R. |
author_sort | Jones, Tim D. |
collection | PubMed |
description | Fungal ribotoxins that block protein synthesis can be useful warheads in the context of a targeted immunotoxin. α-Sarcin is a small (17 kDa) fungal ribonuclease produced by Aspergillus giganteus that functions by catalytically cleaving a single phosphodiester bond in the sarcin–ricin loop of the large ribosomal subunit, thus making the ribosome unrecognisable to elongation factors and leading to inhibition of protein synthesis. Peptide mapping using an ex vivo human T cell assay determined that α-sarcin contained two T cell epitopes; one in the N-terminal 20 amino acids and the other in the C-terminal 20 amino acids. Various mutations were tested individually within each epitope and then in combination to isolate deimmunised α-sarcin variants that had the desired properties of silencing T cell epitopes and retention of the ability to inhibit protein synthesis (equivalent to wild-type, WT α-sarcin). A deimmunised variant (D9T/Q142T) demonstrated a complete lack of T cell activation in in vitro whole protein human T cell assays using peripheral blood mononuclear cells from donors with diverse HLA allotypes. Generation of an immunotoxin by fusion of the D9T/Q142T variant to a single-chain Fv targeting Her2 demonstrated potent cell killing equivalent to a fusion protein comprising the WT α-sarcin. These results represent the first fungal ribotoxin to be deimmunised with the potential to construct a new generation of deimmunised immunotoxin therapeutics. |
format | Online Article Text |
id | pubmed-5081043 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2016 |
publisher | Oxford University Press |
record_format | MEDLINE/PubMed |
spelling | pubmed-50810432016-10-27 A deimmunised form of the ribotoxin, α-sarcin, lacking CD4(+) T cell epitopes and its use as an immunotoxin warhead Jones, Tim D. Hearn, Arron R. Holgate, Robert G.E. Kozub, Dorota Fogg, Mark H. Carr, Francis J. Baker, Matthew P. Lacadena, Javier Gehlsen, Kurt R. Protein Eng Des Sel Original Article Fungal ribotoxins that block protein synthesis can be useful warheads in the context of a targeted immunotoxin. α-Sarcin is a small (17 kDa) fungal ribonuclease produced by Aspergillus giganteus that functions by catalytically cleaving a single phosphodiester bond in the sarcin–ricin loop of the large ribosomal subunit, thus making the ribosome unrecognisable to elongation factors and leading to inhibition of protein synthesis. Peptide mapping using an ex vivo human T cell assay determined that α-sarcin contained two T cell epitopes; one in the N-terminal 20 amino acids and the other in the C-terminal 20 amino acids. Various mutations were tested individually within each epitope and then in combination to isolate deimmunised α-sarcin variants that had the desired properties of silencing T cell epitopes and retention of the ability to inhibit protein synthesis (equivalent to wild-type, WT α-sarcin). A deimmunised variant (D9T/Q142T) demonstrated a complete lack of T cell activation in in vitro whole protein human T cell assays using peripheral blood mononuclear cells from donors with diverse HLA allotypes. Generation of an immunotoxin by fusion of the D9T/Q142T variant to a single-chain Fv targeting Her2 demonstrated potent cell killing equivalent to a fusion protein comprising the WT α-sarcin. These results represent the first fungal ribotoxin to be deimmunised with the potential to construct a new generation of deimmunised immunotoxin therapeutics. Oxford University Press 2016-11 2016-10-22 /pmc/articles/PMC5081043/ /pubmed/27578884 http://dx.doi.org/10.1093/protein/gzw045 Text en © The Author 2016. Published by Oxford University Press. http://creativecommons.org/licenses/by/4.0/ This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | Original Article Jones, Tim D. Hearn, Arron R. Holgate, Robert G.E. Kozub, Dorota Fogg, Mark H. Carr, Francis J. Baker, Matthew P. Lacadena, Javier Gehlsen, Kurt R. A deimmunised form of the ribotoxin, α-sarcin, lacking CD4(+) T cell epitopes and its use as an immunotoxin warhead |
title | A deimmunised form of the ribotoxin, α-sarcin, lacking CD4(+) T cell epitopes and its use as an immunotoxin warhead |
title_full | A deimmunised form of the ribotoxin, α-sarcin, lacking CD4(+) T cell epitopes and its use as an immunotoxin warhead |
title_fullStr | A deimmunised form of the ribotoxin, α-sarcin, lacking CD4(+) T cell epitopes and its use as an immunotoxin warhead |
title_full_unstemmed | A deimmunised form of the ribotoxin, α-sarcin, lacking CD4(+) T cell epitopes and its use as an immunotoxin warhead |
title_short | A deimmunised form of the ribotoxin, α-sarcin, lacking CD4(+) T cell epitopes and its use as an immunotoxin warhead |
title_sort | deimmunised form of the ribotoxin, α-sarcin, lacking cd4(+) t cell epitopes and its use as an immunotoxin warhead |
topic | Original Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5081043/ https://www.ncbi.nlm.nih.gov/pubmed/27578884 http://dx.doi.org/10.1093/protein/gzw045 |
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