Glutaraldehyde-Modified Recombinant Fel d 1: A Hypoallergen With Negligible Biological Activity but Retained Immunogenicity
Background Recombinant allergens are under investigation for replacing allergen extracts in immunotherapy. Site-directed mutagenesis has been suggested as a strategy to develop hypoallergenic molecules that will reduce the risk of side effects. For decades, chemically modified allergen extracts have...
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doaj-4198442143cd4b4186ba538458a7900d2020-11-24T23:49:10ZengElsevierWorld Allergy Organization Journal1939-45512011-01-0147113120Glutaraldehyde-Modified Recombinant Fel d 1: A Hypoallergen With Negligible Biological Activity but Retained ImmunogenicitySerge A. Versteeg, BSc0Ingrid Bulder, BSc1Martin Himly, PhD2Toni M. van Capel, BSc3R. van den Hourt, PhD4Stef J. Koppelman, PhD5Esther C. de Jong, PhD6Fatima Ferreira, PhD7Ronald van Ree, PhD8Department of Experimental Immunology, Academic Medical Center, Amsterdam, The NetherlandsDepartment of Immunopathology, Sanquin Research, Amsterdam, The NetherlandsDepartment of Molecular Biology, University of Salzburg, Salzburg, AustriaDepartment of Cell Biology and Histology, Academic Medical Center, Amsterdam, The NetherlandsR&D Department, HAL Allergy, Leiden, The NetherlandsR&D Department, HAL Allergy, Leiden, The NetherlandsDepartment of Cell Biology and Histology, Academic Medical Center, Amsterdam, The NetherlandsDepartment of Molecular Biology, University of Salzburg, Salzburg, AustriaDepartment of Experimental Immunology, Academic Medical Center, Amsterdam, The Netherlands; Department of Otorhinolaryngology, Academic Medical Center, Amsterdam, The Netherlands; Corresponding author.Background Recombinant allergens are under investigation for replacing allergen extracts in immunotherapy. Site-directed mutagenesis has been suggested as a strategy to develop hypoallergenic molecules that will reduce the risk of side effects. For decades, chemically modified allergen extracts have been used for the same reason.Aim To evaluate whether glutaraldehyde modification is a good strategy to produce hypoallergenic recombinant allergens with retained immunogenicity.Methods Fel d 1 was cloned as a single construct linking both chains of the molecule and expressed in Escherichia coli and Pichia pastoris. After physicochemical purification, recombinant (r) Fel d 1 was chemically modified using glutaraldehyde. The effect of modification on immune reactivity was evaluated using radioallergosorbent test, CAP inhibition, competitive radioimmunoassay, enzyme-linked immunosorbent assay, basophil histamine release, and T-cell proliferation assays. Both natural and recombinant unmodified Fel d 1 were used as controls.Results rFel d 1 demonstrated similar IgE binding and biologic activity as its natural counterpart. Upon modification, IgE-binding potency decreased >1000-fold, translating into a >106-fold reduction in biologic activity assessed by basophil histamine release. In contrast, the modified recombinant did not show a decreased but even a moderately increased capacity (1.5-fold) to stimulate proliferation of T cells (P < 0.01). Finally, it induced specific IgG antibodies in rabbits that recognized the unmodified allergen.Conclusions Chemical modification is a practical and highly effective approach for achieving hypoallergenicity of recombinant allergens with retained immunogenicity. Keywords: allergoid, Felis domesticus, hypoallergen, immunotherapy, rFel d 1http://www.sciencedirect.com/science/article/pii/S1939455119304296 |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Serge A. Versteeg, BSc Ingrid Bulder, BSc Martin Himly, PhD Toni M. van Capel, BSc R. van den Hourt, PhD Stef J. Koppelman, PhD Esther C. de Jong, PhD Fatima Ferreira, PhD Ronald van Ree, PhD |
spellingShingle |
Serge A. Versteeg, BSc Ingrid Bulder, BSc Martin Himly, PhD Toni M. van Capel, BSc R. van den Hourt, PhD Stef J. Koppelman, PhD Esther C. de Jong, PhD Fatima Ferreira, PhD Ronald van Ree, PhD Glutaraldehyde-Modified Recombinant Fel d 1: A Hypoallergen With Negligible Biological Activity but Retained Immunogenicity World Allergy Organization Journal |
author_facet |
Serge A. Versteeg, BSc Ingrid Bulder, BSc Martin Himly, PhD Toni M. van Capel, BSc R. van den Hourt, PhD Stef J. Koppelman, PhD Esther C. de Jong, PhD Fatima Ferreira, PhD Ronald van Ree, PhD |
author_sort |
Serge A. Versteeg, BSc |
title |
Glutaraldehyde-Modified Recombinant Fel d 1: A Hypoallergen With Negligible Biological Activity but Retained Immunogenicity |
title_short |
Glutaraldehyde-Modified Recombinant Fel d 1: A Hypoallergen With Negligible Biological Activity but Retained Immunogenicity |
title_full |
Glutaraldehyde-Modified Recombinant Fel d 1: A Hypoallergen With Negligible Biological Activity but Retained Immunogenicity |
title_fullStr |
Glutaraldehyde-Modified Recombinant Fel d 1: A Hypoallergen With Negligible Biological Activity but Retained Immunogenicity |
title_full_unstemmed |
Glutaraldehyde-Modified Recombinant Fel d 1: A Hypoallergen With Negligible Biological Activity but Retained Immunogenicity |
title_sort |
glutaraldehyde-modified recombinant fel d 1: a hypoallergen with negligible biological activity but retained immunogenicity |
publisher |
Elsevier |
series |
World Allergy Organization Journal |
issn |
1939-4551 |
publishDate |
2011-01-01 |
description |
Background Recombinant allergens are under investigation for replacing allergen extracts in immunotherapy. Site-directed mutagenesis has been suggested as a strategy to develop hypoallergenic molecules that will reduce the risk of side effects. For decades, chemically modified allergen extracts have been used for the same reason.Aim To evaluate whether glutaraldehyde modification is a good strategy to produce hypoallergenic recombinant allergens with retained immunogenicity.Methods Fel d 1 was cloned as a single construct linking both chains of the molecule and expressed in Escherichia coli and Pichia pastoris. After physicochemical purification, recombinant (r) Fel d 1 was chemically modified using glutaraldehyde. The effect of modification on immune reactivity was evaluated using radioallergosorbent test, CAP inhibition, competitive radioimmunoassay, enzyme-linked immunosorbent assay, basophil histamine release, and T-cell proliferation assays. Both natural and recombinant unmodified Fel d 1 were used as controls.Results rFel d 1 demonstrated similar IgE binding and biologic activity as its natural counterpart. Upon modification, IgE-binding potency decreased >1000-fold, translating into a >106-fold reduction in biologic activity assessed by basophil histamine release. In contrast, the modified recombinant did not show a decreased but even a moderately increased capacity (1.5-fold) to stimulate proliferation of T cells (P < 0.01). Finally, it induced specific IgG antibodies in rabbits that recognized the unmodified allergen.Conclusions Chemical modification is a practical and highly effective approach for achieving hypoallergenicity of recombinant allergens with retained immunogenicity. Keywords: allergoid, Felis domesticus, hypoallergen, immunotherapy, rFel d 1 |
url |
http://www.sciencedirect.com/science/article/pii/S1939455119304296 |
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