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Low cross reactivity between wild type and deamidated AAV can lead to false negative results in immune monitoring T-cell assays

During gene therapy trials, immune responses against adeno-associated virus (AAV) vectors are monitored by antibody assays that detect the humoral and T-cell mediated cellular responses to AAV vectors. T cell assays commonly utilize the collection of patients’ peripheral blood mononuclear cells (PBM...

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Autores principales: Bing, So Jin, Warrington, Stephanee, Mazor, Ronit
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10352612/
https://www.ncbi.nlm.nih.gov/pubmed/37469509
http://dx.doi.org/10.3389/fimmu.2023.1211529
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author Bing, So Jin
Warrington, Stephanee
Mazor, Ronit
author_facet Bing, So Jin
Warrington, Stephanee
Mazor, Ronit
author_sort Bing, So Jin
collection PubMed
description During gene therapy trials, immune responses against adeno-associated virus (AAV) vectors are monitored by antibody assays that detect the humoral and T-cell mediated cellular responses to AAV vectors. T cell assays commonly utilize the collection of patients’ peripheral blood mononuclear cells (PBMCs) and stimulation with AAV-derived overlapping peptides. We recently described that spontaneous deamidation coincides with T cell epitopes in AAV capsids and that spontaneous deamidation may enhance or decrease immunogenicity in some individuals. This raised the concern for false negative results of antibody detection and PBMC immune monitoring assays because these assays use wild-type (WT) AAV or WT peptides for T cell re-stimulation and these peptides may not re-activate T cells that were stimulated with deamidated AAV capsid. To investigate this concern, we modeled the scenario by expanding T cells with deamidated peptides and evaluated the cross-reactivity of expanded T cells to WT peptides. In the majority of samples, cells that were expanded with deamidated peptides and restimulated with WT peptide had significantly lowered IL-2 and IFN-γ production. Spiking the four deamidated peptides to the WT peptide pool used for re-stimulation, restored the signal and corrected the performance of the assay. We also evaluated the impact of deamidation on anti AAV binding antibodies and did not observe a major impact on seroprevalence detection of AAV9. These data indicate that a high level of deamidation in AAV therapy may result in underestimation or even failure to detect immune responses against WT peptides during cellular immune monitoring.
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spelling pubmed-103526122023-07-19 Low cross reactivity between wild type and deamidated AAV can lead to false negative results in immune monitoring T-cell assays Bing, So Jin Warrington, Stephanee Mazor, Ronit Front Immunol Immunology During gene therapy trials, immune responses against adeno-associated virus (AAV) vectors are monitored by antibody assays that detect the humoral and T-cell mediated cellular responses to AAV vectors. T cell assays commonly utilize the collection of patients’ peripheral blood mononuclear cells (PBMCs) and stimulation with AAV-derived overlapping peptides. We recently described that spontaneous deamidation coincides with T cell epitopes in AAV capsids and that spontaneous deamidation may enhance or decrease immunogenicity in some individuals. This raised the concern for false negative results of antibody detection and PBMC immune monitoring assays because these assays use wild-type (WT) AAV or WT peptides for T cell re-stimulation and these peptides may not re-activate T cells that were stimulated with deamidated AAV capsid. To investigate this concern, we modeled the scenario by expanding T cells with deamidated peptides and evaluated the cross-reactivity of expanded T cells to WT peptides. In the majority of samples, cells that were expanded with deamidated peptides and restimulated with WT peptide had significantly lowered IL-2 and IFN-γ production. Spiking the four deamidated peptides to the WT peptide pool used for re-stimulation, restored the signal and corrected the performance of the assay. We also evaluated the impact of deamidation on anti AAV binding antibodies and did not observe a major impact on seroprevalence detection of AAV9. These data indicate that a high level of deamidation in AAV therapy may result in underestimation or even failure to detect immune responses against WT peptides during cellular immune monitoring. Frontiers Media S.A. 2023-07-04 /pmc/articles/PMC10352612/ /pubmed/37469509 http://dx.doi.org/10.3389/fimmu.2023.1211529 Text en Copyright © 2023 Bing, Warrington and Mazor https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Immunology
Bing, So Jin
Warrington, Stephanee
Mazor, Ronit
Low cross reactivity between wild type and deamidated AAV can lead to false negative results in immune monitoring T-cell assays
title Low cross reactivity between wild type and deamidated AAV can lead to false negative results in immune monitoring T-cell assays
title_full Low cross reactivity between wild type and deamidated AAV can lead to false negative results in immune monitoring T-cell assays
title_fullStr Low cross reactivity between wild type and deamidated AAV can lead to false negative results in immune monitoring T-cell assays
title_full_unstemmed Low cross reactivity between wild type and deamidated AAV can lead to false negative results in immune monitoring T-cell assays
title_short Low cross reactivity between wild type and deamidated AAV can lead to false negative results in immune monitoring T-cell assays
title_sort low cross reactivity between wild type and deamidated aav can lead to false negative results in immune monitoring t-cell assays
topic Immunology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10352612/
https://www.ncbi.nlm.nih.gov/pubmed/37469509
http://dx.doi.org/10.3389/fimmu.2023.1211529
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