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T-cell tolerance for variability in an HLA class I-presented influenza A virus epitope.

ABSTRACT: To escape immune recognition, viruses acquire amino acid substitutions in class I human leukocyte antigen (HLA)-presented cytotoxic T-lymphocyte (CTL) epitopes. Such viral escape mutations may (i) prevent peptide processing, (ii) diminish class I HLA binding, or (iii) alter T-cell recognition. Because residues 418 to 426 of the hypervariable influenza A virus nucleoprotein (NP(418-426)) epitope are consistently bound by class I HLA and presented to CTL, we assessed the impact that intraepitope sequence variability has upon T-cell recognition. CTL elicited by intranasal influenza virus infection were tested for their cross-recognition of 20 natural NP(418-426) epitope variants. Six of the variant epitopes, of both H1N1 and H3N2 origin, were cross-recognized by CTL while the remaining NP(418-426) epitope variants escaped targeting. A pattern emerged whereby variability at position 5 (P5) within the epitope reduced T-cell recognition, changes at P4 or P6 enabled CTL escape, and a mutation at P8 enhanced T-cell recognition. These data demonstrate that substitutions at P4 and/or P6 facilitate influenza virus escape from T-cell recognition and provide a model for the number, nature, and location of viral mutations that influence T-cell cross-recognition.

SUBMITTER: Wahl A

PROVIDER: S-EPMC2738244 | biostudies-literature | 2009 Sep

REPOSITORIES: biostudies-literature

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T-cell tolerance for variability in an HLA class I-presented influenza A virus epitope.

Wahl Angela A McCoy William WH Schafer Fredda F Bardet Wilfried W Buchli Rico R Fremont Daved H DH Hildebrand William H WH

Journal of virology 20090624 18

To escape immune recognition, viruses acquire amino acid substitutions in class I human leukocyte antigen (HLA)-presented cytotoxic T-lymphocyte (CTL) epitopes. Such viral escape mutations may (i) prevent peptide processing, (ii) diminish class I HLA binding, or (iii) alter T-cell recognition. Because residues 418 to 426 of the hypervariable influenza A virus nucleoprotein (NP(418-426)) epitope are consistently bound by class I HLA and presented to CTL, we assessed the impact that intraepitope s ...[more]

PMID: 19553306

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Project description:BackgroundThe functional integrity of human leukocyte antigen low expression variants is a prerequisite for considering them as essential in the matching process of hematopoietic stem cell donors and recipients to diminish the risk of serious complications such as graft-versus-host disease or graft rejection. The HLA-A*3014L variant has a disulfide bridge missing in the alpha2 domain which could affect peptide binding and presentation to T cells.Design and methodsHLA-A*3014L and HLA-A*3001 were expressed as truncated variants and peptides were eluted and subjected to pool sequencing by Edman degradation as well as to single-peptide sequencing by mass spectrometry. Quantitative analysis of binding peptides presented in vivo was performed by a flow cytometric peptide-binding assay using HLA-A*3001 and HLA-A*3014L-expressing B-LCLs.ResultsThe truncated HLA-A*3014L protein was secreted in the supernatant and it was possible to elute and sequence peptides. Sequence analysis of these eluted peptides revealed no relevant differences to the peptide motif of HLA-A*3001, indicating that the Cys164Ser substitution does not substantially alter the spectrum of presented peptides. Strong binding of one of the shared in vivo identified HLA-A*3001/3014L ligands was confirmed in the peptide-binding assay.ConclusionsThis study is the first to demonstrate that HLA low expression variants are able to present peptides and, thus, can be considered as functionally active. When comparing peptide motifs, it is likely that HLA-A*3014L and HLA-A*3001 represent a permissive mismatch with low allogenicity in hematopoietic stem cell transplantation. These results indicate that surface expression, as well as peptide-binding data of HLA variants with similar disulfide bridge variations (e.g. HLA-A*3211Q) need to be considered as functionally active in an allogeneic hematopoietic stem cell transplantation setting as long as the opposite has not been shown. Otherwise a relevant but not considered HLA mismatch could result in a severe allogeneic T-cell response and graft-versus-host disease.

Project description:BACKGROUND:Immunosuppressive therapy or T-cell depletion in transplant patients can cause uncontrolled growth of Epstein-Barr virus (EBV)-infected B cells resulting in post-transplant lymphoproliferative disease (PTLD). Current treatment options do not distinguish between healthy and malignant B cells and are thereby often limited by severe side effects in the already immunocompromised patients. To specifically target EBV-infected B cells, we developed a novel peptide-selective chimeric antigen receptor (CAR) based on the monoclonal antibody TÜ165 which recognizes an Epstein-Barr nuclear antigen (EBNA)-3C-derived peptide in HLA-B*35 context in a T-cell receptor (TCR)-like manner. In order to attract additional immune cells to proximity of PTLD cells, based on the TÜ165 CAR, we moreover generated T cells redirected for universal cytokine-mediated killing (TRUCKs), which induce interleukin (IL)-12 release on target contact. METHODS:TÜ165-based CAR-T cells (CAR-Ts) and TRUCKs with inducible IL-12 expression in an all-in-one construct were generated. Functionality of the engineered cells was assessed in co-cultures with EBNA-3C-peptide-loaded, HLA-B*35-expressing K562 cells and EBV-infected B cells as PTLD model. IL-12, secreted by TRUCKs on target contact, was further tested for its chemoattractive and activating potential towards monocytes and natural killer (NK) cells. RESULTS:After co-cultivation with EBV target cells, TÜ165 CAR-Ts and TRUCKs showed an increased activation marker expression (CD137, CD25) and release of proinflammatory cytokines (interferon-? and tumor necrosis factor-?). Moreover, TÜ165 CAR-Ts and TRUCKs released apoptosis-inducing mediators (granzyme B and perforin) and were capable to specifically lyse EBV-positive target cells. Live cell imaging revealed a specific attraction of TÜ165 CAR-Ts around EBNA-3C-peptide-loaded target cells. Of note, TÜ165 TRUCKs with inducible IL-12 showed highly improved effector functions and additionally led to recruitment of monocyte and NK cell lines. CONCLUSIONS:Our results demonstrate that TÜ165 CAR-Ts recognize EBV peptide/HLA complexes in a TCR-like manner and thereby allow for recognizing an intracellular EBV target. TÜ165 TRUCKs equipped with inducible IL-12 expression responded even more effectively and released IL-12 recruited additional immune cells which are generally missing in proximity of lymphoproliferation in immunocompromised PTLD patients. This suggests a new and promising strategy to specifically target EBV-infected cells while sparing and mobilizing healthy immune cells and thereby enable control of EBV-associated lymphoproliferation.

Dataset Information

T-cell tolerance for variability in an HLA class I-presented influenza A virus epitope.

Publications

T-cell tolerance for variability in an HLA class I-presented influenza A virus epitope.

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