Flow Cytometric Clinical Immunomonitoring Using Peptide–MHC Class II Tetramers: Optimization of Methods and Protocol Development

Jansen, Diahann T. S. L.; Ramnoruth, Nishta; Loh, Khai Lee; Rossjohn, Jamie; Reid, Hugh Harrington; Nel, Hendrik J.; Thomas, Ranjeny

doi:10.3389/fimmu.2018.00008

Cited by 13 publications

(14 citation statements)

References 26 publications

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“…In the first Phase I proof-of-concept clinical trial of antigen-specific DC immunotherapy, HLA-DR SE + ACPA + RA patients were treated with autologous PB DCs modified with NF-κB inhibitor, BAY11-7082, and exposed to four citrullinated peptide autoantigens. These trials begin to build a mechanistic understanding around the potential for DC-based antigenspecific immunotherapies to rebalance antigen-specific regulatory to effector T cells, and highlight the need for sensitive, standardized and clinically qualified immunological assays, such as tetramers, to determine the pharmacodynamic immunological effects of antigen-specific therapies so that effects can be evaluated and compared in early-stage trials [102]. In treated subjects, ex-vivo IL-6 production to cit-vimentin 447-455 was suppressed [131].…”

Section: Antigen-specific Immunotherapy Targeting the Dc-t Cell Intermentioning

confidence: 99%

“…4a). As the pMHCII multimers are fluorescently labelled, CD4 + T cells specific for the (auto)antigen can be detected and quantified by flow cytometry [101,102] (Fig. 4b).…”

Section: Autoantigen-specific Cd4 + T Cells In Ramentioning

confidence: 99%

“…A second trial of tolerogenic DCs exposed to antigens in autologous synovial fluid also demonstrated safety and feasibility [132], and further DC trials are ongoing (see clinicaltrials.gov). These trials begin to build a mechanistic understanding around the potential for DC-based antigenspecific immunotherapies to rebalance antigen-specific regulatory to effector T cells, and highlight the need for sensitive, standardized and clinically qualified immunological assays, such as tetramers, to determine the pharmacodynamic immunological effects of antigen-specific therapies so that effects can be evaluated and compared in early-stage trials [102].…”

Section: Antigen-specific Immunotherapy Targeting the Dc-t Cell Intermentioning

confidence: 99%

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Dendritic cells, T cells and their interaction in rheumatoid arthritis

Wehr

Purvis

Law

et al. 2019

Clinical and Experimental Immunology

127

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Dendritic cells (DCs) are the key professional antigen-presenting cells which bridge innate and adaptive immune responses, inducing the priming and differentiation of naive to effector CD4 + T cells, the cross-priming of CD8 + T cells and the promotion of B cell antibody responses. DCs also play a critical role in the maintenance of immune homeostasis and tolerance. DC-T cell interactions underpin the generation of an autoimmune response in rheumatoid arthritis (RA). Here we describe the function of DCs and review evidence for DC and T cell involvement in RA pathogenesis, in particular through the presentation of self-peptide by DCs that triggers differentiation and activation of autoreactive T cells. Finally, we discuss the emerging field of targeting the DC-T cell interaction for antigen-specific immunotherapy of RA.

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Section: Antigen-specific Immunotherapy Targeting the Dc-t Cell Intermentioning

confidence: 99%

“…4a). As the pMHCII multimers are fluorescently labelled, CD4 + T cells specific for the (auto)antigen can be detected and quantified by flow cytometry [101,102] (Fig. 4b).…”

Section: Autoantigen-specific Cd4 + T Cells In Ramentioning

confidence: 99%

Section: Antigen-specific Immunotherapy Targeting the Dc-t Cell Intermentioning

confidence: 99%

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Dendritic cells, T cells and their interaction in rheumatoid arthritis

Wehr

Purvis

Law

et al. 2019

Clinical and Experimental Immunology

127

View full text Add to dashboard Cite

show abstract

“…present in many or the majority of patients, and which are private. Such information would be invaluable for further refinement of an immune-monitoring protocol and for designing future immunotherapies based on the exciting progress being made in RA in this regard [59,60].…”

Section: Discussionmentioning

confidence: 99%

Multi-HLA class II tetramer analyses of citrulline-reactive T cells and early treatment response in rheumatoid arthritis

Gerstner

Turcinov

Hensvold

et al. 2020

Preprint

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Background: HLA class II tetramers can be used for ex vivo enumeration and phenotypic characterization of antigen-specific CD4+ T cells. They are increasingly applied in settings like allergy, vaccination and autoimmune diseases. Rheumatoid arthritis (RA) is a chronic autoimmune disorder for which many autoantigens have been described.Results: Using multi-parameter flow cytometry, we developed a multi-HLA class II tetramer approach to simultaneously study several antigen specificities in RA patient samples. We focused on previously described citrullinated HLA-DRB1*04:01-restricted T cell epitopes from α-enolase, fibrinogen-b, vimentin as well as cartilage intermediate layer protein (CILP). First, we examined inter-assay variability and the sensitivity of the assay in peripheral blood from healthy donors (n=7). Next, we confirmed the robustness and sensitivity in a cohort of RA patients with repeat blood draws (n=14). We then applied our method in two different settings. We assessed lymphoid tissue from seropositive arthralgia (n=5) and early RA patients (n=5) and could demonstrate autoreactive T cells in individuals at risk of developing RA. Lastly, we studied peripheral blood from early RA patients (n=10) and found that the group of patients achieving minimum disease activity (DAS28 <2.6) at 6 months follow-up displayed a decrease in the frequency of citrulline-specific T cells. Conclusions: Our study demonstrates the development of a sensitive tetramer panel allowing simultaneous characterization of antigen-specific T cells in ex vivo patient samples including RA ‘at risk’ subjects. This multi-tetramer approach can be useful for longitudinal immune-monitoring in any disease with known HLA-restriction element and several candidate antigens.

show abstract

Section: Discussionmentioning

confidence: 99%

Multi-HLA class II tetramer analyses of citrulline-reactive T cells and early treatment response in rheumatoid arthritis

Gerstner

Turcinov

Hensvold

et al. 2020

Preprint

View full text Add to dashboard Cite

Background: HLA class II tetramers can be used for ex vivo enumeration and phenotypic characterization of antigen-specific CD4+ T cells. They are increasingly applied in settings like allergy, vaccination and autoimmune diseases. Rheumatoid arthritis (RA) is a chronic autoimmune disorder for which many autoantigens have been described. Results: Using multi-parameter flow cytometry, we developed a multi-HLA class II tetramer approach to simultaneously study several antigen specificities in RA patient samples. We focused on previously described citrullinated HLA-DRB1*04:01-restricted T cell epitopes from α-enolase, fibrinogen-b, vimentin as well as cartilage intermediate layer protein (CILP).First, we examined inter-assay variability and the sensitivity of the assay in peripheral blood from healthy donors (n=7). Next, we confirmed the robustness and sensitivity in a cohort of RA patients with repeat blood draws (n=14). We then applied our method in two different settings. We assessed lymphoid tissue from seropositive arthralgia (n=5) and early RA patients (n=5) and could demonstrate autoreactive T cells in individuals at risk of developing RA. Lastly, we studied peripheral blood from early RA patients (n=10) and found that the group of patients achieving minimum disease activity (DAS28 <2.6) at 6 months follow-up displayed a decrease in the frequency of citrulline-specific T cells. Conclusions: Our study demonstrates the development of a sensitive tetramer panel allowing simultaneous characterization of antigen-specific T cells in ex vivo patient samples including RA ‘at risk’ subjects. This multi-tetramer approach can be useful for longitudinal immune-monitoring in any disease with known HLA-restriction element and several candidate antigens.

show abstract

Flow Cytometric Clinical Immunomonitoring Using Peptide–MHC Class II Tetramers: Optimization of Methods and Protocol Development

Cited by 13 publications

References 26 publications

Dendritic cells, T cells and their interaction in rheumatoid arthritis

Dendritic cells, T cells and their interaction in rheumatoid arthritis

Multi-HLA class II tetramer analyses of citrulline-reactive T cells and early treatment response in rheumatoid arthritis

Multi-HLA class II tetramer analyses of citrulline-reactive T cells and early treatment response in rheumatoid arthritis

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