Monitoring regulatory T cells in clinical samples: consensus on an essential marker set and gating strategy for regulatory T cell analysis by flow cytometry

Regulatory T cell (Treg)-mediated immunosuppression is considered a major obstacle for successful cancer immunotherapy. The association between clinical outcome and Tregs is being studied extensively in clinical trials, but unfortunately, no consensus has been reached about (a) the markers and (b) t...

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Vydáno v:	Cancer Immunology, Immunotherapy Ročník 64; číslo 10; s. 1271 - 1286
Hlavní autoři:	Santegoets, Saskia J. A. M., Dijkgraaf, Eveline M., Battaglia, Alessandra, Beckhove, Philipp, Britten, Cedrik M., Gallimore, Awen, Godkin, Andrew, Gouttefangeas, Cecile, de Gruijl, Tanja D., Koenen, Hans J. P. M., Scheffold, Alexander, Shevach, Ethan M., Staats, Janet, Taskén, Kjetil, Whiteside, Theresa L., Kroep, Judith R., Welters, Marij J. P., van der Burg, Sjoerd H.
Médium:	Journal Article
Jazyk:	angličtina
Vydáno:	Berlin/Heidelberg Springer Berlin Heidelberg 01.10.2015 Springer Nature B.V
Témata:	Animals Antigens, CD - metabolism Biomarkers - metabolism Cancer Cancer Research Cell Separation Cells, Cultured Clinical trials Consensus Cytokines Female Flow Cytometry Forkhead Transcription Factors - metabolism Humans Immunology Immunotherapy International Cooperation Ki-67 Antigen - metabolism Lymphocyte Activation Lymphocytes Medical prognosis Medical research Medicine Medicine & Public Health Monitoring, Immunologic Oncology Original Original Article Ovarian Neoplasms - immunology Reference Standards T-Lymphocytes, Regulatory - immunology Tumor Escape Netherlands United Kingdom > UK United States > US Germany Flow cytometry Tregs Consensus Monitoring Phenotyping
ISSN:	0340-7004, 1432-0851
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Abstract	Regulatory T cell (Treg)-mediated immunosuppression is considered a major obstacle for successful cancer immunotherapy. The association between clinical outcome and Tregs is being studied extensively in clinical trials, but unfortunately, no consensus has been reached about (a) the markers and (b) the gating strategy required to define human Tregs in this context, making it difficult to draw final conclusions. Therefore, we have organized an international workshop on the detection and functional testing of Tregs with leading experts in the field, and 40 participants discussing different analyses and the importance of different markers and context in which Tregs were analyzed. This resulted in a rationally composed ranking list of “Treg markers”. Subsequently, the proposed Treg markers were tested to get insight into the overlap/differences between the most frequently used Treg definitions and their utility for Treg detection in various human tissues. Here, we conclude that the CD3, CD4, CD25, CD127, and FoxP3 markers are the minimally required markers to define human Treg cells. Staining for Ki67 and CD45RA showed to provide additional information on the activation status of Tregs. The use of markers was validated in a series of PBMC from healthy donors and cancer patients, as well as in tumor-draining lymph nodes and freshly isolated tumors. In conclusion, we propose an essential marker set comprising antibodies to CD3, CD4, CD25, CD127, Foxp3, Ki67, and CD45RA and a corresponding robust gating strategy for the context-dependent analysis of Tregs by flow cytometry.
AbstractList	Regulatory T cell (Treg)-mediated immunosuppression is considered a major obstacle for successful cancer immunotherapy. The association between clinical outcome and Tregs is being studied extensively in clinical trials, but unfortunately, no consensus has been reached about (a) the markers and (b) the gating strategy required to define human Tregs in this context, making it difficult to draw final conclusions. Therefore, we have organized an international workshop on the detection and functional testing of Tregs with leading experts in the field, and 40 participants discussing different analyses and the importance of different markers and context in which Tregs were analyzed. This resulted in a rationally composed ranking list of “Treg markers”. Subsequently, the proposed Treg markers were tested to get insight into the overlap/differences between the most frequently used Treg definitions and their utility for Treg detection in various human tissues. Here, we conclude that the CD3, CD4, CD25, CD127, and FoxP3 markers are the minimally required markers to define human Treg cells. Staining for Ki67 and CD45RA showed to provide additional information on the activation status of Tregs. The use of markers was validated in a series of PBMC from healthy donors and cancer patients, as well as in tumor-draining lymph nodes and freshly isolated tumors. In conclusion, we propose an essential marker set comprising antibodies to CD3, CD4, CD25, CD127, Foxp3, Ki67, and CD45RA and a corresponding robust gating strategy for the context-dependent analysis of Tregs by flow cytometry.
Author	Beckhove, Philipp Santegoets, Saskia J. A. M. Taskén, Kjetil Battaglia, Alessandra Gouttefangeas, Cecile de Gruijl, Tanja D. Britten, Cedrik M. Godkin, Andrew Dijkgraaf, Eveline M. Koenen, Hans J. P. M. Kroep, Judith R. Whiteside, Theresa L. Welters, Marij J. P. Gallimore, Awen van der Burg, Sjoerd H. Shevach, Ethan M. Scheffold, Alexander Staats, Janet
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BackLink	https://www.ncbi.nlm.nih.gov/pubmed/26122357$$D View this record in MEDLINE/PubMed
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Keywords	Flow cytometry Tregs Consensus Monitoring Phenotyping
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Snippet	Regulatory T cell (Treg)-mediated immunosuppression is considered a major obstacle for successful cancer immunotherapy. The association between clinical...
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SubjectTerms	Animals Antigens, CD - metabolism Biomarkers - metabolism Cancer Cancer Research Cell Separation Cells, Cultured Clinical trials Consensus Cytokines Female Flow Cytometry Forkhead Transcription Factors - metabolism Humans Immunology Immunotherapy International Cooperation Ki-67 Antigen - metabolism Lymphocyte Activation Lymphocytes Medical prognosis Medical research Medicine Medicine & Public Health Monitoring, Immunologic Oncology Original Original Article Ovarian Neoplasms - immunology Reference Standards T-Lymphocytes, Regulatory - immunology Tumor Escape
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Title	Monitoring regulatory T cells in clinical samples: consensus on an essential marker set and gating strategy for regulatory T cell analysis by flow cytometry
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