Quantitative assessment of human T lymphocytes in RAG2(-/-)gammac(-/-) mice: the impact of ex vivo manipulation on in vivo functionality.
OBJECTIVE
Recent clinical trials of adoptive immunotherapy showed diminished reactivity of human T cells upon ex vivo manipulation. For a safe and effective clinical application of human T cells, it is necessary to improve ex vivo manipulation procedures and evaluate their impact on in vivo functionality. However, there is no preclinical model for quantitative assessment of in vivo functionality of human T cells. In this study, we investigated the feasibility of using the huPBMC- RAG2(-/-)gammac(-/-) xenogeneic mouse model. As a first example, we compared 3 different ex vivo culture conditions for human T cells.


METHODS
RAG2(-/-)gammac(-/-) mice received cultured human T cells that were stimulated via CD3 alone or costimulated via CD28 (CD3/28) and/or human 4-1BB (CD3/28/4-1BB). Engraftment levels and survival of the cells were measured. The dynamics of the human T cell phenotypes were analyzed during culture and in vivo, as well as the mechanism of the xenoresponse.


RESULTS
Engraftment potential was improved twofold for costimulation compared to CD3 alone (p < 0.001). Phenotypic analysis showed a strikingly similar pattern of development towards CD4(+) and CD8(+) effector and effector-memory cells, suggesting antigen-driven survival and expansion. All parameters used to analyze different effects on in vivo T-cell functionality, like culture condition, engraftment levels, survival of the cells over time, or xenogeneic graft-vs-host disease were absolutely independent of the distribution of the T cell population in vivo following contact with xeno-antigen.


CONCLUSIONS
The huPBMC-RAG2(-/-)gammac(-/-) xenogeneic transplant model is the most sensitive to date for in vivo functional evaluation of human T cells.
van Rijn
Rozemarijn S
RS
Jordan Laboratory for Hemato-Oncology, Department of Hematology, University Medical Center Utrecht, Utrecht, The Netherlands.
Simonetti
Elles R
ER
Hagenbeek
Anton
A
Bonyhadi
Mark
M
Storm
Gert
G
Martens
Anton C M
AC
Ebeling
Saskia B
SB
D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods; D002470:Cell Survival; D004268:DNA-Binding Proteins / Q000172:deficiency*; D006085:Graft Survival; D006086:Graft vs Host Disease; D006801:Humans; D007142:Immunoglobulin gamma-Chains / Q000235:genetics; D016219:Immunotherapy, Adoptive / Q000379:methods*; D017710:Lymphocyte Transfusion / Q000379:methods; D051379:Mice; D018345:Mice, Knockout; D008822:Mice, Transgenic; D016176:T-Lymphocyte Subsets; D013601:T-Lymphocytes / Q000166:cytology* / Q000637:transplantation*
D004268:DNA-Binding Proteins; D007142:Immunoglobulin gamma-Chains; C499308:Rag2 protein, mouse