Alloantigen-specific regulatory T cells generated with a chimeric antigen receptor
Katherine G. MacDonald, Romy E. Hoeppli, Qing Huang, Jana Gillies, Dan S. Luciani, Paul C. Orban, Raewyn Broady, Megan K. Levings
Katherine G. MacDonald, Romy E. Hoeppli, Qing Huang, Jana Gillies, Dan S. Luciani, Paul C. Orban, Raewyn Broady, Megan K. Levings
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Technical Advance Immunology

Alloantigen-specific regulatory T cells generated with a chimeric antigen receptor

Abstract

Adoptive immunotherapy with regulatory T cells (Tregs) is a promising treatment for allograft rejection and graft-versus-host disease (GVHD). Emerging data indicate that, compared with polyclonal Tregs, disease-relevant antigen-specific Tregs may have numerous advantages, such as a need for fewer cells and reduced risk of nonspecific immune suppression. Current methods to generate alloantigen-specific Tregs rely on expansion with allogeneic antigen-presenting cells, which requires access to donor and recipient cells and multiple MHC mismatches. The successful use of chimeric antigen receptors (CARs) for the generation of antigen-specific effector T cells suggests that a similar approach could be used to generate alloantigen-specific Tregs. Here, we have described the creation of an HLA-A2–specific CAR (A2-CAR) and its application in the generation of alloantigen-specific human Tregs. In vitro, A2-CAR–expressing Tregs maintained their expected phenotype and suppressive function before, during, and after A2-CAR–mediated stimulation. In mouse models, human A2-CAR–expressing Tregs were superior to Tregs expressing an irrelevant CAR at preventing xenogeneic GVHD caused by HLA-A2+ T cells. Together, our results demonstrate that use of CAR technology to generate potent, functional, and stable alloantigen-specific human Tregs markedly enhances their therapeutic potential in transplantation and sets the stage for using this approach for making antigen-specific Tregs for therapy of multiple diseases.

Authors

Katherine G. MacDonald, Romy E. Hoeppli, Qing Huang, Jana Gillies, Dan S. Luciani, Paul C. Orban, Raewyn Broady, Megan K. Levings

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Figure 1

Construction, expression, and antigen specificity of an HLA-A2–specific CAR.

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Construction, expression, and antigen specificity of an HLA-A2–specific ...
(A) Schematic of domains in the A2-CAR. Gly, glycine serine linker; Myc, Myc-tag; TM, transmembrane. (B) Schematic of bidirectional lentiviral vectors encoding the truncated nerve growth factor receptor (ΔNGFR) as a selectable marker under a minimal CMV promoter and CARs specific for HLA-A2 or HER2 under the EF1α promoter. (C) 293T cells were transfected with an empty vector (ΔNGFR), or a vector encoding a HER2-CAR, or the A2-CAR. Surface expression was measured by detection of the Myc-epitope tag by flow cytometry. UT, untransfected. (D) 293T cells expressing the HER2-CAR or A2-CAR were stained with HLA-A2 or HLA-A24 tetramers (tet). Data are representative of 2 independent experiments.