Ex vivo generation of human alloantigen-specific regulatory T cells from CD4(pos)CD25(high) T cells for immunotherapy

PLoS One. 2008 May 21;3(5):e2233. doi: 10.1371/journal.pone.0002233.


Background: Regulatory T cell (Treg) based immunotherapy is a potential treatment for several immune disorders. By now, this approach proved successful in preclinical animal transplantation and auto-immunity models. In these models the success of Treg based immunotherapy crucially depends on the antigen-specificity of the infused Treg population. For the human setting, information is lacking on how to generate Treg with direct antigen-specificity ex vivo to be used for immunotherapy.

Methodology/principal findings: Here, we demonstrate that in as little as two stimulation cycles with HLA mismatched allogeneic stimulator cells and T cell growth factors a very high degree of alloantigen-specificity was reached in magnetic bead isolated human CD4(pos)CD25(high) Treg. Efficient increases in cell numbers were obtained. Primary allogeneic stimulation appeared a prerequisite in the generation of alloantigen-specific Treg, while secondary allogeneic or polyclonal stimulation with anti-CD3 plus anti-CD28 monoclonal antibodies enriched alloantigen-specificity and cell yield to a similar extent.

Conclusions/significance: The ex vivo expansion protocol that we describe will very likely increase the success of clinical Treg-based immunotherapy, and will help to induce tolerance to selected antigens, while minimizing general immune suppression. This approach is of particular interest for recipients of HLA mismatched transplants.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • CD4 Antigens / immunology*
  • Clonal Anergy
  • Flow Cytometry
  • Humans
  • Immunotherapy*
  • Interleukin-2 Receptor alpha Subunit / immunology*
  • Isoantigens / immunology*
  • T-Lymphocytes, Regulatory / immunology*


  • CD4 Antigens
  • Interleukin-2 Receptor alpha Subunit
  • Isoantigens