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1.
J Immunol ; 181(4): 2855-68, 2008 Aug 15.
Article in English | MEDLINE | ID: mdl-18684977

ABSTRACT

The costimulatory requirements required for peripheral blood T regulatory cells (Tregs) are unclear. Using cell-based artificial APCs we found that CD28 but not ICOS, OX40, 4-1BB, CD27, or CD40 ligand costimulation maintained high levels of Foxp3 expression and in vitro suppressive function. Only CD28 costimulation in the presence of rapamycin consistently generated Tregs that consistently suppressed xenogeneic graft-vs-host disease in immunodeficient mice. Restimulation of Tregs after 8-12 days of culture with CD28 costimulation in the presence of rapamycin resulted in >1000-fold expansion of Tregs in <3 wk. Next, we determined whether other costimulatory pathways could augment the replicative potential of CD28-costimulated Tregs. We observed that while OX40 costimulation augmented the proliferative capacity of CD28-costimulated Tregs, Foxp3 expression and suppressive function were diminished. These studies indicate that the costimulatory requirements for expanding Tregs differ from those for T effector cells and, furthermore, they extend findings from mouse Tregs to demonstrate that human postthymic Tregs require CD28 costimulation to expand and maintain potent suppressive function in vivo.


Subject(s)
CD28 Antigens/metabolism , Lymphocyte Activation/immunology , T-Lymphocytes, Regulatory/cytology , T-Lymphocytes, Regulatory/immunology , Adoptive Transfer , Animals , CD28 Antigens/physiology , Cell Culture Techniques , Female , Graft vs Host Disease/immunology , Graft vs Host Disease/therapy , Humans , K562 Cells , Male , Mice , Mice, Inbred NOD , Mice, SCID , Signal Transduction/immunology , T-Lymphocytes, Regulatory/transplantation , Thymus Gland/cytology , Thymus Gland/immunology , Thymus Gland/metabolism
2.
Mol Ther ; 15(5): 981-8, 2007 May.
Article in English | MEDLINE | ID: mdl-17375070

ABSTRACT

To facilitate the therapeutic application of antigen-presenting cells (APCs), we have developed a cell-based artificial APC (aAPC) system by engineering K562 cells with lentiviruses to direct the stable expression and secretion of a variety of co-stimulatory molecules and cytokines. Here we report the use of a combinatorial lentiviral gene transfer approach to achieve long-term stable expression of at least seven genes in the K562 parental cell line. Expression of various combinations of genes on the aAPC enables the precise determination of human T-cell activation requirements, such that aAPCs can be tailored for the optimal propagation of T-cell subsets with specific growth requirements and distinct functions. The aAPCs support ex vivo growth and long-term expansion of functional human CD8 T cells without requiring the addition of exogenous cytokines, in contrast to the use of natural APCs. Distinct populations of T cells can be expanded with aAPCs expressing CD137L (4-1BBL) and/or CD80. Finally, the aAPCs provide an efficient platform to expand genetically modified T cells and to maintain CD28 expression on CD8 T cells. Therefore, K562-based aAPCs have therapeutic potential for adoptive immunotherapies and vaccinations.


Subject(s)
Antigen-Presenting Cells/metabolism , Antigens, Surface/metabolism , Cytokines/metabolism , Antigen-Presenting Cells/cytology , Antigen-Presenting Cells/immunology , Antigens, Surface/genetics , Antigens, Surface/immunology , CD28 Antigens/genetics , CD28 Antigens/immunology , CD28 Antigens/metabolism , CD8-Positive T-Lymphocytes/cytology , CD8-Positive T-Lymphocytes/metabolism , Cell Proliferation , Cells, Cultured , Cytokines/genetics , Cytokines/immunology , Humans , K562 Cells , Lentivirus/genetics , Microscopy, Phase-Contrast , Receptors, IgG/genetics , Receptors, IgG/immunology , Receptors, IgG/metabolism , Transduction, Genetic , Tumor Necrosis Factor Receptor Superfamily, Member 9/genetics , Tumor Necrosis Factor Receptor Superfamily, Member 9/immunology , Tumor Necrosis Factor Receptor Superfamily, Member 9/metabolism
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