RESUMO
In this paper, we describe a new method for preparation of human dendritic cells (DCs) that secrete bioactive IL-12(p70) using synthetic immunostimulatory compounds as TLR7/8 agonists. Monocyte-derived DCs were generated using a procedure that provided mature cells within 3 d. Several maturation mixtures that contained various cytokines, IFN-gamma, different TLR agonists, and PGE(2) were compared for impact on cell recovery, phenotype, cytokine secretion, migration, and lymphocyte activation. Mixtures that included the TLR7/8 agonists R848 or CL075, combined with the TLR3 agonist polyinosinic:polycytidylic acid, yielded 3-d mature DCs that secreted high levels of IL-12(p70), showed strong chemotaxis to CCR7 ligands, and had a positive costimulatory potential. They also had excellent capacity to activate NK cells, effectively polarized CD4(+) and CD8(+) T cells to secrete IFN-gamma and to induce T cell-mediated cytotoxic function. Thereby, mature DCs prepared within 3 d using such maturation mixtures displayed optimal functions required for vaccine development.
Assuntos
Diferenciação Celular/imunologia , Polaridade Celular/imunologia , Células Dendríticas/imunologia , Células Dendríticas/transplante , Quinolinas/farmacologia , Células Th1/imunologia , Tiazóis/farmacologia , Receptor 7 Toll-Like/agonistas , Receptor 8 Toll-Like/agonistas , Adjuvantes Imunológicos/farmacologia , Vacinas Anticâncer/síntese química , Diferenciação Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Movimento Celular/imunologia , Polaridade Celular/efeitos dos fármacos , Células Cultivadas , Técnicas de Cocultura , Citocinas/metabolismo , Células Dendríticas/citologia , Humanos , Imidazóis/agonistas , Imidazóis/farmacologia , Imunoterapia Adotiva/métodos , Interferon gama/metabolismo , Células Matadoras Naturais/efeitos dos fármacos , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/metabolismo , Ligantes , Ativação Linfocitária/efeitos dos fármacos , Ativação Linfocitária/imunologia , Poli I-C/metabolismo , Linfócitos T Citotóxicos/efeitos dos fármacos , Linfócitos T Citotóxicos/imunologia , Linfócitos T Citotóxicos/metabolismo , Células Th1/efeitos dos fármacos , Receptor 3 Toll-Like/metabolismo , Receptor 7 Toll-Like/fisiologia , Receptor 8 Toll-Like/fisiologiaRESUMO
Posttransplantation lymphoproliferative disease (PTLD) associated with Epstein-Barr virus (EBV) is a life-threatening complication after allogeneic hematopoietic stem cell transplantation. PTLD is efficiently prevented by adoptive transfer of EBV-specific T cells from the donor. To make EBV-specific T cells available in urgent clinical situations, we developed a rapid protocol for their isolation by overnight stimulation of donor blood cells with peptides derived from 11 EBV antigens, interferon-gamma surface capture, and immunomagnetic separation. Six patients with PTLD received 1 transfusion of EBV-specific T cells. No response was seen in 3 patients who had late-stage disease with multiorgan dysfunction at the time of T-cell transfer. In 3 patients who received T cells at an earlier stage of disease, we observed complete and stable remission of PTLD. Two patients have remained free from EBV-associated disease for more than 2 years. CD8(+) T cells specific for EBV early antigens rapidly expanded after T-cell transfer, temporarily constituted greater than 20% of all peripheral blood lymphocytes, and were maintained throughout the observation period. Thus, a rapid and sustained reconstitution of a protective EBV-specific T-cell memory occurred after the infusion of small numbers of directly isolated EBV-specific T cells.