A Simple Immunomonitoring Procedure for mRNA-Loaded Dendritic Cell Therapy

Abstract

To develop a simple immunomonitoring method for dendritic cell therapy using messenger RNA (mRNA) as antigen, we evaluated whether Mycobacterium tuberculosis antigen 85A (Ag85A) mRNA-transfected peripheral blood mononuclear cells (PBMCs) could be used to stimulate the induction of interferon (IFN)-γ-producing T cells. PBMCs from 10 healthy donors were cocultured with autologous PBMCs transfected with mRNA overnight, and the number of IFN-γ-producing T cells was measured by flow cytometry. IFN-γ-producing CD4+ and CD8+ T cells were detected in 4 and 5 donors, respectively. PBMCs from 3 donors with negative results were then cocultured with Ag85A mRNA-transfected autologous PBMCs for 1 week to achieve in vitro primary induction of Ag85A-specific T cells. After restimulation with freshly prepared stimulator cells, a small but significant number of IFN-γ-producing CD8+ T cells was detected. The induction of IFN-γ-producing CD8+ T cells by overnight coculture was completely abolished by anti-class II or anti-interleukin-12 antibodies, whereas it was partially inhibited by anti-class I antibody. These data suggest that Ag85A mRNA-transfected PBMCs induce specific IFN-γ-producing T cells and might be applicable for immunomonitoring of mRNA-loaded dendritic cell therapy.