Plasmid DNA adsorbed onto cationic microparticles mediates target gene expression and antigen presentation by dendritic cells

Dendritic cells (DC) play a key role in antigen presentation and activation of specific immunity. Much current research focuses on harnessing the pote ncy of DC for vaccines, gene therapy, and cancer immunotherapy applications . However, DC are not readily transfected in vitro by traditional nonviral techniques. A novel DNA vaccine formulation was used to determine if DC are transfected in vitro. The formulation consists of plasmid DNA adsorbed on to cationic microparticles composed of the biodegradable polymer polylactid e-co-glycolide (PLG) and the cationic surfactant, cetyltrimethylammonium br omide (CTAB). Using preparations of fluorescent-labeled plasmid DNA formula ted on PLG-CTAB microparticles to study internalization by macrophages and dendritic cells in vitro and in vivo, we found that most, but not all, of t he fluorescence was concentrated in endosomal compartments. Furthermore, up take of plasmid DNA encoding HIV p55 gag adsorbed to PLG-CTAB microparticle s by murine bone marrow-derived dendritic cells resulted in target gene exp ression, as detected by RT-PCR. The antigen was subsequently processed and presented, resulting in stimulation of an H-2k(d)-restricted, gag-specific T cell hybridoma. Activation of the hybridoma, detected by IL-2 production, was dose-dependent in the range of 0.1-20 mug DNA (10-2000 mug PLG) and wa s sustained up to 5 days after transfection. Thus, adsorption of plasmid DN A on PLG-CTAB microparticles provides a potentially useful nonviral approac h for in vitro transfection of dendritic cells.