LPD lipopolyplex initiates a potent cytokine response and inhibits tumor growth

Our laboratory has recently developed a lipopolyplex consisting of DOTAP:ch olesterol liposomes, protamine sulfate, and plasmid DNA (LPD) that provides improved systemic gene delivery compared with lipoplex following tail vein injection in mice. Because endothelial cells are the primary cells transfe cted in the lung, it was hypothesized that LPD might be an effective vector for gene therapy of pulmonary metastases. This hypothesis was examined by testing the efficacy of cytokine (IL-12) and tumor suppressor (p53) strateg ies for treatment of an experimental model of pulmonary metastasis in C57Bl /6 mice. Surprisingly, all LPD complexes including those containing an plas mid provided a potent (>50% inhibition) and dose-dependent antitumor effect , compared with dextrose-treated controls. In addition, iv injections of LP D containing 'empty' plasmid also inhibited tumor growth in a subcutaneous model of C3 fibrosarcomma. The antitumor effect correlated well with a stro ng and rapid proinflammatory cytokine (TNF-alpha, IL-12 and IFN-gamma) resp onse. Naked plasmid DNA did not elicit a cytokine response and the response required assembly of DNA into a lipoplex or the LPD lipopolyplex. Except f or the heart, elevated levels of cytokine were observed in all organs (lung , liver, kidney and spleen) where LPD is known to have gene transfer activi ty. Methylation of immune-stimulatory CpG motifs in the plasmid component o f LPD inhibited the proinflammatory cytokine response as well as the antitu mor effect of LPD in both-tumor systems. This suggests that iv administrati on of LPD elicits a systemic proinflammatory cytokine response that mediate s the antitumor activity of the lipopolyplex. In addition, the antitumor ac tivity was not observed in SCID mice suggesting a possible role for B or T lymphocytes in the antitumor response initiated by LPD. This represents the first demonstration that an intravenously administered cationic liposome-b ased nonviral vector can promote a systemic, Th1-like innate immune respons e. The immune adjuvant properties of LPD might prove to be suitable for del ivering tumor-specific antigens in the context of DNA vaccination.