LPD lipopolyplex initiates a potent cytokine response and inhibits tumor growth

Abstract

Our laboratory has recently developed a lipopolyplex response required assembly of DNA into a lipoplex or the consisting of DOTAP:cholesterol liposomes, protamine LPD lipopolyplex. Except for the heart, elevated levels of sulfate, and plasmid DNA (LPD) that provides improved cytokine were observed in all organs (lung, liver, kidney systemic gene delivery compared with lipoplex following and spleen) where LPD is known to have gene transfer tail vein injection in mice. Because endothelial cells are the activity. Methylation of immune-stimulatory CpG motifs in primary cells transfected in the lung, it was hypothesized the plasmid component of LPD inhibited the proinflammathat LPD might be an effective vector for gene therapy of tory cytokine response as well as the antitumor effect of pulmonary metastases. This hypothesis was examined by LPD in both tumor systems. This suggests that i.v. administesting the efficacy of cytokine (IL-12) and tumor suptration of LPD elicits a systemic proinflammatory cytokine pressor (p53) strategies for treatment of an experimental response that mediates the antitumor activity of the lipopomodel of pulmonary metastasis in C57Bl/6 mice. Surprislyplex. In addition, the antitumor activity was not observed ingly, all LPD complexes including those containing an in SCID mice suggesting a possible role for B or T lympho-'empty' plasmid provided a potent (Ͼ50% inhibition) and cytes in the antitumor response initiated by LPD. This repdose-dependent antitumor effect, compared with dextroseresents the first demonstration that an intravenously treated controls. In addition, i.v. injections of LPD containadministered cationic liposome-based nonviral vector can ing 'empty' plasmid also inhibited tumor growth in a subcutpromote a systemic, Th1-like innate immune response. aneous model of C3 fibrosarcomma. The antitumor effectThe immune adjuvant properties of LPD might prove to be correlated well with a strong and rapid proinflammatory suitable for delivering tumor-specific antigens in the context cytokine (TNF-␣, IL-12 and IFN-␥) response. Naked plasof DNA vaccination. mid DNA did not elicit a cytokine response and the

The role of tissue macrophages in the induction of proinflammatory cytokine production following intravenous injection of lipoplexes

“…The peak times for each cytokine were similar to those in previous reports. 4,5 In GdCl 3 -pretreated mice, the amounts of these cytokines were significantly lower than those in the saline-pretreated mice. The amount of TNF-␣, IFN-␥, and IL-12 in serum at the peak times was reduced 25-fold, three-fold, and 23-fold in GdCl 3 -pretreated mice, respectively.…”

“…[6][7][8] It appears that such cytokines activate the immune system and induce strong antitumor effects in mice. 4,5 On the other hand, Li et al have shown that such cytokines, which are secreted after intravenous injection of lipoplexes, can be toxic and cause short-term gene expression and refractory behavior on repeated dosing at frequent intervals. 9,10 This is a potential problem in gene therapy using plasmid DNA.…”

“…injection of the lipoplex, and the amounts of TNF-␣ in the cytoplasm fraction of each organ were examined as described in a previous report. 4 We have focused on TNF-␣ as a proinflammatory cytokine in the following experiments because it is secreted by macrophages after the phagocytosis of parasites and IFN-␥ is secondarily secreted by NK cells following stimulation of IL-12 and TNF-␣. 32 No TNF-␣ was observed in any organs or the serum of untreated mice (data not shown).…”

“…However, intravenous administration of cationic lipid vectors containing plasmid DNA has been shown to initiate potent cytokine responses. 4,5 High amounts of proinflammatory cytokines are produced in blood after lipoplex injection since bacterially derived plasmid DNA is recognized as foreign material by vertebrate cells. Unmethylated CpG sequences in plasmid DNA, occurring at a higher frequency in bacterial DNA, have been reported to have strong stimulatory effects on lymphocytes, NK cells, dendritic cells and macrophages…”

Effective transfer of interleukin-12 gene to solid tumors using a novel gene delivery system, poly [D,L-2,4-diaminobutyric acid]

“…However, such viral vectors can have some harmful antigenicity, and serious concerns have been raised regarding the use of viral vectors, especially in clinical trials. Nonviral carriers, such as cationic liposomes, 14,15 avoid such problems. Furthermore, nonviral vectors may be good clinical options because they may be more efficient and less labor intensive than viral vectors, although the present gene transfection efficiencies still need to be improved.…”