Lipoplexes prepared from cationic liposomes and mammalian DNA induce CpG-independent, direct cytotoxic effects in cell cultures and in mice.
BACKGROUND
Recent studies demonstrated the cytotoxic activity of bacterial DNA (pDNA) complexed with cationic lipids. This cytotoxicity is related to the ability of pDNA to induce potently the immune system, which is associated with release of inflammatory cytokines. Both activities seem to be related to the nonmethylated CpG sequences present in the pDNA. Here we study the cytotoxic activity of nonbacterial DNA complexed with cationic lipids against various tumor cell lines.


METHODS
Various nucleic acids complexed with cationic liposomes were prepared and their cytotoxic activity was studied in cell cultures and in tumor-bearing mice. Cell uptake of lipoplexes was evaluated, and mechanism of DNA cytotoxic activity was studied.


RESULTS
We found that nonbacterial (vertebrate) genomic DNA when complexed with cationic lipids is highly cytotoxic against C-26 and M-109 tumor cells. Cationic lipids alone were not toxic to these cells. The cytotoxic activity does not result from nonspecific acidification of the intracellular milieu, as substitution of DNA by poly-L-glutamate did not result in cytotoxicity, although the level of uptake of anionic charges per cell was similar to that of the nucleic acids, suggesting that this cytotoxic effect is specific to nucleic acids. By studying the nucleic acid fate using confocal microscopy, we found that cytotoxicity correlated with the release of DNA into the cytoplasm following uptake of lipoplexes. Injection of calf thymus DNA-based lipoplexes to mice with peritoneal C-26 metastases resulted in doubling of median survival time and long-term survival in 20% of the tumor-bearing mice. Judging by low levels of IFN-gamma, TNF-alpha and IL-6 in the treated mice, this effect cannot be ascribed to Th-1 inflammation, but rather to a direct cytotoxic effect on the tumor cells.


CONCLUSIONS
The above data provide a new insight into the mechanisms of lipoplex-mediated antitumor effects in vitro and in vivo and new perspectives in cancer therapy.
Khazanov
Elena
E
Laboratory of Membrane and Liposome Research, Department of Biochemistry, The Hebrew University-Hadassah Medical School Jerusalem, Israel. khazanov@md.huji.ac.il
Simberg
Dmitri
D
Barenholz
Yechezkel
Y
D000818:Animals; D000970:Antineoplastic Agents / Q000008:administration & dosage / Q000378:metabolism*; D002412:Cations / Q000737:chemistry; D002417:Cattle; D045744:Cell Line, Tumor; D018899:CpG Islands / Q000235:genetics*; D004247:DNA / Q000737:chemistry / Q000145:classification / Q000235:genetics / Q000378:metabolism* / Q000633:toxicity*; D005229:Fatty Acids, Monounsaturated; D005260:Female; D005456:Fluorescent Dyes; D018014:Gene Transfer Techniques; D015316:Genetic Therapy; D006801:Humans; D066298:In Vitro Techniques; D008081:Liposomes / Q000008:administration & dosage / Q000737:chemistry / Q000378:metabolism*; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D041681:NIH 3T3 Cells; D009362:Neoplasm Metastasis; D009838:Oligodeoxyribonucleotides / Q000737:chemistry / Q000145:classification / Q000235:genetics / Q000378:metabolism / Q000633:toxicity; D010714:Phosphatidylethanolamines; D000644:Quaternary Ammonium Compounds; D012474:Salmon; D013094:Spermatozoa / Q000737:chemistry / Q000145:classification / Q000378:metabolism; D013997:Time Factors
C410262:1,2-dioleoyl-glycero-3-phosphatidyl ethanolamine; D000970:Antineoplastic Agents; D002412:Cations; D005229:Fatty Acids, Monounsaturated; D005456:Fluorescent Dyes; D008081:Liposomes; D009838:Oligodeoxyribonucleotides; D010714:Phosphatidylethanolamines; D000644:Quaternary Ammonium Compounds; D004247:DNA; C427203:calf thymus DNA; C070046:1,2-dioleoyloxy-3-(trimethylammonium)propane