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Although unmethylated CpG dinucleotide-containing oligodeoxynucleotides (CpG ODN) are able to inhibit tumor metastasis through the induction of antitumor immunity, their stability and delivery to antigen presenting cells needs to be improved. In this study, we formulated a CpG ODN complex with cationic liposomes (CpG ODN-lipoplex) and its antitumor activity was evaluated in peritoneal dissemination models of tumor cells stably labeled with firefly luciferase gene. A single intraperitoneal administration of CpG ODN-lipoplex greatly reduced the number of tumor cells to 0.01% or lower compared with that detected in untreated mice, which may be associated with increased production of TNF-α and IL-12. CpG ODN-lipoplex increased the survival time of the tumor-bearing mice, and most long-term survivors rejected rechallenged tumor cells. These results indicate that a single dosing of CpG ODN-lipoplex is effective in inhibiting peritoneal dissemination and inducing long-lasting antitumor immunity.