Summary Dendritic cells (DCs) loaded with tumour antigens have been successfully used to induce protective tumour immunity in murine models and human trials. However, it is still unclear which DC administration route elicits a superior therapeutic effect. Herein, we investigated the vaccine efficiency of DC2.4 cells, a murine dendritic cell line, pulsed with ovalbumin (OVA) in the murine E.G7-OVA tumour model after immunization via various routes. After a single vaccination using 1 × 10 6 OVA-pulsed DC2.4 cells, tumour was completely rejected in the intradermally (i.d.; three of four mice), subcutaneously (s.c.; three of four mice), and intraperitoneally (i.p.; one of four mice) immunized groups. Double vaccinations enhanced the anti-tumour effect in all groups except the intravenous (i.v.) group, which failed to achieve complete rejection. The anti-tumour efficacy of each immunization route was correlated with the OVA-specific cytotoxic T lymphocyte (CTL) activity evaluated on day 7 post-vaccination. Furthermore, the accumulation of DC2.4 cells in the regional lymph nodes was detected only in the i.d.-and s.c.-injected groups. These results demonstrate that the administration route of antigen-loaded DCs affects the migration of DCs to lymphoid tissues and the magnitude of antigen-specific CTL response. Furthermore, the immunization route affects vaccine efficiency.

Early studies in mice demonstrated that tumor-specific cytotoxic T lymphocytes (CTLs) could control tumor growth and metastasis. The identification of T cell-recognizing tumor-associated antigens (TAAs) in human cancer, particularly in melanoma (i.e. MAGE, MART-1, gp100, tyrosinase, and TRP), 1,2) facilitated the development of cancer immunotherapy based on TAA-vaccination with adjuvants to elicit tumor-specific CTLs.3) However, this immunological approach limits the application of this system only to certain cancer patients because TAAs are not yet identified for most of human cancers. Additionally, the expression levels of known TAAs that may be applicable for immunotherapy vary between tumor cells isolated from patients with cancer. 4)Therefore, it is very difficult to predict which TAA would generate an effective anti-tumor immune response that would make it appropriate for use as a vaccine component for a specific patient.To overcome this limitation, several researchers have attempted to develop a vaccine strategy using tumor cell-lysate (TCL) as a possible source of TAA.5) The use of TCL prepared from surgically removed tumors is a promising approach to induce a broader T cell-immune response not only to defined TAAs but also to unknown TAAs. In TCL-based cancer immunotherapy, the development of both an antigendelivery system and an adjuvant that can efficiently prime and propagate CTLs specific for TAAs included in the TCL is required for achieving sufficient therapeutic effect. CTLs are activated by antigen-presenting cells (APCs), including dendritic cells (DCs), through the major histocompatibility complex (MHC) class I-restricted antigen presentation pathway. Peptides presented on MHC class I molecules are derived in most situations exclusively from endogenous antigens synthesized by cells. Antigens in the extracellular fluids fail to gain access to the MHC class I-pathway in most cells, although class I-presentation of endocytosed antigens also occurs in APCs under certain circumstances.6,7) Therefore, if we can introduce the TAA-containing TCL directly into the cytoplasm, the TAAs would be definitively delivered to the MHC class I-antigen presentation pathway, much like cytoplasmic proteins.Fusion active liposomes (fusogenic liposomes; FLs), which are composed of conventional liposomes (CLs) displaying Sendai virus-accessory proteins, retain membranefusion activity derived from Sendai-virus and efficiently introduce its contents into cytoplasm. 8) We have previously reported that direct antigen loading into cytoplasm by FLs is an efficient approach for enhancing antigen-specific CTL induction in mice. [9][10][11] In the present study, in order to evaluate the usefulness of FLs as antigen-delivery carriers for TCL-based cancer immunotherapy, we investigated anti-tumor efficacy of ex vivo vaccination using TCL-containing FLs (TCL/FLs)-pulsed DCs and in vivo direct TCL/FLs-immunization in the murine B16BL6 melanoma model. MATERIALS AND METHODSCells and Mice B16BL6 cells, a C57BL/6-origin melanoma cell ...