Korean J Hematol.  2007 Jun;42(2):136-145. 10.5045/kjh.2007.42.2.136.

Anti-tumor Cytotoxicity of Allogeneic Neuroblastoma Tumor Antigen-loaded Dendiritic Cells

• Park HJ
• Shin HY
• Ahn HS

• KMID: 2083511
• DOI: http://doi.org/10.5045/kjh.2007.42.2.136

Abstract

BACKGROUND: Despite of aggressive treatments, the long-term survival rate is about 30% in stage 4 neuroblastoma (NBL). Recently, dendritic cell (DC)-based immunotherapy is emerging as a promising tool in cancer treatment. But it is very difficult to get sufficient amount of autologous tumor as the source of tumor antigen in DC-based immunotherapy. The purpose of this study was to test whether DCs loaded with allogeneic NBL tumor antigens can prime effective anti-tumor immune responses.
METHODS
Peripheral blood mononuclear cells (PBMCs) were differentiated into immature DCs in the presence of GM-CSF and IL-4. As the source of tumor antigens, human neuroblastoma cell lines, SK-N-MC, SK-N-SH, and IMR-32, were used after induction of apoptosis by UV irradiation. The immature DCs were loaded with apoptotic tumor cells, and then cultured with PBMCs for priming the tumor antigen-specific T lymphocytes. The tumor-specific cytotoxicity of T lymphocytes against NBL cells was analysed after coculture.
RESULTS
Incubation of DCs with apoptotic tumor cells effectively loaded DCs with tumor antigens and induced maturation of DCs. The tumor antigen-challenged T lymphocytes effectively killed the NBL cells, which were used as tumor antigens. Furthermore, the T lymphocytes showed a broad ranged cytotoxicity to all of the NBL cell lines, which were not challenged as tumor antigens.
CONCLUSION
This study showed that the apoptotic NBL tumor cells induced maturation of DCs and could be used as tumor antigens, and DCs loaded with apoptotic NBL tumor cells could induce effective anti-tumor specific cytotoxic T lymphocytes to allogeneic NBL tumors.

Keyword

Dendritic cell; Immunotherapy; Allogeneic tumor antigen; Neuroblastoma

MeSH Terms

Antigens, Neoplasm
Apoptosis
Cell Line
Coculture Techniques
Dendritic Cells
Granulocyte-Macrophage Colony-Stimulating Factor
Humans
Immunotherapy
Interleukin-4
Neuroblastoma*
Survival Rate
T-Lymphocytes
T-Lymphocytes, Cytotoxic
Antigens, Neoplasm
Granulocyte-Macrophage Colony-Stimulating Factor
Interleukin-4

Figure

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  Fig. 1 Scheme of the experimental design for stimulation of T lymphocytes by tumor antigen-loaded dendritic cells.

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  Fig. 2 Photomicrograph of peripheral blood monocyte- derived immature (left, ×400) and mature human dendritic cells (right, ×1,000), Papanicolaou staining.

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  Fig. 3 Phenotypic analysis of peripheral blood monocyte-derived dendritic cells (PB-DC) by two color flow cytometry analysis using MoAb combinations of CH6, a new monoclonal antibody against human dendritic cells, and other markers of human dendritic cells.

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  Fig. 4 UV-induced apoptosis of tumor cells (SK-N-MC). Apoptosis of the tumor cells was induced by UV irradiation for 2 hours at room temperature. Apoptosis and cell death were analysed by staining the cells with annexin V.

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  Fig. 5 Immunophenotypic changes of peripheral blood monocyte-derived dendrtitic cells before and after tumor antigen loading with apoptotic tumor cells (SK-N-SH).

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  Fig. 6 Tumor-specific cytotoxicity by tumor antigen- challenged T lymphocytes. T lymphocytes were stimulated by tumor (SK-N-MC) antigen-loaded dendritic cells, and analysed for the tumor-specific cytotoxicity against three NBL cell lines; SK-N-MC (●—●), SK-N-SH (■—■), and IMR-32 (▲—▲), and two control cell lines; SK-BR-3 (△—△) and K-562 (○—○) at various effecto r：target (E：T) ratios.

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  Fig. 7 Tumor-directed cytotoxicity of T lymphocytes against various tumor cell lines at various effector：target (E：T) ratios. T lymphocytes from different donors were stimulated by DCs loaded with apoptotic SK-N-MC cells, and tumor-specific cytotoxic effector function was measured.

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  Fig. 8 Tumor-directed cytotoxicity of T lymphocytes against various tumor cell lines at various effector：target (E：T) ratios. T lymphocytes were stimulated by DCs loaded with apoptotic SK-N-SH cells, and tumor-specific cytotoxic effector function was measured.

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  Fig. 9 Expression of MHC class I and II MoAb on various tumor cell lines.

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  Fig. 10 Comparison of the extent of cytotoxicity in relation with different antigen sources.

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