Generation of tumor cell lysate-loaded dendritic cells preprogrammed for IL-12 production and augmented T cell response

Abstract.

Dendritic cells (DC) loaded with tumor associated antigens (TAA) are often used for the vaccination of cancer patients; however methodologies for the vaccine preparation have not yet been standardized. The purpose of this work was to optimize the ex-vivo production of functional TAA-loaded DC that would produce interleukin-2 (IL-12) and enhance the T cell response. We generated ex-vivo DC from human monocytes with granulocyte macrophage-colony stimulating factor (GM-CSF) and IL-4, and whole necrotic tumor cells (cell lysates) of cancer cell lines were used as model TAA. DC were loaded with lysates without or with additional tumor necrosis factor-α (TNF-α), or cytokine combination treatments and tested for functional ability in vitro. Tumor cell lysates alone did not fully mature DC either phenotypically or functionally. After antigen uptake additional maturation signals were necessary. TNF-α matured DC phenotypically, but additional interferon-γ (IFN-γ) treatment was necessary to achieve functional maturation, the production of significant amounts of IL-12. Since IL-12 production by DC increased during the first 24 h of maturation and declined by 48 h, proper timing of the ex-vivo DC treatment was crucial for the generation of functionally mature antigen-loaded DC. Our results suggest that after allowing 4 h of tumor lysate uptake by immature DC, further treatment with TNF-α and IFN-γ for 24 h provides the optimal conditions to obtain functional TAA-loaded DC. These TAA-loaded cytokine pretreated DC then prime naïve T cells, and enhance both T helper 1 (Th1), Th2 and cytotoxic T lymphocyte (CTL) responses, that are necessary to achieve an effective, specific anti-tumor response.