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Abstract: Purpose. : To evaluate a protocol that allowed the successful generation of DC and OVCA cells, fusion of these two cell types and assessment of stimulatory ability of the fusion cells for clinical use. Patients and methods. : Ovarian cancer (OVCA) cells and dendritic cells (DC) were isolated or generated from 22 patients with OVCA and subsequently fused with PEG. The stimulatory ability of fusion cells including T cell proliferation and induction of cytotocic T lymphocytes (CTL) was assessed. In addition, the impact of radiation, freezing and thawing of the fusion cells was evaluated. Results. : OVCA cells derived from 22 patients were successfully fused with autologous DC. The created heterokaryons expressed tumor-associated antigens, such as MUC1 and CA-125, and DC-derived MHC class II and costimulatory molecules. The fusion cells were functional in stimulating the proliferation of autologous T cells. In addition, CD4 and CD8 T cells derived from patients with ovarian cancer were stimulated by fusion cells and produced IFN-γ as demonstrated with intracellular staining. Significantly, T cells primed by fusion cells produced MHC class I-dependent lysis of autologous ovarian tumor cells. One cycle of fusion-cell stimulation can maintain the CTL activity up to 25 days. Conclusions. : The fusion of human OVCA cells and DC created immunogenic cells capable of stimulating CD4 and CD8 T cells. The effects of the processes required for preparing a vaccine for clinical use, including freezing and thawing and irradiation, do not interfere with the immunogenic properties of the fusion cells. [Copyright &y& Elsevier]

Fusion of human dendritic cells (DC) with tumor cells is an effective approach for delivering tumor antigens to DC, and DC/tumor fusion cells are potent stimulators of autologous T cells. However, the integration and morphology of DC/tumor fusion cells has not been examined. In the present study, we fused patient-derived DC to autologous breast or ovarian carcinoma cells. The fusion cells possessed the properties of both parent cells. After fusion, the cytoplasm of the two cells was integrated, whereas their nuclei remained separate entities. Colocalization of MUC1 peptide and HLA-DR molecules was observed on fusion cells under the immunoelectron microscope. Coculture of patient-derived peripheral blood mononuclear cells (PBMC) with DC/tumor fusion cells resulted in activation of CD4 and CD8 T cells as assessed by IFN-γ secretion, HLA-A*0201-MUC1 tetramer, and standard cytotoxic T lymphocyte (CTL) assays. The present study provides first evidence of integration of human DC and tumor cells and links their properties to T cell activation. [Copyright &y& Elsevier]