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Human U251MG glioma cells (HLA-A2+) retrovirally transduced with the human gene for the membrane form of macrophage colony stimulating factor (mM-CSF) were investigated. The clones, MG-2F11 and MG-2C4, that expressed the most mM-CSF, but not the viral vector or the parental U251MG cells, were killed by both murine and human monocyte/macrophages in cytotoxicity assays. MG-2F11 cells failed to form subcutaneous tumors in either nude or NIH-bg-nu-xidBR mice, while mice inoculated with the U251MG viral vector (MG-VV) cells developed tumors. Electron microscopy studies showed that four hours after subcutaneous injection, the mM-CSF-transduced cells began dying of a process that resembled paraptosis. The dying tumor cells were swollen and had extensive vacuolization of their mitochondria and endoplasm reticulum. This killing process was complete within 24 hours. Macrophage-like cells were immediately adjacent to the killed MG-2F11 cells. Immunohistological staining for the heat shock proteins, HSP60, HSP70 and GRP94(gp96) showed that 18 hours after inoculation into nude mice, the MG-2F11 injection site were two to four times more intensely stained than the MG-VV cells. SCID/beige mice depleted of their mouse macrophages, allowed the U251-2F11 cells to form subcutaneous tumors. Injecting HLA-A2 matched PBMC allowed us to investigate whether human immune responses could be generated in vivo against the mM-CSF transduced U251 cells. In 3 experiments, human immune responses could be induced which limited the growth of the U251 tumors in vivo. Histology confirmed loss of tumor cells and activated lymphocytes were present. Human CD4+ T cells could be isolated and grown out of the tumors that were vaccinated by the mM-CSF U251 tumor cells. This study shows that human gliomas transduced with mM-CSF have the potential to be used as a safe live tumor cell vaccine.

[Proc Amer Assoc Cancer Res, Volume 46, 2005]