MAGE-A1, MAGE-A3, and NY-ESO-1 can be upregulated on neuroblastoma cells to facilitate cytotoxic T lymphocyte-mediated tumor cell killing

Approximately half of patients with stage IV neuroblastoma are expected to relapse despite current therapy, and when this occurs, there is little likelihood of achieving a cure. Very few clinical trials have been conducted to determine whether cellular immune responses could be harnessed to fight this tumor, largely because potential tumor antigens for cytotoxic T lymphocytes (CTL) are limited. MAGE-A1, MAGE-A3, and NY-ESO-1 are cancer-testis (CT) antigens expressed on a number of malignant solid tumors, including neuroblastoma, but many tumor cell lines down-regulate the expression of CT antigens as well as major histocompatibility (MHC) antigens, precluding recognition by antigen-specific T cells. If expression of cancer antigens on neuroblastoma could be enhanced pharmacologically, CT antigen-specific immunotherapy could be considered for this tumor. We have demonstrated that the expression of MAGE-A1, MAGE-A3, and NY-ESO-1 can be upregulated on neuroblastoma cells following exposure to pharmacologic levels of the demethylating agent 5-aza-2'-deoxycytidine (decitabine, DAC). Expression of NY-ESO-1, MAGE-A1, or MAGE-A3 was induced in 10/10 neuroblastoma cell lines after 5 days of exposure to DAC. Culture of neuroblastoma cell lines with IFN-gamma was also associated with an increased expression of either MHC Class I or II by cytofluorometry, as reported by other groups. MAGE-A1, MAGE-A3, and NY-ESO-1-specific CTL were cultured from volunteer donors by stimulating peripheral blood mononuclear cells with dendritic cells pulsed with overlapping peptide mixes derived from full-length proteins, and these CTL preferentially lysed HLA partially matched, DAC-treated neuroblastoma and glioblastoma cell lines. These studies show that demethylating chemotherapy can be combined with IFN-gamma to increase the expression of CT antigens and MHC molecules on neuroblastoma cells, and pre-treatment with these agents makes tumor cell lines more susceptible to CTL-mediated killing. These data provide a basis to consider the use of demethylating chemotherapy in neuroblastoma patients, in conjunction with immune therapies that facilitate the expansion of CT antigen-specific CTL.