Treatment of ovarian cancer cell lines with 5-aza-2′-deoxycytidine upregulates the expression of cancer-testis antigens and class I major histocompatibility complex-encoded molecules
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To test the hypothesis that decrease in DNA methylation will increase the expression of cancer-testis antigens (CTA) and class I major histocompatibility complex (MHC)-encoded molecules by ovarian cancer cells, and thus increase the ability of these cells to be recognized by antigen-reactive CD8+ T cells.
Human ovarian cancer cell lines were cultured in the presence or absence of varying concentrations of the DNA demethylating agent 5-aza-2′-deoxycytidine (DAC) for 3–7 days. The expression levels of 12 CTA genes were measured using the polymerase chain reaction. The protein expression levels of class I MHC molecules and MAGE-A1 were measured by flow cytometry. T cell reactivity was determined using interferon-γ ELISpot analysis.
DAC treatment of ovarian cancer cell lines increased the expression of 11 of 12 CTA genes tested including MAGE-A1, MAGE-A3, MAGE-A4, MAGE-A6, MAGE-A10, MAGE-A12, NY-ESO-1, TAG-1, TAG-2a, TAG-2b, and TAG-2c. In contrast, DAC treatment decreased the already low expression of the MAGE-A2 gene by ovarian cancer cells, a finding not previously observed in cancers of any histological type. DAC treatment increases the expression of class I MHC molecules by the cells. These effects were time-dependent over a 7-day interval, and were dose-dependent up to 1–3 μM for CTA and up to 10 μM for class I MHC molecules. Each cell line tested had a unique pattern of gene upregulation after exposure to DAC. The enhanced expression levels increased the recognition of 2 of 3 antigens recognized by antigen-reactive CD8+ T cells.
These results demonstrate the potential utility of combining DAC therapy with vaccine therapy in an attempt to induce the expression of antigens targeted by the vaccine, but they also demonstrate that care must be taken to target inducible antigens.
KeywordsOvarian cancer Class I MHC molecules Cancer-testis antigens DNA methylation 5-Aza-2′-deoxycytidine
Cytotoxic T lymphocyte
Major histocompatibility complex
Polymerase chain reaction
This work was supported by grant W81XWH-05-1-0012 from the United States Department of Defense to K. T. Hogan. The authors wish to thank Dr. Craig L. Slingluff Jr. for reviewing the manuscript and the members of the UVA Human Immune Therapy Center core laboratory for performing the ELISpot analyses.
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