Filho Pedro A Andrade, López-Albaitero Andrés, Xi Liqiang, Gooding William, Godfrey Tony, Ferris Robert L
Department of Otolaryngology, University of Pittsburgh Cancer Institute, Pittsburgh, PA, USA.
Int J Cancer. 2009 Oct 15;125(8):1912-20. doi: 10.1002/ijc.24590.
The MAGE antigens are frequently expressed cancer vaccine targets. However, quantitative analysis of MAGE expression in upper aerodigestive tract (UADT) tumor cells and its association with T-cell recognition has not been performed, hindering the selection of appropriate candidates for MAGE-specific immunotherapy. Using quantitative RT-PCR (QRT-PCR), we evaluated the expression of MAGE-3/6 in 65 UADT cancers, 48 normal samples from tumor matched sites and 7 HLA-A0201+ squamous cell carcinoma of the head and neck (SCCHN) cell lines. Expression results were confirmed using Western blot. HLA-A0201:MAGE-3- (271-279) specific cytotoxic T lymphocytes (MAGE-CTL) from SCCHN patients and healthy donors showed that MAGE-3/6 expression was highly associated with CTL recognition in vitro. On the basis of the MAGE-3/6 expression, we could identify 31 (47%) of the 65 UADT tumors, which appeared to express MAGE-3/6 at levels that correlated with efficient CTL recognition. To confirm that the level of MAGE-3 expression was responsible for CTL recognition, 2 MAGE-3/6 mRNA(high) SCCHN cell lines, PCI-13 and PCI-30, were subjected to MAGE-3/6-specific knockdown. RNAi-transfected cells showed that MAGE expression and MAGE-CTL recognition were significantly reduced. Furthermore, treatment of cells expressing low MAGE-3/6 mRNA with a demethylating agent, 5-aza-2'-deoxycytidine (DAC), increased the expression of MAGE-3/6 and CTL recognition. Thus, using QRT-PCR UADT cancers frequently express MAGE-3/6 at levels sufficient for CTL recognition, supporting the use of a QRT-PCR-based assay for the selection of candidates likely to respond to MAGE-3/6 immunotherapy. Demethylating agents could increase the number of patients amenable for targeting epigenetically modified tumor antigens in vaccine trials.
黑色素瘤相关抗原（MAGE）是常用的癌症疫苗靶点。然而，尚未对上消化道（UADT）肿瘤细胞中MAGE表达进行定量分析及其与T细胞识别的关联，这阻碍了MAGE特异性免疫疗法合适候选者的选择。我们采用定量逆转录聚合酶链反应（QRT-PCR），评估了65例UADT癌症、48例肿瘤匹配部位的正常样本以及7株HLA-A0201+头颈部鳞状细胞癌（SCCHN）细胞系中MAGE-3/6的表达。通过蛋白质免疫印迹法确认表达结果。来自SCCHN患者和健康供体的HLA-A0201:MAGE-3-（271-279）特异性细胞毒性T淋巴细胞（MAGE-CTL）表明，MAGE-3/6表达与体外CTL识别高度相关。基于MAGE-3/6表达，我们在65例UADT肿瘤中鉴定出31例（47%），其MAGE-3/6表达水平似乎与有效的CTL识别相关。为了证实MAGE-3表达水平是CTL识别的原因，对2株MAGE-3/6 mRNA（高表达）SCCHN细胞系PCI-13和PCI-30进行了MAGE-3/6特异性敲低。RNAi转染细胞显示MAGE表达和MAGE-CTL识别显著降低。此外，用去甲基化剂5-氮杂-2'-脱氧胞苷（DAC）处理低表达MAGE-3/6 mRNA的细胞，可增加MAGE-3/6的表达和CTL识别。因此，使用QRT-PCR检测发现UADT癌症中MAGE-3/6的表达水平通常足以进行CTL识别，这支持使用基于QRT-PCR的检测方法来选择可能对MAGE-3/6免疫疗法有反应的候选者。去甲基化剂可增加疫苗试验中适合靶向表观遗传修饰肿瘤抗原的患者数量。
