5-氮雜-2’脫氧胞苷處理誘導(dǎo)NY-ESO-1的表達(dá)和T淋巴細(xì)胞介導(dǎo)的促腫瘤細(xì)胞殺傷

Treatment with 5-Aza-2'-Deoxycytidine Induces Expression of NY-ESO-1 and Facilitates Cytotoxic T Lymphocyte-Mediated Tumor Cell Killing
Background
NY-ESO-1 belongs to the cancer/testis antigen (CTA) family and represents an attractive
target for cancer immunotherapy. Its expression is induced in a variety of solid tumors via DNA demethylation of the promoter of CpG islands. However, NY-ESO-1 expression is usually very low or absent in some tumors such as breast cancer or multiple myeloma. Therefore, we established an optimized in vitro treatment protocol for up-regulation of NYESO-1 expression by tumor cells using the hypomethylating agent 5-aza-2'-deoxycytidine(DAC).
Methodology/Principal Findings
We demonstrated de novo induction of NY-ESO-1 in MCF7 breast cancer cells and significantly increased expression in U266 multiple myeloma cells. This effect was time- and dose-dependent with the highest expression of NY-ESO-1 mRNA achieved by the incubation of 10 μM DAC for 72 hours. NY-ESO-1 activation was also confirmed at the protein level as shown by Western blot, flow cytometry, and immunofluorescence staining. The detection and quantification of single NY-ESO-1 peptides presented at the tumor cell surface in the context of HLA-A*0201 molecules revealed an increase of 100% and 50% for MCF7 and U266 cells, respectively. Moreover, the enhanced expression of NY-ESO-1 derived peptides at the cell surface was accompanied by an increased specific lysis of MCF7 and U266 cells by HLA-A*0201/NY-ESO-1(157–165) peptide specific chimeric antigen receptor (CAR) CD8+ T cells. In addition, the killing activity of CAR T cells correlated with the secretion of higher IFN-gamma levels.
Conclusions/Significance
These results indicate that NY-ESO-1 directed immunotherapy with specific CAR T cells
might benefit from concomitant DAC treatment.
5-氮雜-2’脫氧胞苷處理誘導(dǎo)NY-ESO-1的表達(dá)和T淋巴細(xì)胞介導(dǎo)的促腫瘤細(xì)胞殺傷
背景：
NY-ESO-1屬于腫瘤、抗原（CTA）家族，在癌癥的免疫治療中，是一個令人矚目的靶點。在一些實體腫瘤中，通過CpG島啟動子對DNA的去甲基化，NY-ESO-1的表達(dá)被誘導(dǎo)。然而，在某些腫瘤中，如癌和多發(fā)性骨髓瘤中，NY-ESO-1表達(dá)很低或者不表達(dá)。因此，我們建立了一個優(yōu)化的體外治療方案，使用去甲基化劑5-氮雜-2'-脫氧胞苷（DAC），來增加腫瘤細(xì)胞NYESO-1的表達(dá)上調(diào)。
方法/主要的結(jié)果：
我們證實了NY-ESO-1會在MCF7癌細(xì)胞中重新誘導(dǎo)的和在U266多發(fā)骨髓瘤中表達(dá)明顯增加。這種效果是時間和劑量依賴的，NY-ESO-1 mRNA會在10μM DAC處理72小時之后表現(xiàn)出zui高的表達(dá)量。NY-ESO-1的活化也同樣由western印跡、流式細(xì)胞儀檢測、免疫熒光染色在蛋白質(zhì)水平的監(jiān)測得以確定。在HLA-A*0201分子中，對癌細(xì)胞表面的NY-ESO-1單鏈多肽進(jìn)行檢測與定量，MCF7和U266 細(xì)胞分別顯示出了100%和50%的增長。此外，細(xì)胞表面NY-ESO-1來源多肽的表達(dá)增強，伴隨著HLA-A*0201/NY-ESO-1(157–165)多肽特異性嵌合抗原（CAR）CD8+T細(xì)胞具有一個增強的特異性溶解MCF7 和 U266細(xì)胞的能力。此外，CAR-T細(xì)胞的殺傷能力與高水平干擾素的分泌有關(guān)。
結(jié)論：
這個結(jié)果表明，NY-ESO-1指導(dǎo)的CAR-T細(xì)胞免疫治療可能會從同時使用5-氮雜-2’脫氧胞苷（DAC）中受益。