car-t免疫技术治疗肿瘤好文分享

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文章来源: http://www.ncbi.nlm.nih.gov/pubmed/26447882

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-氮杂-2’脱氧胞苷处理诱导NY-ESO-1的表达和T淋巴细胞介导的促肿瘤细胞杀伤

背景：

NY-ESO-1属于肿瘤、睾丸抗原（CTA）家族，在癌症的免疫治疗中，是一个令人瞩目的靶点。在一些实体肿瘤中，通过CpG岛启动子对DNA的去甲基化，NY-ESO-1的表达被诱导。然而，在某些肿瘤中，如乳腺癌和多发性骨髓瘤中，NY-ESO-1表达很低或者不表达。因此，我们建立了一个优化的体外治疗方案，使用去甲基化剂5-氮杂-2'-脱氧胞苷（DAC），来增加肿瘤细胞NYESO-1的表达上调。

方法/主要的结果：

我们证实了NY-ESO-1会在MCF7乳腺癌细胞中重新诱导的和在U266多发骨髓瘤中表达明显增加。这种效果是时间和剂量依赖的，NY-ESO-1 mRNA会在10μM DAC处理72小时之后表现出最高的表达量。NY-ESO-1的活化也同样由western印迹、流式细胞仪检测、免疫荧光染色在蛋白质水平的监测得以确定。在HLA-A*0201分子中，对癌细胞表面的NY-ESO-1单链多肽进行检测与定量，MCF7和U266 细胞分别显示出了100%和50%的增长。此外，细胞表面NY-ESO-1来源多肽的表达增强，伴随着HLA-A*0201/NY-ESO-1(157–165)多肽特异性嵌合抗原（CAR）CD8+T细胞具有一个增强的特异性溶解MCF7 和 U266细胞的能力。此外，CAR-T细胞的杀伤能力与高水平干扰素的分泌有关。

结论：

这个结果表明，NY-ESO-1指导的CAR-T细胞免疫治疗可能会从同时使用5-氮杂-2’脱氧胞苷（DAC）中受益。