目的: 评价修饰的CEA610D抗原肽疫苗体外抗肿瘤免疫的有效性,为其临床应用提供依据。方法： 多肽固相合成法合成HLAA2 CEA修饰肽(CEA610D)、HLAA2天然肽CEA605613（天然肽）和HLAA2无关肽MAGE3（无关肽），采用同种异体混合淋巴细胞与肽共孵育的方法，诱导肽特异性细胞毒性T淋巴细胞(CTL)，利用MTT法检测CTL的增殖和杀伤活性；流式细胞术观察细胞表型；RTPCR检测细胞穿孔素的表达；ELISA法检测CTL上清中IFNγ的水平。结果：CEA610D疫苗产生肽特异性CTL的能力最强（P<0.05），其CD8+T细胞数也高于天然肽组和无关肽组；在效靶比为40〖DK〗∶1时，CEA610D和天然肽所诱导的CTL对CEA+HLAA2限制性人结肠癌细胞T84的杀伤活性可达(56.7±3.73)%和(51.2±1.86)%，而3种肽特异性CTL细胞对CEA+HLAA2人结肠癌lovo细胞杀伤活性维持在本底水平；CEA610D组的CTL所释放的杀伤相关介质穿孔素和上清中IFNγ的水平也显著高于其余两组（P<0.01）。结论：与天然肽相比，CEA610D疫苗可打破自身表位的免疫耐受，在体外抗肿瘤免疫中更具优势。

Objective:To investigated the in vivo efficacy of altered CEA610D peptide vaccine against tumor, so as to provide a basis for its clinical use.Methods:Altered CEA peptide CEA610D, natural CEA peptide CEA605613 and MAGE3 peptide were synthesized by polypeptide solidphase synthesis system. The cytotoxicity T lymphocytes (CTLs) were induced by autologous mixed lymphocytes reaction implused with above peptides; proliferation and cytotoxicity of different CTLs were measured by MTT; phenotypes of the CTLs were detected by flow cytometry; expression of perforin in different CTLs were assayed by RTPCR; IFNγ levels in the supernatants of different CTLs were detected by ELISA.Results: CEA610D peptide more efficiently induced CTL than CEA605613 and MAGE3 peptide did (P<0.05). The number of CD8+T cells in CEA610D group was significantly larger than those in CEA605613 and MAGE3 groups (P<005). When the E∶T was 40∶1, the cytotoxicity of CTLs induced by CEA610D and CEA605613 against CEA+HLAA2+ T84 cells were (56.7±373)% and (51.2±1.86)%, respectively. But the CTL cytotoxicities induced by the three peptides against CEA+HLAA2Lovo cells were all at the background level. Perforin expression and IFNγ level of CTLs in CEA610D group were significantly higher than those in the other two groups (P<0.01).Conclusion:Compared with natural CEA605613 peptide, altered CEA610D peptide can effectively break the immune tolerance to self peptide, and thus has a stronger antitumor activity in vitro.

山东省医学科学院科研项目(No. 200625)