一种特异性靶向性淋巴细胞对荷瘤裸鼠的治疗观察

Observation of targeting lymphocytes to therapy nude mice bearing KATOⅢ gastric cancer cell

目的观察癌胚抗原(CEA)阳性靶向性淋巴细胞对胃癌细胞体内及体外靶向性治疗作用。方法从健康人外周血中分离单个核细胞(PBMC),通过脂质体lipofectamine 2000介导的细胞转染将重组真核表达载体anti-CEA-sc Fv-CD3ζ-pc DNA3.0介导的细胞转染导入PBMC,以获取CEA阳性靶向性淋巴细胞(以下简称靶淋巴细胞);将不含anti-CEA-sc Fv-CD3ζ-pc DNA3.0融合基因片段的pc DNA3.0真核表达载体转染入PBMC,获取的淋巴细胞简称为空载体淋巴细胞。将上述两种淋巴细胞分别与CEA阳性KATOⅢ胃癌细胞和CEA阴性BGC-823胃癌细胞共培养,观察不同靶淋巴细胞与胃癌细胞共培养(24 h或36 h)后的识别情况或其对胃癌细胞凋亡的影响。并将上述已获取的两种淋巴细胞分别通过尾静脉注入荷CEA阳性KATOⅢ胃癌细胞裸鼠和荷CEA阴性BGC-823胃癌细胞裸鼠体内,以观察其对荷胃癌裸鼠肿瘤生长情况的影响。结果 (1)靶淋巴细胞与胃癌细胞共培养24 h时,靶淋巴细胞对KATOⅢ胃癌细胞的识别率为72.3%,淋巴细胞对KATOⅢ胃癌细胞的识别能力较强;其对BGC-823胃癌细胞识别率为7.8%,其对BGC-823胃癌细胞识别能力较弱。(2)当靶淋巴细胞、空载体淋巴细胞分别与KATOⅢ和BGC-823胃癌细胞分别共培养36 h时,靶淋巴细胞+KATOⅢ胃癌细胞组的凋亡率明显高于空载体淋巴细胞+KATOⅢ胃癌细胞组(P=0.032),而靶淋巴细胞+BGC-823胃癌细胞组和空载体淋巴细胞+BGC-823胃癌细胞组的凋亡率比较差异无统计学意义(P=0.118)。(3)在观察40 d内,靶淋巴细胞+KATOⅢ胃癌细胞组的肿瘤体积明显小于空载体淋巴细胞+KATOⅢ胃癌细胞组(F=5.010,P<0.01)和空白对照组(F=7.560,P<0.01);靶淋巴细胞+BGC-823胃癌细胞组与空载体淋巴细胞+BGC-823胃癌细胞组间比较差异无统计学意义(F=1.210,P>0.05)。靶淋巴细胞+KATOⅢ胃癌细胞组的肿瘤体积明显小于靶淋巴细胞+BGC-823胃癌细胞组(F=4.982,P<0.01)。结论 CEA阳性靶向性淋巴细胞可特异性促进CEA阳性胃癌细胞凋亡,抑制荷CEA阳性胃癌细胞裸鼠肿瘤的生长。

Objective To study effect of carcinoembryonic antigen （CEA） positive targeted lymphocytes on gastric cancer ceils in vitro and in vivo. Methods The peripheral blood mononuclear cells （PBMCs） were isolated from the peripheral blood of healthy volunteers. The recombinant vector anti-CEA-scFv-CD3（-pcDNA3.0 was transfected into the PBMCs by lipofectamine 2000, by this means, the CEA special lymphocytes were obtained. Meanwhile, the PBMCs transfected with empty plasmid pcDNA3.0 were used as control （empty vector lymphocytes）. The different lymphocytes and gastric cancer calls （CEA positive KATO HI gastric cancer cells and CEA negative BGC-823 gastric cancer cells） were co-cultured, then the ability to identify the gastric cancer cells and it＇s effect on apoptosis of gastric cancer ceUs were observed at 24 h or 36 h later respectively. The CEA special lymphocytes and empty vector lymphocytes were injected by the tail vein of nude mice bearing gastric cancer cells, then it＇s effect on the tumor was observed. Results ① The CEA special lymphocytes could strongly identify the KATO HI gastric cancer cells （identification rate was 72.3%）, which could weakly identify the BGC-823 gastric cancer cells （identification rate was 7.8%）. ② The apoptosis rate of the co- culture of CEA special lymphocytes and KATO Ⅲ gastric cancer cells was significantly higher than that of the co-culture of empty vector lymphocytes and KATO Ill gastric cancer cells （P=0.032）, which had no significant difference between the co-culture of CEA special lymphocytes and BGC-823 gastric cancer cells and the co-culture of empty vector lymphocytes and BGC-823 gastric cancer cells （P=0.118）. ③ The tumor volume of the co-culture of CEA special lymphocytes and KATO Ⅲ gastric cancer cells was significantly smaller than that of the co-culture of empty vector lymphocytes and KATO Ⅲ gastric cancer cells （F=5.010, P〈0.01） or the co-culture of CEA special lymphocytes and BGC-823 gastric cancer cells （F=4.982, P〈0.01）, which had no significant difference between the co-culture of CEA special lymphocytes and BGC- 823 gastric cancer cells and the co-culture of empty vector lymphocytes and BGC-823 gastric cancer cells （F= 1.210, P〉0.05）. Conclusion CEA special lymphocytes can promote cell apoptosis and inhabit tumor reproduction of CEA positive gastric cancer cells in vitro and in vivo.