腺病毒介导人IL-2基因治疗的抗肿瘤作用及机理分析

Antitumor Effect of Gene Therapy with Recombinant Adenovirus-mediated Human Interleukin-2 Gene Transfer and Its Mechanism

目的 观察重组人白介素 2腺病毒 (advh IL- 2 )在小鼠肝癌 H2 2中的转染效率、表达时程、体内抗肿瘤作用及其免疫机理。方法 小鼠皮下接种肝癌 H2 2 ,将肿瘤生长至体积为 10 0～ 2 0 0 mm3的荷瘤小鼠随机分组 ,经肿瘤局部注射给药。用 X- gal染色法检测重组腺病毒载体 adv L ac Z的转染效率 ;用 RT- PCR检测人 IL- 2基因的持续表达时间 ;以 H2 2瘤体生长及荷瘤小鼠存活时间评价 advh IL - 2抗肿瘤作用 ;以 51 Cr 4小时释放法测定治疗小鼠脾细胞的 L AK与 CTL 杀伤活性 ,免疫荧光法分析肿瘤组织内 CD4 +与 CD8+ T细胞浸润情况。结果 肿瘤局部单次注射 1× 10 9pfu重组腺病毒 ,可在肿瘤组织中进行有效的转染及表达 ,人 IL- 2基因的表达时间长于 12 d。AdvhIL- 2的抗肿瘤作用具有剂量依赖性 ,与 PBS组比较 ,2× 10 9pfu advh IL- 2可非常明显地抑制 H2 2皮下瘤的生长、延长荷瘤小鼠的生存时间 (P<0 .0 1) ,同时明显增强脾细胞 L AK与 CTL杀伤活性 (P<0 .0 1) ,增加肿瘤组织内CD4 + 与 CD8+ T细胞的浸润。结论 Advh IL - 2通过介导人 IL - 2基因在小鼠 H 2 2肿瘤组织中的持续表达 ,产生明显的体内抗肿瘤作用 。

Objective To explore the transfection, expression, antitumor effect and mechanism of a replication deficient adenovirus vector expressing human IL 2 gene(advhIL 2) in a murine H22 hepatocellular carcinoma model. Methods KM mice bearing tumor 100 200 mm 3 were injected intratumorally with advhIL 2, adv LacZ or with PBS alone. The tumor evolution was recorded every 3 days. The transfection and expression of the recombinant adenovirus were examined with X gal staining and RT PCR respectively. The splenic LAK and CTL activities were assayed with 51 Cr 4 hours release. The infiltration of CD 4 + and CD 8 + T cells in the tumors were analyzed with immunofluorescence. Results It was found that the recombinant adenovirus can bring about in vivo effective transfection and expression. The continuosly expressive time of advhIL 2 is longer than 12 days. AdvhIL 2 has dose dependent anti tumor effect. The mice received a dose of 2×10 9 pfu advhIL 2 divided into halves for two injections developed tumor more slowly and survived much longer than the mice treated with PBS ( P <0.01). Moreover, advhIL 2 increased the splenic LAK and CTL activities, CD 4 + and CD 8 + T cell infiltration in the H22 tumors significantly. Conclusion Adenovirus mediated hIL 2 gene treatment has significant antitumor activity in pre established hepatocellular carcinoma in mice. Antitumor immunity might be responsible for the therapeutic effect.