He Y K, Lui V W, Baar J, Wang L, Shurin M, Almonte C, Watkins S C, Huang L
Department of Pharmacology, University of Pittsburgh, School of Medicine, PA 15261, USA.
Gene Ther. 1998 Nov;5(11):1462-71. doi: 10.1038/sj.gt.3300769.
Down-regulation of oncogene expression by antisense-based gene therapy has been extensively studied, and in some cases, therapeutic effects have been demonstrated. We have previously shown that down-regulation of HPV16 E6 and E7 gene expression inhibited HPV DNA-positive C3 mouse tumor growth. Although not all of the tumor cells were transfected by pU6E7AS plasmid, complete tumor regression was achieved if the tumor size was small at the start of therapy in a syngeneic host. This suggests that some other antitumor mechanisms may be involved in addition to the direct down-regulation of HPV16 E7 oncogene expression by the antisense effect of E7AS. In the current study, we demonstrated that E7AS induces tumor cell apoptosis. More importantly, a strong antitumor immune response was elicited in the pU6E7AS-treated and tumor-regressed mice. There was no tumor growth after rechallenging the tumor-regressed mice with 1 million C3 cells. This E7AS-induced antitumor immune response was augmented by co-delivery of mIL-12 cytokine gene. The combination therapy strategy resulted in complete regression of 26 of 28 (93%) tumors. Only 12 of 31 (38%) tumors from the group treated with pU6E7AS alone and 14 of 28 (50%) tumors from the group treated with pCMVmIL-12 alone had completely regressed. Complete regression was also demonstrated in tumors located 1 cm from the treated tumors, which indicates that a systemic antitumor effect was induced by E7AS and mIL-12. Immunohistochemistry demonstrated that a significant amount of CD4+ and CD8+ cells infiltrated into tumors treated with pU6E7AS, pCMVmIL-12 and pU6E7AS+pCMVmIL-12. These data indicate that host immunity is an important factor for antisense-based gene therapy approach which can be further enhanced by combination with cytokine gene therapy.
基于反义技术的基因疗法对癌基因表达的下调作用已得到广泛研究，在某些情况下，已证实具有治疗效果。我们之前已经表明，HPV16 E6和E7基因表达的下调抑制了HPV DNA阳性的C3小鼠肿瘤生长。尽管并非所有肿瘤细胞都被pU6E7AS质粒转染，但在同基因宿主中，如果在治疗开始时肿瘤体积较小，仍可实现肿瘤完全消退。这表明，除了E7AS的反义效应直接下调HPV16 E7癌基因表达外，可能还涉及其他一些抗肿瘤机制。在当前研究中，我们证明E7AS可诱导肿瘤细胞凋亡。更重要的是，在用pU6E7AS治疗且肿瘤消退的小鼠中引发了强烈的抗肿瘤免疫反应。用100万个C3细胞再次攻击肿瘤消退的小鼠后，未出现肿瘤生长。通过共递送mIL-12细胞因子基因可增强这种E7AS诱导的抗肿瘤免疫反应。联合治疗策略使28个肿瘤中的26个（93%）完全消退。单独用pU6E7AS治疗的组中，31个肿瘤只有12个（38%）完全消退，单独用pCMVmIL-12治疗的组中，28个肿瘤只有14个（50%）完全消退。在距离治疗肿瘤1厘米处的肿瘤中也证实了完全消退，这表明E7AS和mIL-12诱导了全身抗肿瘤效应。免疫组织化学显示，大量CD4+和CD8+细胞浸润到用pU6E7AS、pCMVmIL-12和pU6E7AS + pCMVmIL-12治疗的肿瘤中。这些数据表明，宿主免疫是基于反义技术的基因治疗方法的一个重要因素，与细胞因子基因治疗联合可进一步增强这种作用。
