Establishment of novel gene therapy targeting gynecologic tumors

我们建立了一种新的方法，利用2-5腺苷酸连接的反义DNA抑制人类端粒酶RNA成分（hTR）在癌细胞中抑制端粒酶活性。该反义DNA可有效阻断宫颈癌ME180细胞的端粒酶活性。2-5A抗htr能显著抑制ME180细胞的体外生长。令人惊讶的是，在治疗后24-48小时观察到细胞生长抑制，比预期的要早得多。端粒长度在短时间内没有缩短。这些发现表明，端粒酶的阻断通过不依赖端粒的机制导致细胞生长抑制。我们证实2-5A抗htr对细胞生长的抑制是由于诱导细胞凋亡。2-5A抗htr诱导细胞凋亡的机制正在进一步分析中。建立了肿瘤基因治疗的新型载体体系。我们之前克隆了人类端粒酶逆转录酶（hTERT）的启动子，它对癌细胞具有高度特异性。因此，我们利用该启动子在基因传递系统中表达癌症特异性基因。将caspase-8、FADD等多种凋亡诱导基因与hTERT启动子结合，作为肿瘤基因治疗的载体。这些载体的引入有效地诱导了癌细胞的凋亡，但周围的正常组织。

We established novel methods that inhibit telomerase activity in cancer cells using 2-5Adenylate-linked antisense DNA against human telomerase RNA component (hTR). This antisense DNA effectively blocked telomerase activity in cervical cancer ME180 cells. Growth of ME180 cells in vitro was significantly inhibited by the treatment with 2-5A anti-hTR. Surprisingly, inhibition of cell growth was observed in 24-48 hr after treatment, quite earlier than expected. Telomere length was not shortened in this short period. These findings suggest that blockade of telomerase led to cell growth inhibition via telomere-independent mechanisms. We confirmed that growth inhibition of cells by the treatment with 2-5A anti-hTR was due to induction of apoptosis. The further anaylsis of mechanisms how 2-5A anti-hTR induces apoptosis is on going.We also established novel vector system for cancer gene therapy. We previously cloned promoter of human telomerase reverse transcriptase (hTERT), which is highly specific to cancer cells. We thus used this promoter for cancer-specific gene expression in gene delivery system. Various apoptosis-inducible genes, such as caspase-8 and FADD, were combined with hTERT promoter and used as vectors for cancer gene therapy. Introduction of these vectors effectively induced apoptosis of cancer cells but of surrounding normal tissues.