Establishment of novel molecular cancer therapy targeting telomerase and its clinical appication to gynecologic tumors

端粒酶激活在肿瘤的生长和发展中起着至关重要的作用，部分通过维持端粒结构。事实上，端粒酶在人类癌症中的普遍表达使端粒酶成为癌症治疗的一个有希望的靶点。描述了抑制端粒酶的遗传、药理学和反义方法；然而，在大多数情况下，癌细胞只有在多次细胞分裂后才会死亡。在这里，我们使用逆转录病毒递送人类端粒酶逆转录酶（hTERT）特异性的小干扰RHAs，成功地抑制了宫颈癌细胞系的端粒酶活性。缺乏hTERT表达的细胞端粒酶活性显著降低，端粒缩短，端粒3'-悬垂。这些细胞经过相当数量的细胞分裂后进入复制衰老。值得注意的是，与端粒酶活性的对照细胞相比，这些细胞的增殖率明显受损，即使在低传代细胞中也是如此（PD 5）。同样，在缺乏hTERT的低传代细胞中，小鼠的集落形成能力和致瘤性都减弱了。我们进一步研究了化疗和电离辐射对hTERT表达被抑制的细胞的影响。缺乏hTERT的细胞对诱导DNA双链断裂的电离辐射或化疗药物（如拓扑异构酶抑制剂或博来霉素）的敏感性明显高于对照细胞。这些发现表明，基于sirna的策略可以应用于新型端粒酶抑制剂的开发，其抗肿瘤作用可能与电离辐射和化疗联合使用。

Telomerase activation plays critical roles in tumor growth and progression in part through the maintenance of telomere structure. Indeed, the ubiquitous expression of telomerase in human cancers makes telomerase a promising target for cancer therapy. Genetic, pharmacologic and antisense methods to inhibit telomerase have been described ; however, in most cases, cancer cell death was observed only after many cell divisions. Here, using retroviral delivery of small interfering RHAs specific for the human telomerase reverse transcriptase (hTERT), we successfully inhibited telomerase activity in cervical cancer cell lines. Cells lacking hTERT expression exhibited significantly decreased telomerase activity and showed shortened telomeres and telomeric 3'-overhangs with passage. These cells entered the replicative senescence after considerable number of cell divisions. Notably, the proliferative rate of these cells was significantly impaired, compared to control cells with telomerase activity, even in low passage cells (PD 5). Likewise, colony-forming ability and tumorgenicity in mice were attenuated in low passage cells lacking hTERT. We further examined the effects of chemotherapy and ionizing radiation of cells in which hTERT expression is suppressed. Cells lacking hTERT showed a significantly increased sensitivity than control cells to ionizing radiation or chemotherapeutic agents that induce DNA double strand breaks, such as topoisomerase inhibitors or bleomycin. These findings suggest that a siRNA-based strategy can be applied to the development of novel telomerase inhibitors, whose anti-tumor effects may be enhanced in combination with ionizing radiation and chemotherapy.