Qu Y, OuYang X, Liu S, Peng X, Liu B
Laboratory of Medical Molecular Biology, West China University of Medical Sciences, Chengdu, 610041 P. R. China.
Zhonghua Yi Xue Yi Chuan Xue Za Zhi. 1999 Jun;16(3):133-7.
To evaluate the possibility of using ribozyme technology for telomerase inhibition and cancer therapy.
A hammer head ribozyme (telomerase ribozyme, teloRZ) directed against the RNA component of human telomerase (hTR) was designed and synthesized to serve as a telomerase inhibitor. An in vitro transcription plasmid and a eukaryotic expression plasmid containing teloRZ gene were constructed. In vitro cleavage reaction was carried out by mixing the ribozyme RNA with DIG-labeled-hTR in different reaction conditions. Cleavage bands were detected by digoxin chemilumines- cent assay. The eukaryotic expression plasmid was inducted into HeLa cells by lipofectamine; the telomerase activities and bio-characteristics of HeLa cells were detected continuously.
teloRZ showed a specific cleavage activity against the telomerase RNA component used as template. The in vitro cleavage ratio reached about 60%. The telomerase activities of cells expressing teloRZ dropped to eight times; the doubling times became longer and apoptosis ratios became higher with increasing population doublings (PDS); at 19-20 PDS 95% cells showed apoptosis.
These findings support the potential use of this ribozyme against immortalized cancer cells.
评估利用核酶技术抑制端粒酶及用于癌症治疗的可能性。
设计并合成一种针对人端粒酶RNA组分（hTR）的锤头状核酶（端粒酶核酶，teloRZ）作为端粒酶抑制剂。构建含teloRZ基因的体外转录质粒和真核表达质粒。将核酶RNA与地高辛标记的hTR在不同反应条件下混合进行体外切割反应。用地高辛化学发光法检测切割条带。通过脂质体转染将真核表达质粒导入HeLa细胞；连续检测HeLa细胞的端粒酶活性及生物学特性。
teloRZ对用作模板的端粒酶RNA组分表现出特异性切割活性。体外切割率达约60%。表达teloRZ的细胞端粒酶活性降至八分之一；随着群体倍增数（PDS）增加，倍增时间延长，凋亡率升高；在19 - 20个PDS时，95%的细胞发生凋亡。
这些结果支持这种核酶在针对永生化癌细胞方面的潜在应用。
