摘要
设计合成能切割人ｍｄｒ－１ｍＲＮＡ的Ｒｉｂｏｚｙｍｅ基因并构建成真核细胞表达克隆（ｐＨβＡｐｒ－１ｎｅｗ／５ｍＲ３），以靶向切割耐药癌细胞株（ＭＣＦ－７／Ａｄｒ）的ｍｄｒ－１ｍＲＮＡ，抑制Ｐ－糖蛋白的表达，逆转多药抗药性。方法将含Ｒｉｂｏｚｙｍｅ基因的真核细胞表达克隆转染ＭＣＦ－７／Ａｄｒ细胞，采用斑点杂交和Ｎｏｒｔｈｅｒｎ杂交技术测得Ｒｉｂｏｚｙｍｅ能在细胞中稳定表达，并使细胞中ｍｄｒ－１ｍＲＮＡ降解８３．５％，Ｐ－糖蛋白表达明显下降（ＡＢＣ法）。结果Ｒｉｂｏｚｙｍｅ基因转染的ＭＣＦ－７／Ａｄｒ细胞耐药性下降至敏感细胞的６倍，而无Ｒｉｂｏｚｙｍｅ基因转染的ＭＣＦ－７／Ａｄｒ耐药性是敏感细胞的１０００倍。结论这种基因调控技术介入可能将为肿瘤耐药的治疗开辟一个新领域。
Objective To construct a specific hammerhead ribozyme possessing catalytic activity that cleaves the mdr1 mRNA for reversing the resistant phenotype in tumour cell line. Methods A DNA sequence encoding the ribozyme gene was incorporated into a eukaryotic expression vector(pHβ Apr 1 neu) and transfected into the human breast carcinoma cell line MCF 7/Adr, which is resistant to adriamycin and expresses the MDR phenotype. Results The ribozyme was stably expressed in the cell line, and decreased the level of mdr1 mRNA expression by 83.5%. The ribozyme inhibited the formation of P glycoprotein and reduced the cell's resistance to adriamycin. Conclusion The resistant cells were 1,000 fold more resistant than the parental cell line(MCF 7), whereas those cell clones that showed ribozyme expression were only 6 fold more resistant than the parental cell line.
出处
《中华医学杂志》
CAS
CSCD
北大核心
1997年第7期494-496,共3页
National Medical Journal of China
基金
国家自然科学基金
