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为逆转多药耐药ｍｄｒ－１基因产物Ｐ－ｇｐ蛋白所介导的肿瘤细胞对多种化疗药物的耐受性，作者设计合成了一种能切割ｍｄｒ－１ｍＲＮＡ第１９６密码子ＧＵＣ序列的锤头状核酶（Ｒｉｂｏｚｙｍｅ），并定向克隆于逆转录病毒载体。经ＰＡ３１７包装后，病毒上清感染ＢＥＬ－７４０２／ＤＯＸ细胞株。经Ｎｏｒｔｈｅｒｎｂｌｏｔ杂交证实，ＰＡ３１７及转化的ＢＥＬ－７４０２／ＤＯＸ细胞中均有病毒的高表达，ＲＴ－ＰＣＲ结果表明，转化细胞中ｍｄｒ－１ｍＲＮＡ与未转化细胞相比明显减少甚至不能扩增出来，流式细胞技术检测发现转化细胞表面Ｐ－ｇｐ的表达与非转化细胞相比明显下降。ＭＴＴ法检测发现转化细胞对多种化疗药物重新产生较高的敏感性。结果提示，此Ｒｉｂｏｚｙｍｅ转化ＢＥＬ－７４０２／ＤＯＸ细胞后能有效抑制ｍｄｒ－１基因的表达，使已产生耐药的肝癌细胞的多药耐药表型发生逆转。
In order to reverse the tolerance of tumor cells to various chemotherapeutic drugs mediated by multidrug resistance mdr-1 gene product P-gp protein, the authors designed and synthesized a hammer capable of cleaving the mdr-1 mRNA codon GUC sequence. Ribozymes are directional and cloned in retroviral vectors. After packaging with PA317, the virus supernatant was infected with the BEL-7402/DOX cell line. Northern blot hybridization confirmed that the virus was highly expressed in both PA317 and transformed BEL-7402/DOX cells. RT-PCR results showed that the mdr-1 mRNA was significantly reduced or even not amplified in the transformed cells. The expression of P-gp on the surface of transformed cells was significantly lower than that of non-transformed cells. The MTT assay showed that the transformed cells regenerated higher sensitivity to various chemotherapeutic drugs. The results suggest that this ribozyme-converted BEL-7402/DOX cell can effectively inhibit the expression of mdr-1 gene and reverse the multidrug resistance phenotype of the drug-resistant hepatocellular carcinoma cells.