慢性粒细胞白血病bcr-abl基因反义核酸的初步研究

Antisense Studies toward her-abl Gene in Chronic Myelogenous Leukemia

探讨用逆转录病毒表达载体携带反义核酸进入 ＣＭＬ细胞并作用于 ｂｃｒ－ａｂ１基因的方法。方法：反义序列用ＲＴ－ＰＣＲ法，取自Ｋ５６２细胞系，使之成功地构建了正反义ＲＮＡ的逆转录病毒表达载体。结果：与正义链相比，反义ｂｃｒ－ａｂ１序列显著抑制Ｋ５６２及ＣＭＬ细胞生长，其抑制程度与剂量呈正相关，其细胞形态学改变支持其进入凋亡进程。结论：此反义核酸是探索ＣＭＬ基因治疗的重要材料。

Background and Purpose:Antisense strategy is becoming an important way in the regulation and control of many dis- eases. Chronic myelogenous leukemia (CML) is a mlyeloproliferative disorder in which the abnormal hematopoitic stem cells contain bcr-abl fusion gene, which is the pathogen in CML. As the results of gene therapy such as antisense oligde- oxynuleotide, rybozym and P210 inhibitors, this gene is the most important target in the gene therapeutic studies. Methods: The retrovirus-expressing vector was used to bring an antisense sequence to the abnormal cells. The antisense sequence was isolated from K562 cells by RT-PCR. This 0. 3kb sequence targets the junction region of bcr-abl mRNA, which contains 73 nucleotides of bcr exon 2,75 nucleotides of bcr exon 3 and 161 nucleotides of abl exon 2. Results: In comparison with the sense chain, it was found that the antisense chain inhibited the growth of K562 and CML cells. After treatment, the prolifer- ation of the K652 cells showed remarkable inhibitionl with antisense dose-related response. After exposing to the antisense for 72 hours,more than 50% of these cells were not resistant to trepan blue with typical morphological changes indicating apop- tosis process. Conclusion: This antisense fragment may be useful for CML gene therapy studies.