Basic Study for The Gene-Targeted Therapy in Lung cancer

我们研究了L-myc反义DNA对高表达L-myc的人小细胞肺癌细胞株NCI-H209的抗增殖作用。该细胞系经反义DNA处理后，其翻译起始位点L-myc的表达量明显增加，主要定位于细胞核，细胞质中的表达量较低，myc蛋白在1.6 μ M~10 μ M浓度范围内呈浓度依赖性抑制细胞增殖，在三种不同的SCLC细胞系中，包括NCI-L-myc，该反义DNA的生长抑制与L-myc的表达水平相关。H209.与反义OPT核苷酸长度相同但顺序随机的混合序列对照OPT对细胞增殖无影响，正义OPT处理引起细胞增殖的一定抑制，这种抑制可能是OPT的非序列特异性抑制，不依赖于常规的反义基因抑制，在反义处理的NCI-H209细胞中观察到L-myc mRNA的大量表达和L-myc蛋白的表达降低，表明覆盖翻译起始位点的反义OPT通过抑制靶mRNA的核糖体翻译而不是抑制基因的转录来起作用。包括1）在SCLC中频繁扩增和过表达的状态，和2）在人成人组织中非常有限和低水平表达，目前的数据表明L-myc是基于分子生物学诊断的SCLC反义DNA治疗的靶基因的良好候选者。

We evaluated the antiproliferative effect of L-myc antisense DNA in NCI-H209, a human small cell lung cancer (SCLC) cell line overexpressing the L-myc gene.The synthetic DNA used in the present study was 15mer oligodeoxynucleoside phosphorothioate (OPT), which showed rapid incorporation into NCI-H209 cells, and which localized mainly in the cell nucleus and weakly in the cytoplasm.The exposure of this cell line to s L-myc antisense DNA convering the translational initiation site L-myc proteins inhibited the cell proliferation in a concentration-dependent manner through the concentrations of 1.6muM to 10muM.Furthermore, the growth inhibition by this antisense DNA was correlated with the level of L-myc expression in three different SCLC cell lines including NCI-H209.A mixed sequence control OPT having the same nucleotide length as the antisense OPT but in random, order, did not show any effect on cell proliferation.The sense OPT treatment induced the some inhibition of cell proliferation, which migh be non-sequence-specific inhibition by OPT independent of conventional antisense gene inhibition, which was reported in other systems.Abundant expression of L-myc mRNA and decreased expression of L-myc protein were observed in the antisense-treated NCI-H209 cells, suggesting that the antisense OPT covering the translational initiation site acted by inhibiting ribosomal translation of the target mRNA,rather than by inhibiting transcription from the gene.Together with unique characteristics of the L-myc gene, including 1) a frequently amplified and overexpressed state in SCLC,and 2) very restricted and low level expression in human adult tissues, the present data indicate that L-myc is a good candidate for the target gene for antisense DNA therapy in SCLC based on molecular biological diagonosis.