OLIGONUCLEOTIDE INHIBITION OF ONCOGENE EXPRESSION

这项工作的目的是阐明 反义寡核苷酸抑制细胞增殖 由人类c-myc和原认知字符转化而成。 人类cMYC编码一种保守的核蛋白p65，它 可能参与基因表达的调控，c-Ha-ras 编码一种保守的内质膜蛋白p21，它 可能参与了生长因子信号的传导。细胞 变形可能是由不恰当的表达或 癌基因产物在细胞增殖过程中的反应，从而 降低cMYC和CHA-ras的表达水平可能在 原则上足以进行反向转换。计算出 CMYC和charas mRNAs的二级结构预测 在帽子和起始密码子之间有许多环状和凸起， 易受反义寡核苷酸杂交停滞影响。 在HL60细胞中加入抗cMYC的十五核苷酸 培养抑制细胞增殖并刺激细胞分化 剂量依赖和序列特异性的方式和一种抗癌 HARAS十五核苷酸抑制人卵巢癌细胞增殖 T24转化NIH3T3细胞。在这个项目中，剂量 反义寡核苷酸对cMYC的依赖作用 和cha-ras mRNA转录、稳定性、翻译、DNA 复制，细胞形态将在正常和 转化的细胞。寡核苷酸摄取， 分隔和退化将在同一课程中进行研究 细胞系。然而，寡核苷酸还不够稳定。 用于静脉或口服给药。低聚脱氧核苷 甲基膦酸盐不带电荷，对核酸酶有抵抗力，进入 动物细胞并保护它们免受病毒的挑战，但 在体外的效果出乎意料地低于正常 寡聚脱氧核苷酸。R和S立体异构体分别出现在 磷原子，但只有S形式具有相同的构象 作为正常的寡聚脱氧核苷酸。立体专门化ALL 将合成低聚脱氧核苷甲基膦酸酯，并 测试它们在阻止杂交方面的有效性。

The goal of this work is to elucidate the mechanism by which antisense oligodeoxynucleotides inhibit proliferation of cells transformed by human c-myc and cHaras proto-oncogneses. Human cmyc codes for a conserved nuclear protein, p65, which may be involved in regulation of gene expression, and c-Ha-ras codes for a conserved inner plasma membrane protein, p21, which may be involved in transducing growth factor signals. Cell transformation may be caused by inappropriate expression or response of oncogene products during cell proliferation, so that reducing the level of cmyc and cHa-ras expression may in principle be sufficient to reverse transformation. Calculated secondary structures of cmyc and cHaras mRNAs predict many loops and bulges between the cap and the initiation codon, susceptible to antisense oligodeoynucleotide hybridization arrest. Addition of an anti-cmyc pentadecanucleotide to HL60 cells in culture inhibits proliferation and stimulates differentiation in a dose dependent and sequence specific manner and an antic- Haras pentadecanucleotide inhibits proliferation of T24transformed NIH 3T3 cells. In this project, the dose dependent effects of antisense oligodeoxynucleotides on cmyc and cHa-ras mRNA transcription, stability, translation, DNA replication, and cell morphology will be determined in normal and transformed cells. Oligodeoxynucleotide uptake, compartmentalization and degradation will be studied in the same cell lines. Oligodeoxynucleotides, however, are not stable enough for intravenous or oral administration. Oligodeoxynucleoside methylphosphonates are uncharged and nuclease resistant, enter animal cells and protect them from viral challenge, but are unexpectedly less effective in vitro than normal oligodeoxynucleotides. R and S stereoisomers occur at each phosphorus atom, but only the S form has the same conformation as a normal oligodeoxyonucleotide. Stereospecific allS oligodeoxynucleoside methylphosphonates will be synthesized and tested for their efficacy in hybridization arrest.