MODIFIELD OLIGONUCLEOTIDES IN TRIPLE HELIX FORMATION

这项研究的主要目标是从化学上提炼一类序列-- 特异性、DNA结合、三螺旋形成的寡核苷酸(TFOS)。这个 这些TFOS独特特征是将第三链结合到其 双链DNA靶标与pH无关，由GGC和TAT稳定 三元组，这样的首选取向将绑定的TFO 相对于更多富含嘌呤的潜在链的反平行 双面打印。我们对38个碱基长的寡核苷酸(HIV38P)的初步研究， 设计成与长末端重复序列中的启动子区域结合 人类免疫缺陷病毒(HIV)的序列分析表明，HIV38p具有 由于高G含量而有自我联想的倾向。后续研究 用HIV38P研究表明，G被2‘-脱氧-7-取代。 去氮鸟苷残基显著降低了自结合作用。因此， HIV38P中部分或全部鸟嘌呤残基的部分或全部替换 鸟嘌呤类似物可能会减少或消除自联想并改善 三联体队形。 我们现在建议制备含有不同水平取代基的TFOS 含2‘-脱氧-7-脱氮鸟苷或2’-脱氧-6- 采用固相亚磷酰胺化学的硫代鸟苷。这个 这样得到的修饰寡核苷酸将得到提纯和充分 特色化的。这些新型TFOS形成稳定三链能力 将使用频带偏移分析、DNase足迹进行评估 实验、熔化曲线和Cd测量。希望这样的一个 取代改善了三螺旋结构，减少了自缔合， 提高结合亲和力和抗病毒(艾滋病毒和单纯疱疹病毒)活性。详细 将在第二阶段评估这种经修饰的TFOS的生物效应 学习。

The primary goal of this study is to chemically refine a class of sequence- specific, DNA binding, triple helix-forming oligonucleotides (TFOs). the unique feature of these TFOs is that binding of the third strand to its duplex DNA target is pH independent and is stabilized by GGC and TAT triplets such that the preferred orientation places the bound TFO antiparallel with respect to more purine rich strand of the underlying duplex. Our initial studies with a 38 base long oligonucleotide (HIV38p), designed to bind to the promoter region in the long terminal repeat sequence of human immunodeficiency virus (HIV), indicated that HIV38p has a tendency to self-associate due to high G content. Subsequent studies with HIV38p have shown that substitution of G with 2'-deoxy-7- deazaguanosine residues reduces the self-association considerably. Thus, substitution of some or all guanine residues in HIV38p with selected guanine analogs may reduce or eliminate self-association and improve triplex formation. We now propose to prepare TFOs containing various levels of substitutions ranging from 10 to 50% with 2'-deoxy-7-deazaguanosine or 2'-deoxy-6- thioguanosine employing solid-phase phosphoramidite chemistry. The modified oligonucleotides thus obtained will be purified and adequately characterized. The ability of these novel TFOs to form stable triplexes will be evaluated using band shift analysis, DNase footprinting experiments, melting curves and CD measurements. It is hoped that such a substitution improves triple helix structure, reduces self-association, increases binding affinity and antiviral (HIV and HSV) activity. Detailed biological effects of such modified TFOs will be assessed in Phase II studies.