MODIFIED NUCLEOSIDES IN TRIPLEX FORMING OLIGONUCLEOTIDES

描述（改编自申请人的摘要）：本研究的主要目标 这项研究是从化学上提炼一类序列特异性DNA结合， 三螺旋形成寡核苷酸（TFO）。 这些独特的功能 TFOs是第三条链与其双链体DNA靶的结合是pH 是独立的，并通过GGC和达特三联体稳定， 优选的取向将结合的TFO相对于 底层双链体的嘌呤更丰富的链。 符合雅阁 分子模拟，将2 '-脱氧甲氧霉素（DF）引入到3'-胺中， 在进行氢键合的位点处的改性TFO， 双链体靶中的多嘌呤/多嘧啶运行被CG中断 反演 这种dF-TFO的结合亲和力通过条带来评估。 移位分析和定量DNA酶I足迹技术。 的 通过局部高亲和力评估dF对三链体结构的影响 铜菲咯啉裂解。 在初步研究中，3 'A-PRE 2ap的同源物显示出10倍的 由于在三个位点包含dF， 与其未修改的对应物相比。 在I期研究中，我们将 证实了这些发现，并扩展了在TFO中使用dF取代 其对表皮生长因子受体（EGFR）（EG 36 ap）具有特异性， 小鼠胰岛素受体（IR）启动子（IR 2ap）和HIV-1（HIV 38 p）结构域。 如果dF取代改善了三螺旋结构和结合亲和力， 在II期研究中，将评估此类TFO的生物效应。

DESCRIPTION (Adapted from applicant's abstract):The primary goal of this study is to chemically refine 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 the more purine rich strand of the underlying duplex. In accord with molecular modeling, 2'-deoxyformycin (df) is introduced into 3'-amine modified TFOs at sites engaged in H-bonding where polypurine/polypyrimidine runs in a duplex target are interrupted by a CG inversion. The binding affinity of such a dF-TFO is assessed by band shift analysis and by quantitative DNase I footprinting techniques. The effect of dF upon triplex structure is assessed by local high affinity Cuphenanthroline cleavage. In preliminary studies, a homologue of 3'A-PRE2ap displayed a 10-fold increase in binding affinity due to inclusion of dF at three sites as compared to its unmodified counterpart. In Phase I studies we will confirm these findings and extend the use of dF substitution within TFOs which are specific for epidermal growth factor receptor (EGFR) (EG36ap), mouse insulin receptor (IR) promoter (IR2ap) and HIV-1 (HIV38p) domains. If dF substitution improves triple helix structure and binding affinity, in Phase II Studies the biological effects of such TFOs will be assessed.