Control of Gene Expression by Functional and Artificial Nucleic Acids

为了开发一种控制基因表达的新策略，合成了一种寡核苷酸功能分子偶联物，并对其生物学特性进行了研究。结果总结如下。1)设计合成了一些杂环化合物，并将其整合到桥式核酸骨架中。将获得的带有非天然核碱基的桥接核酸引入寡核苷酸中，并对其三联体形成能力进行了研究。非常有趣的是，我们发现桥接的核酸衍生物成功地实现了嘧啶-嘌呤中断位点的识别/2)为了方便地将官能团引入到聚核苷酸中，我们开发了基于Click化学的后延伸修饰方法。此外，还成功地制备了一些与转录因子结合位点（双链DNA）结合的寡核苷酸。3)将上述制备的含有非天然农村核碱基的寡核苷酸转染到活细胞中，并评估其控制基因表达的能力。含有非天然核碱基的寡核苷酸，无论是单独使用还是与本小组开发的抗原阻断策略一起使用，都能有效地调控基因表达。此外，与转录因子结合位点结合的寡核苷酸促进了靶基因的表达。这些结果是非常有趣的，将有助于进一步的生物学应用。4)详细评价了寡核苷酸类似物的化学结构与其三联体形成能力之间的关系，成功开发出比pinto型BNA具有更高三联体形成能力的新型桥接核酸。

In order to develop a novel strategy to control a gene expression, an olignnucleotidc-functional molecule conjugate has been synthesized and its biological properties were also studied. The results are summarized below.1) Some heterocyclic compounds were designed, synthesized and incorporated into a bridged nucleic acid skeleton. The obtained bridged nucleic acids bearing unnatural nucleobases were also introduced into oligonucleotides, and their triplex-forming ability was investigated. Very interestingly, it was found that the recognition of pyrimicline-purine interruption site was successfully achieved by the bridged nucleic acid derivatives /2) As a convenient method to introduce a functional group into oligonucleotides, a post-elongation modification method based on Click chemistry was developed. Furthermore, some oligonucleotides conjugated with a transcription factor-binding site (double stranded DNA) were also successfully prepared.3) The oligonucleotides bearing unnatrural nucleobases prepared above were transfected into living cells, and their ability to control gene expression was evaluated. The oligonucleotides bearing unnatural nucleobases, by themselves or together with the antigene-block strategy developed by our group, showed effective regulation of gene expression. Furthermore, the oligonucleotides conjugated with a transcription factor-binding site promoted the target gene expression. These results are of great interest and would be useful for further biological application.4) A relationship between chemical structures of oligonucleotide analogues and their triplex-forming ability was evaluated in detail, and the novel bridged nucleic acid, of which triplex-forming ability was higher than that of the pinto-type BNA, was successfully developed.