Recognition of Chromosomal DNA by Peptide Nucleic Acids

描述(申请人提供)：肽核酸(PNA)是一种通过Watson-Crick碱基配对识别互补序列的DNA模拟物。PNA的优势之一是它们能够通过链入侵识别双链DNA中的位点。我们已经开发了合成PNA并将其引入细胞的有效方法。我们还描述了提高链入侵效率的策略。我们现在建议结合这些进展来开发反基因PNA，作为在染色体水平上操纵基因表达的试剂。 这项建议的目的是了解反基因PNA的性质，并验证反基因PNA可以识别细胞内染色体DNA的假设。具体地说，我们建议1)表征和优化PNA的细胞内定位，2)制定使用反义PNA操纵基因表达的规则，以及3)探索反义PNA作为识别双链DNA的一般策略的价值，通过靶向在癌症进展中至关重要的基因，特别是c-myc和端粒酶hter T的逆转录酶成分。 为了实现这些目标，我的实验室将利用我们快速合成PNA和PNA-肽结合物的能力，以及我们在细胞内使用PNA的经验。初步实验已经表明，PNA可以作为抗基因试剂，为本提案中描述的实验提供了一个强有力的起点。 序列特异性识别染色体DNA的化合物具有巨大的治疗疾病的潜力，并成为强大的研究工具。这类药物可能会阻止基因表达，激活基因表达，或者使有害的突变得以纠正。尽管染色体DNA作为靶标很有吸引力，但抗基因药物的发展一直很缓慢。这项建议中描述的研究严格测试了抗基因PNA识别染色体的价值，我们的数据将决定使用PNA进行实验室研究和临床开发的决定。

DESCRIPTION (provided by applicant): Peptide nucleic acid (PNA) is a DNA mimic that recognizes complementary sequences by Watson-Crick base-pairing. One of the strengths of PNAs is their ability to recognize sites within duplex DNA by strand invasion. We have developed efficient methods for synthesizing PNAs and introducing them into cells. We have also characterized strategies for improving the efficiency of strand invasion. We now propose to combine these advances to develop antigene PNAs as agents for manipulating gene expression at the level of the chromosome. The objective of this proposal is to understand the properties of antigene PNAs and test the hypothesis that antigene PNAs can recognize chromosomal DNA inside cells. Specifically, we propose to 1) Characterize and optimize the intracellular localization of PNAs, 2) Develop rules for using antisense PNAs to manipulate gene expression, and 3) Explore the value of antigene PNAs as a general strategy for recognizing duplex DNA by targeting genes that are important in the progression of cancer, specifically c-myc and the reverse transcriptase component of telomerase h TER T. To achieve these goals my laboratory will take advantage of our ability to rapidly synthesize PNAs and PNA-peptide conjugates and our experience with using PNAs inside cells. Preliminary experiments have already shown that PNAs can act as antigene agents, providing a powerful starting point for the experiments described in this proposal. Compounds that sequence-specifically recognize chromosomal DNA have enormous potential to treat disease and be powerful research tools. Such agents might block gene expression, activate gene expression, or enable harmful mutations to be corrected. In spite of the attractiveness of chromosomal DNA as a target, the development of antigene agents has been slow. The studies described in this proposal rigorously test the value of antigene PNAs for the recognition of chromosomes and our data will shape decisions regarding the use of PNAs for laboratory studies and clinical development.