Development of Cell-Permeable Peptide Nucleic Acid

描述（由申请人提供）：最近，我们已经开发了一类新的肽核酸，称为GPNA，其容易被人类体细胞和胚胎干细胞摄取，并且序列特异性地结合DNA和RNA。与未修饰的PNA及其其他衍生物相比，GPNA被设计为结合到基因转录物的转录和翻译起始位点，引起了有效的反义作用，具有异常低的细胞毒性。拟议的研究将探索这类特定分子的范围，并建立对控制细胞摄取，杂交和细胞毒性的因素和机制的基本理解，包括人类ES细胞在内的各种细胞系。具体而言，我们的目标是在未来五年内实现四个具体目标。目的1（第D.1节）：评价GPNA对人体细胞和ES细胞的反义作用;目的2（第D.2节）：优化细胞摄取、杂交和细胞毒性。目的3（第D.3节）：确定GPNA摄取和细胞毒性的机制。目标4（第D.4节）：确定GPNA在调节基因表达中的作用范围。 这项研究对未来设计和开发用于动物和人类的安全有效的核酸模拟物至关重要。据我们所知，GPNA是人类ES细胞摄取的第一个核酸类似物的例子。这很有趣，因为这些原始细胞非常难以穿透，并且对环境线索非常敏感。人类胚胎干细胞是了解人类早期发育的关键，既不能直接在子宫内研究，也不能用动物模型完全理解，更不用说它在再生医学方面的巨大潜力了。关于人ES细胞增殖、分化、细胞寿命等许多有趣的问题尚未得到解决。此外，最近的证据表明，癌症可能源于干细胞。如果这被证明是正确的，它将改变癌症治疗的过程。在我们开始解决这些问题之前，必须开发一种有效的方法来调节这些和相关细胞类型中的基因表达-这是我们研究的目的。

DESCRIPTION (provided by applicant): Recently we have developed a new class of peptide nucleic acid called GPNA that is readily taken up by both human somatic and embryonic stem cells, and binds sequence specifically to DNA and RNA. GPNA designed to bind to the transcription and translation start site of the gene transcript elicited potent antisense effect with unusually low cytotoxicity compared to the unmodified PNA and its other derivatives. The proposed research will explore the scope of this particular class of molecules and establish a basic understanding of the factors and mechanisms that control cellular uptake, hybridization and cytotoxicity over a wide range of cell lines, including human ES cells. Specifically, we aim to accomplish four specific objectives within the next five years. Aim 1 (Section D.1): Evaluate the antisense effects of GPNA with human somatic and ES cells; Aim 2 (Section D.2): Optimize cellular uptake, hybridization and cytotoxicity. Aim 3 (Section D.3): Determine the mechanism of GPNA uptake and cytotoxicity. And Aim 4 (Section D.4): Determine the scope of GPNA in regulating gene expression. The proposed study is crucial to the future design and development of nucleic acid mimics for a safe and effective use in animals and in humans. GPNA is, to the best of our knowledge, the first example of nucleic acid analogue taken up by human ES cells. This is intriguing because these primitive cells are extremely difficult to penetrate and are sensitive to the environmental cues. Human ES cells hold the key to understanding early human development that can neither be studied directly in utero nor fully understood with animals model, not to mention its enormous potential for regenerative medicine. Many fascinating questions concerning human ES cell proliferation, differentiation, cellular lifespan and so forth have not yet been addressed. Furthermore, recent evidence suggests that cancers may arise from stem cells. If this proves to be correct, it will alter the course of cancer treatment. Before we can begin to address these questions, an effective method must be developed to regulate gene expression in these and related cell types - this is the aim of our research.