Chemically Modified siRNAs for the Treatment of Gastrointestinal Cancer

胃肠道恶性肿瘤是英国第二常见的癌症，占英国癌症死亡人数的25%。食道腺癌和胰腺癌的预后特别差，5年存活率仅为5%。胃泌素基因在胃肠腺癌的早期表达，促进癌前病变的进展。除了作为生长激素，它还通过增加侵袭性和粘附性来保护细胞免受凋亡、刺激血管形成和增加转移的可能性。胃泌素抗体能够抑制这些生物效应，最近成功地完成了用于治疗胰腺癌的第三阶段临床试验。虽然这是概念的证明，但它们只部分中和了胃泌素基因的产物。这种反义方法提供了下调参与疾病建立和发展的关键癌基因的希望。传统的反义技术已经成功地进入临床，作为CMV视网膜炎的局部治疗，但它的成功受到了限制，因为需要使用高全身剂量来达到局部有效剂量。胃泌素反义寡核苷酸已被证明能有效阻断胃泌素介导的致癌作用。由于使用了细胞内自然发生的催化过程，siRNA是一种更有效的下调基因的方法。因此，这种方法的成功机会相当高，前提是能够开发出稳定并在体内传递siRNA的方法。本提案中描述的工作旨在为与基于RNA的治疗剂相关的稳定性和传递问题提供化学解决方案。我们将使用胃泌素基因作为测试系统来研究骨架修饰对siRNAs水解稳定性的影响，并评估它们在肿瘤细胞系中启动siRNA反应的能力。我们还将解决细胞特异性靶向和细胞质递送的问题，方法是将一些针对胃泌素/CCK-2受体的配体与已知的分子转运蛋白多肽结合在一起。

Gastrointestinal (GI) malignancy is the second most common cancer in the UK and accounts for 25% of UK cancer deaths. Oesophageal and pancreatic adenocarcinomas have a particularly poor prognosis, with 5-year survival rates of only 5%. The gastrin gene is expressed early in the development of gastrointestinal adenocarcinomas, promoting the progression of premalignant lesions. In addition to acting as a growth hormone, it protects,cells against apoptosis, stimulates blood vessel formation and increases the potential for metastasis by increasing invasion and adhesion. Antibodies to gastrin are able to inhibit these biological effects and have recently successfully completed a Phase III clinical trial for use in the treatment of pancreatic cancer. Whilst this is proof of concept, they only partially neutralise the products of the gastrin gene. The antisense approach offers hope of downregulating key oncogenes involved in disease establishment and progression. Traditional antisense technology has successfully reached the clinic as a topical treatment for CMV retinitis but its success has been limited by the need to use high systemic doses to achieve effective doses locally. Gastrin antisense has proven to be effective in blocking gastrin-mediated carcinogenesis. siRNAs are a more potent means of downregulating genes due to use of a naturally-occurring catalytic process within the cell. Thus the chances of success with this approach are considerably higher, provided that methods of stabilising and delivering siRNAs'in vivo' can be developed.The work described in this proposal aims to provide chemistry solutions to both the stability and delivery problems associated with RNA-based therapeutic agents. We will study the effects of backbone modifications upon the hydrolytic stability of siRNAs using the gastrin gene as the test system and assess their ability to initiate an siRNA response in tumour cell lines. We will also address the problem of cell-specific targeting and cytoplasmic delivery by covalent attachment of a number of ligands targeted at the gastrin/CCK-2 receptor in conjunction with known 'molecular transporter' peptides.