Matrilysin-Sensing Gene Delivery Vectors for Colorectal Cancer Therapy

项目摘要 结直肠癌一旦转移，就会成为一种致命的疾病，5年生存率为 大约10%。可以特异性靶向转移性结直肠肿瘤细胞的有效治疗剂是 非常需要。递送核酸（例如基因或RNAi）以对抗癌症是非常有前途的方法。 不幸的是，将基因载体靶向递送到肿瘤细胞在很大程度上是困难的。 实现。迄今为止，大多数载体靶向方法依赖于在细胞表面上过表达的细胞表面受体。 一些靶癌细胞亚群。不幸的是，没有独特的细胞表面生物标志物， 能特异性识别肿瘤中的所有细胞为了克服这一限制，我们建议开发蛋白酶- 在转移性结直肠肿瘤中使用过表达的细胞外蛋白酶的可活化病毒（PAV） 微环境作为生物标志物来实现靶向递送。具体而言，基质溶解素（也称为 基质金属蛋白酶7，MMP 7）在结肠直肠癌中过表达。高水平的 肿瘤微环境中的MMP 7将以局部方式激活PAV，并使载体能够 结合广泛表达的细胞受体，包括结肠直肠癌细胞，并介导有效的 基因传递我们的PAV技术基于临床上有前途的腺相关病毒（AV），该病毒 最近被批准为欧洲第一个人类基因治疗产品。我们有关键的飞行员数据 这表明我们已经创造了MMP 7传感PAVs，大大提高了它们的基因传递效率， 一旦暴露于蛋白酶。此外，在原位癌症模型中，PAV原型能够 显著增加肿瘤中转基因递送和表达。在目标1中，我们将综合和 表征一组MMP 7感测PAV。我们的设计过程将利用理性和组合 方法，以加快实现设计解决方案。在目标2中，我们将测试基因递送 PAV在体外对结直肠癌细胞的作用，并将进行机制研究以探索PAV在结直肠癌细胞中的作用。 PAV与细胞的相互作用。最后，我们将在转移性结直肠癌的原位模型中测试PAV。 以确定它们的体内特异性和治疗功效。如果成功，该项目将 产生蛋白酶响应性AAV载体，其可能成为转移性肿瘤的可行治疗选择， 结肠直肠癌

PROJECT SUMMARY Once colorectal cancer metastasizes, it becomes a lethal disease with a 5-year survival rate of approximately 10%. Effective therapeutics that can specifically target metastatic colorectal tumor cells are sorely needed. Delivery of nucleic-acids (e.g. genes or RNAi) to combat cancer is a highly promising therapeutic approach; unfortunately, targeted delivery of gene vectors to tumor cells has been largely difficult to achieve. Most vector targeting approaches to date have relied on cell surface receptors overexpressed on some subpopulation of target cancer cells. Unfortunately, there is no unique cell surface biomarker that specifically identifies all cells in a tumor. To overcome this limitation, we propose to develop protease- activatable viruses (PAVs) that use extracellular proteases overexpressed in metastatic colorectal tumor microenvironments as the biomarkers to achieve targeted delivery. Specifically, matrilysin (also known as matrix metalloproteinase 7, MMP7) has been shown to be overexpressed in colorectal cancer. High levels of MMP7 in the tumor microenvironment will activate the PAVs in a localized manner and enable the vectors to bind cellular receptors that are broadly expressed, including on colorectal cancer cells, and mediate efficient gene delivery. Our PAV technology is based on the clinically promising adeno-associated virus (AAV), which has recently been approved as the first human gene therapy product in Europe. We have key pilot data demonstrating we have created MMP7-sensing PAVs that dramatically increase their gene delivery efficiency once exposed to the protease. Moreover, in an orthotopic cancer model, a PAV prototype is able to significantly increase transgene delivery and expression in tumors. In aim 1, we will synthesize and characterize a panel of MMP7-sensing PAVs. Our design process will harness both rational and combinatorial approaches in order to expedite achievement of the design solution. In aim 2, we will test the gene delivery performance of PAVs in vitro on colorectal cancer cells, and mechanistic studies will be done to probe the interaction of PAVs with the cells. Finally, we will test the PAVs in an orthotopic model of metastatic colorectal cancer in order to determine their in vivo specificity and therapeutic efficacy. If successful, this project will generate protease-responsive AAV vectors that may become viable therapeutic options for metastatic colorectal cancer.