Optimization of Tumor Targeted HSV for Human Use

项目1有两个主要目标。第一个是提高候选治疗的有效性 通过缺失特定基因构建的病毒。这些病毒在治疗中更有效 将病毒注射到肿瘤和放射治疗相结合。尽管如此，联合治疗的有效性 治疗依赖于肿瘤的基因型别。在过去的5年里，我们与项目2合作：(A)确定 电离辐射(IR)增强减毒病毒的分子基础和(B)鉴定 细胞基因(MEK)，其产物当与IR正确结合使用时，可以克服 肿瘤基因型别对减弱病毒复制的限制。这些研究的结果是 在项目2的进度报告中描述。我们的第二个目标是构建仅能 感染癌细胞，但不感染正常细胞。从本质上讲，我们削弱了HSV-1与肝素的结合能力 通过其天然的蛋白受体HveA和Nectin 1进入细胞。因此， 工程病毒R5141仅通过IL13a2受体感染细胞，而R5181通过IL13a2受体进入细胞 尿激酶型纤溶酶原激活剂受体。我们的发现极大地有助于理解 糖蛋白(G)D的生物学和生物化学我们的新目标如下：目标1将开发 临床级靶向病毒的生产方法。由于目标病毒必须在 稳定表达新型受体的非肿瘤细胞(S)，我们提出了几种方法，可以 产生临床级病毒。AIM 2的目标是开发出更有效的病毒用于治疗 Gbm与AY34.5突变病毒。目前正在进行临床试验的治疗性病毒延长了患者的存活时间 一小部分治疗患者以肿瘤基因依赖的方式进行治疗。我们已经构建了一种病毒 组成活性的MEK基因由IR可诱导启动子(R2660)驱动。在预赛中 R2660加IR的研究阻止了对病毒或单独IR的肿瘤的生长。这款车的安全特性 以及其他变异病毒将被研究。AIM 3的目标是呈现针对特定目标的病毒 细胞表面的受体更有效。我们已经建立了原则证明，但不考虑 当前世代的最佳选择。我们已经确定了这些病毒的具体缺点和改进方法 AIM 4的目标是基于一项新的观察结果，即 尿激酶型纤溶酶原激活剂受体与GD胞外区的羧基末端相互作用。这个 尿激酶型纤溶酶原激活剂的氨基末端含有一个与赖氨酸有亲和力的Kringle结构域。因此， 我们将确定人纤溶酶原的Kringle结构域是否可用于靶向GD，并通过 延伸，靶向HSV-1的配体，葡萄糖调节蛋白78。 相关性(请参阅说明)： 项目1是计划项目赠款的组成部分，旨在治愈或至少有效地延长质量 恶性胶质瘤患者的生活质量。在这一点上，原则的证明已经建立在 无论是基础水平还是临床水平。项目1面临的任务是设计、建造和测试下一个 产生以增强的治疗轮廓为特征的治疗性病毒，并开发方法 它们在GMP设施中生产。新病毒的验证将与项目2合作完成 和3，最终是项目4。

Project 1 has had two major objectives. The first was to enhance effectiveness of candidate therapeutic viruses constructed by deletion of specific genes. These viruses are more effective in treatment that combines virus administration to the tumor and radiation. Nevertheless, the effectiveness of combined therapy is tumor genotype-dependent. In the past 5 years, we have collaborated with Project 2 to (a) identify the molecular basis for enhancement of attenuated viruses by ionizing radiation (IR) and (b) identify a cellular gene (MEK) whose product, when used properly in conjunction with IR, could overcome the restriction to attenuated virus replication imposed by tumor genotypes. The results of these studies are described in the Progress Report for Project 2. Our second objective was to construct viruses that can only infect cancer cells but not normal cells. In essence we ablated the ability of HSV-1 to attach to heparin sulfate proteoglycans and to enter cells by way of its natural protein receptors, HveA and nectin 1. Thus, the engineered virus R5141 infects cells solely via the IL13 a2 receptor while R5181 enters cells via the urokinase plasminogen activator receptor. Our findings have significantly contributed to understanding the biology and biochemistry of glycoprotein (g) D. Our new objectives are as follows: AIM 1 will develop methods for production of clinical grade targeted viruses. Since targeted viruses must be produced in noncancerous cells that stably express the novel receptor(s), we propose several ways in which we can produce clinical grade viruses. The objective of AIM 2 is to develop more effective viruses for therapy of GBM with AY34.5 mutant viruses. Therapeutic viruses currently in clinical trials extend survival time in a small fraction of treated patients in a tumor genotype dependent manner. We have constructed a virus in which the constitutively active MEK gene is driven by an IR inducible promoter (R2660). In preliminary studies R2660 plus IR blocked growth of a tumor resistant to virus or IR alone. The safety features of this and other mutant viruses will be studied. The objective of AIM 3 is to render the viruses targeting specific receptors on cell surfaces more effective. We have established proof of principle but do not consider the current generation optimal. We have identified specific shortcomings and ways to improve these viruses The objective of AIM 4 is based on the novel observation that a component of the amino-terminal domain of the urokinase plasminogen activator receptor interacts with the carboxyl-terminus of the gD ectodomain. The amino terminus of urokinase plasminogen activator contains a Kringle domain with affinity for lysines. Thus, we will determine whether the Kringle domain of human plasminogen can be used to target gD and by extension, target HSV-1 to its ligand, the glucose regulatory protein 78. RELEVANCE (See instructions): Project 1 is a component of a Program Project Grant designed to cure or at least effectively prolong quality of life of patients with malignant gliomas. At this point in time the proof of principle has been established at both basic and clinical levels. The task confronting Project 1 is to design, construct and test the next generation of therapeutic viruses characterized by enhanced therapeutic profile and to develop methods for their production in GMP facilities. Validation ofthe novel viruses will be done in collaboration with Projects 2 and 3 and, ultimately, with Project 4.