Adenovirus E1B55K Functions Related to Oncolytic Replication

描述（由申请人提供）：所有癌细胞的两个主要“特征”包括细胞周期失调和细胞凋亡抑制，这两者也与腺病毒（Ad）感染有关。这两个过程主要由病毒癌蛋白E1A和E1B进行。由于病毒E1蛋白的功能与癌细胞因子类似，具有促进增殖、抑制凋亡的功能，因此致癌蛋白E1A和E1B突变的病毒可以在癌细胞中选择性复制。其中一种E1B55K缺失的突变体dl1520已被用于临床试验，记录了溶瘤作用。然而，病毒溶瘤复制的机制尚未明确，治疗效果有待提高。一般认为E1B55K的主要作用是结合并抑制p53的激活，但许多研究证明E1B55K介导的p53失活并不是病毒复制所必需的。本实验室研究表明：(1)突变病毒在大肿瘤中的有限传播是导致治疗效果下降的关键因素；(2) Ad - E1A表达增加可促进病毒溶瘤复制；(3) E1B缺失导致的细胞凋亡可以部分减少病毒复制，但不改变病毒介导的癌症杀伤的最终结果。我们最近的研究发现，E1B55K在诱导细胞周期蛋白E和其他细胞周期相关基因中具有新的功能。最重要的是，我们还观察到周期蛋白E表达的增加与病毒复制效率相关。在正常细胞中，e1b55k诱导的周期蛋白E表达是病毒复制所必需的，但在癌细胞中则不是必需的。我们假设E1B55K可能靶向细胞因子来增加cyclin E的表达，而该因子可能已经在癌细胞中被激活。因此，癌细胞中细胞周期蛋白E的失调可能是e1b55k缺失病毒溶瘤复制的分子基础。我们的研究团队将(1)确定E1B55K诱导cyclin E的靶向细胞因子，(2)明确E1B55K激活cyclin E表达的机制，(3)确定cyclin E过表达与e1b缺失病毒溶瘤复制的关系。如果我们证实溶瘤复制依赖于细胞周期蛋白E的表达和细胞增殖，恶性肿瘤生长和细胞周期蛋白E失调的患者应该从这种腺病毒治疗中获益。这项工作的长期目标是提高溶瘤性癌症基因治疗的疗效。公共卫生相关性：缺乏一种重要的调节蛋白e1b55k的腺病毒仍然可以在一些癌细胞中扩增。因此，E1B55K突变体dl1520已被用于临床试验。了解E1B55K的功能和E1B55K缺失的dl1520在癌细胞中的选择性复制是很重要的。我们最近发表的研究表明，E1B55K在诱导细胞周期蛋白E表达方面具有新的功能，这对DNA复制至关重要。癌细胞通常表达高水平的细胞周期蛋白E或有基因失调。我们认为病毒E1B55K可能激活一些细胞因子，增加周期蛋白E的表达，从而促进病毒DNA复制。癌细胞可能已经激活了这些因子；因此癌细胞不需要E1B55K的功能。研究这种可能性是非常重要的。如果我们证实突变的病毒复制依赖于细胞周期蛋白E的表达，恶性肿瘤和细胞周期蛋白E失调的患者应该从这种腺病毒治疗中获益。这项工作的长期目标是提高溶瘤性癌症基因治疗的疗效。

DESCRIPTION (provided by applicant): Two major "hallmarks" of all cancer cells include dysregulated cell cycle and inhibited apoptosis, both of which are also involved in adenovirus (Ad) infection. These two processes are primarily conducted by viral oncoproteins E1A and E1B. Since the function of viral E1 protein is similar to that of cancer cellular factors that promote proliferation and inhibit apoptosis, viruses with mutations in oncoprotein E1A and E1B can selectively replicate in cancer cells. One such mutant with deletion of E1B55K, known as dl1520, has been used in clinical trials documenting oncolytic effects. However, the mechanism of viral oncolytic replication has not been well characterized, and the therapeutic efficacy needs improvement. It is generally believed that the major role of E1B55K is to bind to and inhibit p53 activation, but many studies have documented that E1B55K-mediated p53 inactivation is not required for virus replication. Our laboratory has shown: (1) that the limited spread of mutant viruses in large tumors is a key factor in decreased therapeutic efficacy; (2) increased Ad E1A expression enhances virus-oncolytic replication; and (3) apoptosis caused by E1B deletion can partially decrease virus replication but does not change the final outcome of virus-mediated cancer killing. Our recent studies have revealed that E1B55K has a novel function in the induction of cyclin E and other cell cycle-related genes. Most importantly, we also observed that increased cyclin E expression is correlated with virus replication efficiency. E1B55K-induced cyclin E expression is required for virus replication in normal cells, but is not necessary in cancer cells. We hypothesize that E1B55K may target cellular factor(s) to increase cyclin E expression, and this factor(s) may already be activated in cancer cells. Thus, cyclin E dysregulation in cancer cells may be the molecular basis for oncolytic replication of E1B55K-deleted viruses. Our research team will (1) identify cellular factors targeted by E1B55K for cyclin E induction, (2) define the mechanism by which E1B55K activates cyclin E expression, and (3) determine the relationship between cyclin E overexpression and oncolytic replication of E1B-deleted viriuses. If we confirm that oncolytic replication relies on cyclin E expression and cell proliferation, patients with aggressively growing tumors and dysregulated cyclin E should greatly benefit from this adenoviral therapy. The long-term goal of this work is to increase the efficacy of oncolytic cancer gene therapy. PUBLIC HEALTH RELEVANCE: Adenoviruses lacking an important regulative protein-E1B55K-still can amplify in some cancer cells. Therefore, the E1B55K mutant dl1520 has been used in clinical trials. It is important to understand the E1B55K function and the selective replication of E1B55K-deleted dl1520 in cancer cells. Our recently published studies have revealed that E1B55K has a novel function in the induction of cyclin E expression, which is crucial for DNA replication. Cancer cells generally express high levels of cyclin E or have a dysregulation of the gene. We reason that viral E1B55K may activate some cellular factors that increase cyclin E expression for viral DNA replication. Cancer cells may already have the factors activated; therefore cancer cells do not require the E1B55K function. The study of this possibility is very important. If we confirm that mutated virus replication relies on cyclin E expression, patients with aggressively growing tumors and dysregulated cyclin E should greatly benefit from this adenoviral therapy. The long-term goal of this work is to increase the efficacy of oncolytic cancer gene therapy.