Studies in Poxvirus Host Range Genes and Tropism

描述（由申请人提供）：我们建议检查介导一种特定痘病毒（即粘液瘤病毒（MV））的种属特异性和嗜性的分子途径。MV是一种兔特异性痘病毒，根据特定的兔物种诱导明显不同的疾病谱，但对包括人在内的所有其他宿主物种无致病性。我们实验室广泛研究了MV及其在兔中引起的疾病，称为粘液瘤病，作为研究痘病毒发病机制基本原理的模型系统，特别是通过利用我们大量且不断增长的靶向MV基因敲除构建体的集合。几年前，我们意外地发现，原代小鼠细胞，这是通常不允许MV感染，可以通过中断细胞干扰素（IFN）的反应，使完全允许。这项工作随后导致发现，大多数测试的人类癌细胞对MV是完全允许的，并且MV作为溶瘤疗法在各种动物模型中治疗人类癌症异种移植物是非常有效的。我们还发现，两种病毒宿主范围因子（M-T5和M063）对于许多人类癌细胞中的容许MV复制也是关键的，并且我们可以通过这些病毒蛋白及其宿主细胞蛋白靶标来操纵病毒容许性和溶瘤作用。最后，我们最近的观察与这一建议是，原代人类细胞的保护MV感染协同IFN和肿瘤坏死因子（TNF）。我们提出：1-评估病毒宿主范围因子在MV对人类癌细胞的嗜性中的作用。我们建议通过诱变、siRNA敲除、信号抑制剂和蛋白质芯片来分析MV宿主范围蛋白与宿主细胞信号分子（如Akt）的相互作用。我们将测试一些辅助策略，如用于人类癌症化疗的信号调节剂药物，以增加MV对更广泛的人类癌细胞的溶瘤潜力。2-研究宿主IFN和TNF应答在MV嗜性中的作用。我们将研究MV基因敲除病毒，这些病毒在几个关键的病毒宿主范围基因中被删除，以研究它们在抑制IFN或TNF反应，调节MV嗜性和溶瘤中的作用。这一新的信息将允许更合理的方法来优化MV病毒治疗对更广泛的人类癌症，并控制MV复制在原代非癌人类细胞。公共卫生相关性：以前，我们对痘病毒发病机制的研究更多地集中在研究一种称为粘液瘤病毒（MV）的特定兔特异性痘病毒在兔宿主中引起疾病的基本机制。最近，我们发现MV也感染和杀死广泛的人类癌细胞，我们现在已经使用MV成功地治疗了动物模型中的几种类型的癌症。在这项提议中，我们将研究两个关键的病毒宿主范围因子及其细胞信号传导通路靶点，以协助MV作为人类癌症新的溶瘤治疗药物的临床前开发。

DESCRIPTION (provided by applicant): We propose to examine the molecular pathways that mediate the species specificity and tropism of one particular poxvirus, namely myxoma virus (MV). MV is a rabbit-specific poxvirus that induces distinctly different disease profiles depending on the specific rabbit species but is nonpathogenic for every other host species, including man. Our lab has extensively studied MV and the disease it causes in rabbits, called myxomatosis, as a model system to investigate the fundamental principles of poxvirus pathogenesis, particularly by exploiting our large and growing collection of targeted MV gene knockout constructs. Several years ago, we unexpectedly discovered that primary mouse cells, which are normally nonpermissive for MV infection, could be rendered fully permissive by interrupting the cellular interferon (IFN) responses. This work then led to the discovery that the majority of human cancer cells tested were fully permissive for MV and that MV is remarkably effective as oncolytic therapy for the treatment of human cancer xenografts in a variety of animal models. We also discovered that two viral host range factors (M-T5 and M063) were also critical for permissive MV replication in many human cancer cells and we could manipulate viral permissiveness and oncolysis through these viral proteins and their host cell protein targets. Finally, our most recent observation related to this proposal is that primary human cells are protected from MV infection synergistically by IFN and tumor necrosis factor (TNF). We propose to: 1- Evaluate the roles of viral host range factors in MV tropism for human cancer cells. We propose to analyze the interactions of MV host range proteins with host cell signaling molecules (like Akt) by mutagenesis, siRNA knockdowns, signaling inhibitors and protein microarrays. We will test a number of adjunct strategies, like signaling modifier drugs used for cancer chemotherapy in man, to increase MV oncolytic potential for a wider spectrum of human cancer cells. 2- Investigate the role of host IFN and TNF responses in MV tropism. We will study MV knockout viruses that are deleted in several key viral host range genes to investigate their roles in inhibiting IFN or TNF responses, and modulating MV tropism and oncolysis. This new information will allow for more rational approaches to optimizing MV virotherapy against a wider spectrum of human cancers, and for controlling MV replication in primary noncancerous human cells. PUBLIC HEALTH RELEVANCE: Previously, our studies on poxvirus pathogenesis were more focused on studying the basic mechanisms by which one particular rabbit-specific poxvirus called myxoma virus (MV) causes disease in the rabbit host. Recently, we discovered that MV also infects and kills a wide spectrum of human cancer cells and we have now used MV to successfully treat several types of cancers in animal models. In this proposal, we will study two key viral host range factors and their cellular signaling pathway targets in order to assist the preclinical development of MV as a new oncolytic therapeutic for cancer in man.