Development of an effective DISC vaccine strategy against congenital CMV

说明(申请人提供：由于先天性感染的风险，开发巨细胞病毒(CMV)疫苗是一项主要的公共卫生优先事项。CMV临床致病株与实验室适应株的不同之处在于，它们保留了感染上皮细胞和内皮细胞的能力。根据胎盘的结构，内皮细胞可能是启动病毒通过胎盘转移到胎儿的潜在重要部位。病毒进入内皮细胞的机制不同于进入成纤维细胞的gB途径，因为它需要病毒蛋白Gh/Gl/UL128-131形成内吞复合体才能使病毒进入细胞。豚鼠是目前唯一可以用种特异性豚鼠巨细胞病毒(GPCMV)研究先天性巨细胞病毒的小型动物模型。我们最近调查了复制受损的GPCMV活疫苗的使用，该疫苗需要补充细胞系来生产传染性病毒。这种失效的传染性单周期(圆盘)疫苗策略导致了VIRS，它可以感染细胞表达一系列病毒抗原，并诱导免疫反应，但不产生传染性病毒。这种疫苗策略在对重要病毒抗原产生抗体和T细胞反应的豚鼠中具有高度的免疫原性。然而，我们最初的圆盘疫苗缺乏一个新发现的与人CMV UL128-131(GP128-131)同源的基因座。因此，这种疫苗策略缺乏针对潜在的GPCMV内吞复合体产生免疫反应的能力。在这项申请中，我们建议定义GPCMV进入内皮细胞的要求，并另外开发携带必要抗原的第二代圆盘疫苗，以诱导有效的中和免疫反应，对抗表观/内皮细胞的病毒感染。作为第二代圆盘疫苗策略的一部分，病毒将被进一步减毒，以提高免疫原性，并通过敲除pp71同源物来降低病毒建立潜伏期的能力。第二代圆盘疫苗预防先天性GPCMV的能力将被调查，并通过将其与先前在该模型中调查的重组GB疫苗策略进行比较来确定有效性。

DESCRIPTION (provided by applicant: Development of a cytomegalovirus (CMV) vaccine is a major public health priority due to the risk of congenital infection. Pathogenic clinical strains o CMV differ from lab adapted strains in that they retain the ability to infect epithelial and endothelial cells. Based on the structure of the placenta endothelial cells could be a potentially important site for initiation of virus transfer across the placenta to the fetus. The mechanism of virus entry into epi/endothelial cells is different from the gB pathway of entry into fibroblast cels as it requires viral proteins gH/gL/UL128-131 forming an endocytic complex to enable viral cell entry. The guinea pig is the only small animal model that allows the study of congenital CMV by using species specific guinea pig CMV (GPCMV). We recently investigated the use of a replication-impaired live GPCMV vaccine that requires a complementing cell line for production of infectious virus. This disabled infectious single cycle (DISC) vaccine strategy results in a virs that can infect cells to express an array of viral antigens and induce an immune response but does not produce infectious virus. This vaccine strategy was highly immunogenic in guinea pigs producing antibody and T cell responses to important viral antigens. However, our original DISC vaccine lacked a newly identified homolog locus to human CMV UL128-131 (GP128-131). Consequently, this vaccine strategy lacks the ability to generate an immune response against a potential GPCMV endocytic complex. In this application we propose to define requirements for GPCMV entry into endothelial cells and additionally develop a 2nd generation DISC vaccine carrying the essential antigens necessary to induce an effective neutralizing immune response against viral infection of epi/endothelial cells. As part of this 2nd generation DISC vaccine strategy the virus will be further attenuated to increase immunogenicity and decrease the ability of the virus to establish latency by the knockout of the pp71 homolog. The ability of second generation DISC vaccines to protect against congenital GPCMV will be investigated and efficacy determined by comparing it with a recombinant gB vaccine strategy previously investigated in this model.