Vaccine against CMV endothelial tropism & congenital infection

描述（由申请方提供）：内皮细胞和上皮细胞是HCMV的主要体内靶点，并且可能是先天性感染中病毒通过胎盘转移至胎儿的重要起始部位。在HCMV中，gH/gL糖蛋白复合物和来自UL 128 -131基因座的病毒蛋白是内吞病毒进入内皮/上皮细胞所必需的。该病毒进入途径与成纤维细胞中发现的gB进入途径不同。针对UL 128-UL 131基因座和内吞复合物的蛋白质的抗体可以中和内皮/上皮细胞的病毒感染。我们假设，包括内吞病毒进入途径的组分作为中和靶抗原的疫苗策略将是针对先天性感染的更有效的疫苗。由于HCMV的种属特异性，将使用豚鼠巨细胞病毒（GPCMV）进行研究。豚鼠是唯一的小动物模型，允许先天性巨细胞病毒感染的研究。研究真正的全长CMV和病毒进入内皮/上皮细胞的另一个复杂性是当病毒在成纤维细胞系上生长时UL 128 -131基因座缺失的倾向。这在HCMV和GPCMV中都发生，这使得对内皮/上皮细胞的感染研究复杂化。我们建议使用一种新的方法，慢病毒转导tet-off互补细胞系系统表达缺失的GPCMV基因（GP 128 -131），以产生假型GPCMV编码部分或全部的假定的内吞途径的组成部分。我们的假设是，我们将能够确定相关的蛋白质所必需的内吞途径的GPCMV感染的内皮细胞通过这一战略。此外，我们假设，一旦内吞途径蛋白复合物的最小组分被鉴定，它们可以被包括作为基于复制受损（DISC）病毒的新型疫苗策略的一部分，与缺乏该位点的DISC病毒相比，所述DISC病毒将增强针对先天性CMV的保护性免疫应答。

DESCRIPTION (provided by applicant): Endothelial and epithelial cells are a major target in vivo for HCMV and potentially an important site for initiation of virus transfer across the placenta to the fetus in congenital infection. In HCMV, the gH/gL glycoprotein complex and viral proteins from the UL128-131 locus are required for endocytic viral entry into endothelial/epithelial cells. This viral entry pathway is separate from the gB entry pathway found in fibroblast cells. Antibodies against proteins of the UL128-UL131 locus and endocytic complex can neutralize virus infection of endothelial/epithelial cells. We hypothesize that a vaccine strategy which includes components of the endocytic viral entry pathway as neutralizing target antigens would be a more effective vaccine against congenital infection. The studies will be carried out using guinea pig cytomegalovirus (GPCMV) because of species specificity of HCMV. The guinea pig is the only small animal model that allows the study of congenital cytomegalovirus infection. An added complication of studying bona fide full length CMV and virus entry into endothelial/epithelial cells is the propensity for deletion of the UL128-131 locus when virus is grown on fibroblast cell lines. This happens in both HCMV and GPCMV, which complicates infection studies on endo/epithelial cells. We propose to use a novel approach of a lentivirus transduced tet-off complementing cell line system expressing missing GPCMV genes (GP128-131) to produce pseudotyped GPCMV encoding part or all of the components of the presumed endocytic pathway. Our hypothesis is that we will be able to determine the relevant proteins necessary for the endocytic pathway of GPCMV infection of endothelial cells via this strategy. Furthermore, we hypothesize that once the minimum components of the endocytic pathway protein complex are identified they can be included as part of a novel vaccine strategy based on a replication impaired (DISC) virus which will enhance the protective immune response against congenital CMV compared to a DISC virus lacking this locus.