GP350 AS A NOVEL VACCINE PROTEIN CARRIER

GP350 作为新型疫苗蛋白载体

• 批准号: 7349607
• 负责人: CLIFFORD M SNAPPER
• 金额: $ 3.5万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-05-01 至 2007-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7349607
• 关键词: GP350; NOVEL; VACCINE; PROTEIN; CARRIER

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Purpose of the experiment: gp350 is an Epstein-Barr virus (EBV) protein that binds to CD21 on human and monkey, but not mouse, B cells. Previous studies have demonstrated that the co-crosslinking of CD21 and the B cell antigen receptor (BCR) on the surface of the B cell is highly synergistic for B cell activation and significantly lowers the amount of antigen required for optimal immunogenicity. CD21 is also expressed on follicular dendritic cells (FDC), present within germinal centers, where it is used to trap antigen (via bound complement) on the surface of the FDC and thus augment the germinal center reaction and subsequent immunity. A recent study demonstrated that linkage of C3d to a polysaccharide antigen leads to improved anti-polysaccharide responses in vivo. Thus, we propose that gp350 can be used as a novel carrier protein for polysaccharide conjugate vaccines in which gp350 not only serves to recruit T cell help (on the basis of it being a foreign protein) for the anti-polysaccharide response but provides an adjuvanting affect as well (on the basis of its ability to bind CD21), potentially allowing for the use of much lower doses of antigen in vivo. Further, we are constructing a dimer of gp350 that can be conjugated to peptides or proteins and used in a similar fashion to augment anti-peptide or anti-protein Ig responses. Preparation of conjugates: We have covalently linked gp350 to pneumococcal capsular polysaccharide, serotype 14 (PPS14). As a control we use the same chemistry and conjugation conditions to produce a similar conjugate of diphtheria toxoid (DT) covalently linked to PPS14. Further, we confirmed that the gp350-PPS14 conjugate retained its binding activity to CD21, using a CD21-expressing cell line. We also demonstrated that both conjugates are immunogenic in mice for both the anti-PPS14 and anti-protein response. Finally, we confirmed that the gp350 also specifically binds to peripheral blood B cells derived from either Cynomolgus or rhesus monkeys. Goals of current experiment: The goal of this initial experiment is a PROOF of PRINCIPLE, and are to directly compare the two conjugates using what historically should be optimal doses of antigen. This will give us baseline data that we can indeed elicit specific Ig responses in monkeys using these conjugates and to directly compare the quantitative responses between these two conjugates.

这个子项目是利用由NIH/NCRR资助的中心拨款提供的资源的许多研究子项目之一。子项目和调查员(PI)可能从另一个NIH来源获得了主要资金，因此可能会出现在其他CRISE条目中。列出的机构是针对中心的，而不一定是针对调查员的机构。实验目的：gp350是一种EB病毒(EBV)蛋白，可与人和猴子的CD21结合，但不能与小鼠的B细胞结合。以往的研究表明，CD21与B细胞表面的B细胞抗原受体(BCR)的共交联对B细胞的激活具有高度的协同作用，并显著降低了最佳免疫原性所需的抗原量。CD21也表达在毛囊树突状细胞(FDC)上，存在于生发中心，在那里它被用来捕获FDC表面的抗原(通过结合的补体)，从而增强生发中心反应和随后的免疫。最近的一项研究表明，C3d与多糖类抗原的连接可以改善体内的抗多糖类反应。因此，我们建议gp350可以作为一种新的载体蛋白用于多糖结合疫苗，其中gp350不仅为抗多糖反应招募T细胞帮助(基于它是外源蛋白)，而且还提供佐剂作用(基于其结合CD21的能力)，潜在地允许在体内使用更低剂量的抗原。此外，我们正在构建一种gp350二聚体，它可以与多肽或蛋白质偶联，并以类似的方式用于增强抗肽或抗蛋白Ig反应。偶联物的制备：我们已将gp350共价连接到肺炎链球菌14型衣壳多糖(PPS14)上。作为对照，我们使用相同的化学和连接条件来产生类似的与PPS14共价连接的白喉类毒素(DT)的连接物。此外，我们利用表达CD21的细胞系证实了gp350-PPS14结合物保持了其与CD21的结合活性。我们还证明了这两种结合物对小鼠的抗PPS14和抗蛋白质反应都是免疫原性的。最后，我们证实gp350也能与食蟹猴或恒河猴外周血中的B细胞特异性结合。当前实验的目标：这个初始实验的目标是证明原理，并使用历史上应该是最佳剂量的抗原直接比较这两种结合物。这将给我们提供基线数据，我们确实可以使用这些结合物在猴子身上诱导特异性Ig反应，并直接比较这两种结合物之间的定量反应。