Generating Native Env Glycosylation in HIV Vaccine Candidates

 描述(由申请人提供)：通过接种疫苗预防人类免疫缺陷病毒(HIV-1)传播是一个非常可取但难以实现的目标。疫苗诱导的抗体结合HIV-1包膜糖蛋白(Env)尖峰并中和病毒被认为是保护性免疫反应的理想组成部分；因此，含有Env外部部分的重组表达的免疫原通常被认为是预防性疫苗的基本成分。HIV-1env是高度糖基化的，糖链约占糖蛋白质量的50%。这些多糖对许多病毒功能有很大贡献，包括HIV-1逃避抗体反应的能力。基于环境病毒的候选疫苗上的多糖的存在和类型影响蛋白质的免疫原性和抗原性，其中几个gp120和gp41多糖作为有效的HIV-1中和抗体靶向的表位或表位的组成部分。如果在疫苗接种期间有希望激发这些类型的抗体，那么Env疫苗上的葡聚糖必须与病毒上发现的相同类型。因此，HIV-1疫苗开发领域面临的两个关键问题是：(1)天然HIV-1 Env的糖链是什么？以及(2)如何在以环境为基础的疫苗中重现这种糖链特征？回答这两个基本问题将代表着HIV-1疫苗研究的重大进步：如果候选疫苗的糖链图谱与本地病毒的糖链图谱相匹配，那么产生包含基于糖链表位的有效抗体的机会将会增加。这项应用的两个目标是表征来自T淋巴细胞的最自然来源的Env的糖基化，并确定在更容易处理的细胞系中表达具有正确糖基化的Env的方法，该方法可用于生产用于人类临床试验的Env的GMP。这个项目将由一个协同、协作、优势互补的团队来运行。该小组最近发表了HIV-1膜环境三聚体的糖基化图谱，利用了在环境蛋白生产、样品制备和糖基化分析方面高度相关的技术创新；这些最新进展为本文提出的研究提供了坚实的基础。该项目的成功完成将为所有疫苗开发商提供他们所需的工具，以便在他们选择的环境中生成本地糖基化特征。

 DESCRIPTION (provided by applicant): Preventing human immunodeficiency virus (HIV-1) transmission by vaccination is a highly desirable, but elusive, goal. Vaccine-induced antibodies that bind the HIV-1 envelope glycoprotein (Env) spike and neutralize the virus are thought to be a desirable component of a protective immune response; as a consequence, recombinantly expressed immunogens containing the exterior portion of Env are generally regarded as an essential element of a prophylactic vaccine. The HIV-1 Env is highly glycosylated, with glycans accounting for about 50% of the glycoprotein mass. These glycans contribute significantly to a number of viral functions, including the ability of HIV-1 to evade antibody responses. The presence and type of glycans on Env-based vaccine candidates influence both immunogenicity and antigenicity of the protein, and several of the gp120 and gp41 glycans serve as epitopes or components of epitopes targeted by potent HIV-1-neutralizing antibodies. If there is any hope of eliciting these types of antibodies during vaccination, the glycans on the Env vaccine must be of the same type as those found on the virus. Therefore, two critical questions facing the field of HIV-1 vaccine development with respect to Env glycosylation are: (1) What is the glycan profile of native HIV-1 Env? and (2) How can that glycan profile be recapitulated in an Env-based vaccine? Answering these two fundamental questions would represent a significant advance in HIV-1 vaccine research: If the glycan profile of a vaccine candidate could match that of native virus, the chances of eliciting effective antibodies that contain glycan-based epitopes would be increased. The two goals of this application are to characterize the glycosylation of Env from the most native source possible, from T-lymphocytes, and to identify ways to express Env with the correct glycosylation in a more tractable cell line that could be used in GMP manufacturing of Env for human clinical trials. This project will be run by a synergistic, collaborative team wit complementary strengths. The group has recently published the glycosylation profile of an HIV-1 membrane Env trimer, taking advantage of highly relevant technical innovations in Env glycoprotein production, sample preparation, and glycosylation analysis; these recent developments provide a strong foundation for the studies proposed herein. The successful completion of the project will provide all vaccine developers with the tools they need to generate native glycosylation profiles on their Env of choice.