Mature virus-like particles as a new strategy for dengue virus vaccines

描述（由申请人提供）：尽管在开发针对四种血清型登革热病毒（DENV）的四价疫苗方面取得了相当大的努力和进展，但尚未解决的主要挑战之一是难以激发针对所有四种血清型的平衡中和（NT）抗体（Abs）并降低抗体依赖性增强（ADE）的风险。主要由交叉反应性和弱或非NT抗体介导。对DENV感染后的人抗体的研究表明，识别包膜(E)蛋白融合环（FL）的交叉反应性和弱或非NT抗体比类型特异性和强效NT抗体具有免疫优势。以及交叉反应性和弱或非NT抗前膜（prM）抗体的存在。这种免疫优势是否可以被调节以诱导强效NT抗体，而不存在交叉反应性和弱或非NT抗体，目前尚不清楚。我们最近发现一种表达成熟DENV颗粒的DNA疫苗诱导了强效NT抗体，而交叉反应最小，弱或非NT抗体，这表明DENV颗粒的特异性调节可能诱导了更好的抗体反应。我们的长期目标是开发一种安全有效的DENV疫苗。本研究的目的是了解DENV E蛋白的免疫优势性，并测试成熟或fl修饰的成熟DENV颗粒是否能诱导强效NT抗体，而不产生交叉反应，弱或非NT抗体。中心假设是成熟DENV颗粒诱导强效NT抗体，较少的抗fl抗体和无抗prm抗体，从而与混合DENV颗粒相比降低ADE的风险。目标将通过三个特定目标来实现：第一个特定目标是确定E蛋白表位以及大量抗E单抗在成熟、混合和未成熟DENV颗粒上的可及性、结合亲和度和NT效力。我们假设不同的抗原表位可及性和抗E抗体对不同DENV颗粒的结合亲和力可能解释了E蛋白的免疫优势。第二个目的是证明表达成熟DENV颗粒的DNA疫苗比表达混合颗粒的DNA疫苗在诱导强效NT抗体和最小潜力增强抗体方面的优势，并在远交种小鼠和AG129小鼠（一种已建立的登革热小鼠模型）中提供保护。第三个目的是证明表达fl修饰的成熟DENV颗粒的DNA疫苗比表达未修饰的成熟DENV颗粒的DNA疫苗在激发强效NT抗体和最小潜力增强抗体方面的优势，并在小鼠模型中提供保护。该研究的意义在于详细了解了DENV E蛋白的免疫优势，并证明了这种免疫优势可以通过成熟的和fl修饰的DENV颗粒来调节，以诱导有效的NT抗体和最小的感染增强抗体。这代表了DENV疫苗设计的一种新策略，目前所有基于DENV颗粒的候选疫苗都无法实现。拟议的研究可以转化为几种更先进的DENV候选疫苗，作为第二代“安全有效”的DENV疫苗。

DESCRIPTION (provided by applicant): Despite considerable effort and progress in developing tetravalent vaccines against the four serotypes of dengue virus (DENV), one of the major unmet challenges is the difficulty in eliciting balanced neutralizing (NT) antibodies (Abs) against all four serotypes and to lower the risk of antibody-dependent enhancement (ADE), mediated mainly by cross-reactive and weakly or non-NT Abs. Studies of human Abs after DENV infection have shown the immunodominance of cross-reactive and weakly or non-NT Abs recognizing the fusion loop (FL) of envelope (E) protein over the type-specific and potent NT Abs, and the presence of cross-reactive and weakly or non-NT anti-precursor membrane (prM) Abs. Whether such immunodominance can be modulated to induce potent NT Abs without cross-reactive and weakly or non-NT Abs remains unknown. We recently found a DNA vaccine expressing mature DENV particles induced potent NT Abs with minimal cross-reactive, weakly or non-NT Abs, suggesting that specific modulation of DENV particles might induce superior Ab responses. Our long-term goal is to develop a safe and effective DENV vaccine. The objective of the proposed research is to understand the immunodominance of DENV E protein and to test whether mature or FL-modified mature DENV particles can induce potent NT Abs without cross-reactive, weakly or non-NT Abs. The central hypothesis is that mature DENV particles induce potent NT Abs, less anti-FL Abs and no anti-prM Abs, thus reducing the risk of ADE compared with mixed DENV particles. The objective will be achieved by the three specific aims: The first specific aim is to define the E protein epitopes and the accessibility, binding avidity and NT potency of a large panel of anti-E mAbs on mature, mixed and immature DENV particles. We hypothesize that differential epitope accessibility and binding avidity of anti-E Abs to different DENV particles may account for the immunodominance of E protein. The second aim is to demonstrate the superiority of DNA vaccines expressing mature DENV particles, over DNA vaccines expressing mixed particles, in eliciting potent NT Abs and minimal potential enhancing Abs and providing protection in outbred mice and AG129 mice, a well-established dengue murine model. The third aim is to demonstrate the superiority of DNA vaccines expressing FL-modified mature DENV particles, over DNA vaccines expressing non-modified mature DENV particles, in eliciting potent NT Abs and minimal potential enhancing Abs and providing protection in murine models. The significance of the proposed research rests on its detailed understanding of the immunodominance of DENV E protein and on the demonstration that such immunodominance can be modulated by mature and FL-modified DENV particles to induce potent NT Abs and minimal infection-enhancing Abs. This represents a novel strategy for DENV vaccine design and cannot be achieved by all current DENV particle-based candidate vaccines. The proposed research can be translated to several more advanced DENV vaccine candidates as the second generation of "safe and effective" DENV vaccines.