Broad-Range VLP Vaccine Against H5N1 Influenza

描述（由申请人提供）：高致病性禽流感（HPAI）病毒是C类NIAID优先病原体，是全球生物防御威胁。提供针对高致病性禽流感病毒的保护的疫苗是生物防御和大流行准备所必需的。在过去的十年中，H5 N1高致病性禽流感病毒的基因和抗原性多样化，导致需要多种H5 N1疫苗。虽然已经制备了H5 N1进化枝1型疫苗，但它们不太可能对其他H5 N1进化枝产生保护作用。此外，准备 针对每种潜在威胁病毒的疫苗成本昂贵，在经济上不可行，并可能引起严重的生物安全和生物安保问题。重组病毒样颗粒（VLP）是预防高致病性禽流感的一种很有前途的策略。VLP在安全性、有效性和制造方面具有优势，因为它们避免了病毒生长缓慢、产量不可预测和宿主适应性突变等问题。已经表明，VLP比标准流感疫苗诱导针对不同毒株的更广泛的免疫，特别是如果粘膜施用（Bright et al.，2007; Perrone等人，2009年）。此外，我们最近制备了多亚型VLP，其中几种不同的血凝素（HA）亚型共定位在相同的VLP结构内，从而提供针对多种攻击病毒的保护（Pushko et al.，2011; Tretyakova等人，2013年）。多亚型疫苗具有诱导针对几种流感亚型的大范围但传统的病毒中和免疫应答的优势。 在此，我们提出使用多HA VLP平台来开发多H5 VLP作为独特的多进化枝H5 N1 VLP疫苗。多H5 VLP将被合理地设计以在VLP内共定位源自三个不同H5 N1进化枝的三种H5蛋白。预期所得三重H5 VLP引发针对多种H5 N1 HPAI病毒的有效保护，从而产生安全且有效的、广泛保护性的H5 N1疫苗。特别地，我们提出制备共定位来自进化枝1、进化枝2.2.1.1和进化枝2.3.2.1病毒的H5蛋白的VLP，所有这些都是WHO推荐用于H5 N1疫苗开发的。将使用重组杆状病毒表达制备Multi-H5 VLP，并评价VLP的遗传稳定性、生化、结构和抗原特性（Sp. Aim 1）。三重H5 VLP的免疫原性和功效将与疾病控制和预防中心（CDC）合作在实验性雪貂攻击模型中进行评估。为了评价这种新型广谱H5 N1疫苗，提出了同源（Sp. Aim 2）和异源（Sp. Aim 3）H5 N1 HPAI挑战。年轻和年老的雪貂将被纳入挑战研究，以模拟最脆弱的人群。将评价肌内和鼻内接种途径对免疫应答的影响。广泛的免疫机制将被阐明，包括体液，细胞介导的，和粘膜免疫。将进行工艺开发和技术转移到制造环境中。完成临床前试验和工艺开发活动后，将计划与FDA召开IND前会议。如果成功，这项新技术可能代表一个创新平台，用于快速和成本效益高的制备多价流感疫苗，用于生物防御和大流行的准备。

DESCRIPTION (provided by applicant): Highly pathogenic avian influenza (HPAI) viruses are Category C NIAID priority pathogens and a global biodefense threat. Vaccines that provide protection against HPAI viruses are required for biodefense and pandemic preparedness. During the past decade, the H5N1 HPAI viruses have diversified genetically and antigenically leading to the need for multiple H5N1 vaccines. Although H5N1 clade 1 vaccines have been prepared, it is unlikely that they will protect against other H5N1 clades. In addition, preparation of vaccines for each potential threat virus is expensive, economically not feasible and can raise serious biosafety and biosecurity concerns. Recombinant virus-like particles (VLPs) represent a promising strategy for prevention of HPAI. VLPs have advantages in safety, efficacy, and manufacturing because they circumvent problems like slow virus growth, unpredictable yields, and host-adaptive mutations. It has been shown that VLPs induce broader immunity against divergent strains than standard influenza vaccines, especially if administered mucosally (Bright et al., 2007; Perrone et al., 2009). Furthermore, we have recently prepared multi-subtype VLP, in which several distinct subtypes of hemagglutinin (HA) were co-localized within the same VLP structure thus providing protection against multiple challenge viruses (Pushko et al., 2011; Tretyakova et al., 2013). The multi-subtype vaccines have advantage of inducing broad-range, yet traditional virus-neutralizing immune responses directed against several influenza subtypes. Here we propose the use of multi-HA VLP platform for the development of multi-H5 VLPs as a unique multi-clade H5N1 VLP vaccine. The multi-H5 VLPs will be rationally designed to co-localize within the VLP the three H5 proteins derived from three distinct H5N1 clades. The resulting triple-H5 VLPs are expected to elicit effective protection against multiple H5N1 HPAI viruses resulting in a safe and effective, broadly protective H5N1 vaccine. Particularly, we propose preparation of VLPs that co-localize H5 proteins derived from Clade 1, Clade 2.2.1.1, and Clade 2.3.2.1 viruses, all recommended by the WHO for H5N1 vaccine development. Multi-H5 VLPs will be prepared by using recombinant baculovirus expression, and genetic stability, biochemical, structural, and antigenic characteristics of VLPs will be evaluated (Sp. Aim 1). Immunogenicity and efficacy of triple-H5 VLPs will be evaluated in experimental ferret challenge model in collaboration with the Centers for Disease Control and Prevention (CDC). Both the homologous (Sp. Aim 2) and heterologous (Sp. Aim 3) H5N1 HPAI challenges are proposed in order to evaluate this novel broad-spectrum H5N1 vaccine. Young and aged ferrets will be included into the challenge studies to mimic the most vulnerable human populations. The effects of intramuscular and intranasal routes of vaccination on immune responses will be evaluated. Mechanisms of broad immunity will be elucidated including humoral, cell-mediated, and mucosal immunity. Process development and technology transfer into manufacturing environment will be carried out. Following completion of preclinical trials and process development activities, a pre-IND meeting with FDA will be planned. If successful, this novel technology may represent an innovative platform for rapid and cost-effective preparation of multivalent influenza vaccines for biodefense and pandemic preparedness.