Development of a novel and broadly applicable thermostable bacteriophage VLPs platforms for vaccine design, drug delivery, and imaging

项目总结 病毒样颗粒(VLP)在大小、结构和免疫原性上类似于病毒，病毒的外壳或 包膜蛋白(S)的来源除了它们缺乏病毒基因组这一事实外；VLP是非传染性的和 是安全的。这些特征已被利用来开发基于VLP的人乳头瘤病毒疫苗 和乙肝病毒；此外，目前正在探索来自~70种病毒的外壳蛋白，以开发VLP- 基于这些病毒的疫苗。其中一些病毒的VLP也被用作展示 开发嵌合VLP的平台，展示来自其他感染性病原体、肿瘤- 相关抗原和其他代谢性疾病。这些嵌合的VLP的目标是诱导抗体 对抗展示在平台上的异源抗原，而不是平台。除了担任 除了展示平台外，VLP还被用于将药物或货物定向输送到特定的癌细胞。 而显示异源多肽的候选VLP疫苗在动物身上非常有效 研究，在大多数研究中，VLP平台(包括腺病毒VLP或十二面体)是衍生出来的 从感染人类的病毒，在某些情况下，研究使用了以前 用于为普通人群接种疫苗；一个很好的例子是乙肝疫苗，在全球范围内为婴儿接种疫苗 2015年的覆盖率为84%。基于其中一些平台的疫苗，其中包括预先存在的抗体 在一般人群中，对人类的免疫原性可能较低。此外，对大小有限制 可以在某些VLP平台上以基因方式显示的异源抗原，这使得它具有挑战性 在相同的VLP上显示具有多个表位的单肽。此外，大部分VLP平台都是 它们对温度敏感，不太适合制冷设施较差的发展中国家。 在这项提案中，PI将开发和表征新型耐热噬菌体VLP平台，使用 嗜热病毒P23-77和ΦIN93的外壳蛋白。P23-77和ΦIN93是从 在70-75°C下生长。因此，来自这些病毒的VLP在室温(RT)或 高于RT。这些VLP平台的另一个好处是，因为这些病毒不会感染人类，所以 人类缺乏针对VLP平台的预先存在的中和抗体；因此， 这些平台的免疫原性不会受到预先存在的抗体的影响。此外，许多表面上- 衣壳上暴露的环可以耐受更大的异源抗原插入。公安部将联合表达 三个来自P23-77的外壳蛋白和两个来自ΦIN93的外壳蛋白。私家侦探将评估大衣是否 蛋白质可以组装成VLP，如果它们VLP是耐热的，可以耐受异源多肽插入 来自人类乳头瘤病毒的病毒，以及与病毒相比VLP是否具有免疫原性。

PROJECT SUMMARY Virus-like particles (VLPs) resemble - in size, structure, and immunogenicity - the virus from which the coat or envelope protein(s) are derived from except for the fact they lack a viral genome; VLPs are non-infectious and are safe. These features have been exploited to develop VLP-based vaccines against human papillomaviruses and hepatitis B virus; furthermore, coat proteins from ~70 viruses are currently being explored to develop VLP- based vaccines against these viruses. VLPs from some of these viruses have also been used as display platforms to develop chimeric VLPs displaying heterologous peptides from other infectious agents, tumor- associated antigens and other metabolic diseases. The goal of these chimeric VLPs is to induce antibodies against the heterologous antigen displayed on the platforms and not the platforms. In addition to serving as display platforms, VLPs have also been used for targeted delivery of drugs or cargo to specific cancer cells. While the candidate VLPs-based vaccines displaying heterologous peptides are very effective in animal studies, in the majority of studies, VLPs platforms (including adenoviral VLPs or dodecahedron) are derived from viruses that infect humans and in some cases, studies used VLPs platforms that had previously been used to immunize the general population; a good example is HBV vaccine, with a global infant vaccination coverage of 84% in 2015. Vaccines based on some of these platforms, with pre-existing antibodies in the general population, are likely to be less immunogenic in humans. Additionally, there is a limitation on the size of heterologous antigens that can be genetically displayed on some VLPs platforms making it challenging to display a single peptide with multiple epitopes on the same VLPs. Moreover, most of the VLPs platforms are temperature-sensitive making them less suitable in developing countries with poor refrigeration facilities. In this proposal, the PI will develop and characterize novel thermostable bacteriophage VLPs platforms using coat proteins from thermophilic viruses P23-77 and ΦIN93. P23-77 and ΦIN93 was isolated from bacteria that grow at 70-75 °C. Thus VLPs derived from these viruses are likely to be stable at room temperature (RT) or above RT. Additional benefit of these VLPs platforms is that because these viruses do not infect humans, the human population lacks pre-existing neutralizing antibodies against the VLPs platforms; thus, the immunogenicity of the platforms cannot be compromised by pre-existing antibodies. Also, many surface- exposed loops on the capsid may tolerate larger insertions of heterologous antigens. The PI will co-express three coat proteins from P23-77 and two coat proteins from ΦIN93. The PI will assess whether the coat proteins can assemble into VLPs, if they VLPs are thermostable, can tolerate heterologous peptide insertions from human papillomaviruses, and if VLPs are immunogenic in comparison to the virus(es).