Nanoparticle Delivery of DNA Vaccine Against Zika Virus

纳米颗粒递送抗寨卡病毒 DNA 疫苗

• 批准号: 9379639
• 负责人: Hai-Quan Mao
• 金额: $ 20.43万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-05-19 至 2019-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9379639
• 关键词: Address; Alexa594; Amino Acid Sequence; Amino Acids; Analysis of Variance; Antibodies; Antibody Formation; Antibody titer measurement; Antigen Presentation; Antigen Presentation Pathway; Antigen-Presenting Cells; Antigens; Asia; B-Cell Activation; Benchmarking; Bone Marrow; Brazil; CD4 Positive T Lymphocytes; CD8-Positive T-Lymphocytes; CD8B1 gene; Cells; Centers for Disease Control and Prevention (U.S.); Chimera organism; Cytotoxic T-Lymphocytes; DNA; DNA Vaccines; DNA delivery; Dendritic Cells; Development; Dose; Drainage procedure; Electroporation; Engineering; Enzyme-Linked Immunosorbent Assay; Enzymes; Epidemic; Esters; Europe; Exhibits; Flow Cytometry; Frequencies; Gene Expression; Generations; Glycolates; Helper-Inducer T-Lymphocyte; Histocompatibility Antigens Class II; Immune response; Immunoglobulin G; Immunologic Memory; Infection; Injection of therapeutic agent; Intramuscular; Label; Laboratories; Life; Link; Lymph Node Drainage; Lymphocyte; MHC Class II Genes; Mediating; Medicine; Membrane; Membrane Proteins; Methods; Mus; Neutralization Tests; North America; Ovalbumin; Phosphate Buffer; Physical condensation; Polyethylene Glycols; Polyethyleneimine; Polymerase Chain Reaction; Polypropylenes; Process; Production; Protein Subunits; Race; Reporting; Reproducibility; Reverse Transcription; Saline; Serum; Site; South America; Subunit Vaccines; Sulfides; Surface Properties; T-Cell Activation; Testing; Th2 Cells; Time; Tissues; Transfection; Transmission Electron Microscopy; Vaccination; Vaccines; World Health Organization; Zika Virus; Zika virus vaccine; base; copolymer; cytokine; disorder prevention; env Gene Products; fetal bovine serum; fight against; innovation; light scattering; long term memory; lymph nodes; lymphatic drainage; nanoparticle; neutralizing antibody; next generation; nonhuman primate; novel strategies; pathogen; plasmid DNA; response; subcutaneous; transgene expression; uptake; vaccine candidate; vaccine delivery; whole body imaging

PROJECT SUMMARY The objective of this project is to develop DNA nanoparticles with efficient delivery to the local draining lymph nodes (dLN) via subcutaneous (s.c.) administration to promote dendritic cell (DC) transfection, antigen presentation, and T cell activation in the lymph nodes, and to elicit robust antibody titers and immunological memory against Zika virus. As the Zika virus (ZIKV) epidemic in Brazil spread around south and north Americas, Europe, and Asia, teams around the world have been racing to develop ZIKV vaccines. DNA vaccines offer many advantages in terms of ease of production, excellent stability with long shelf life, multivalent capability, and fast development cycle. The first generation of vaccine candidates is built with the pre-membrane and envelope sequences from a Brazilian ZIKV strain as the dominant immunogen. They have been tested as naked plasmid DNA vaccines administered by intramuscular (i.m.) injection, and generated effective neutralizing antibodies in mice and nonhuman primates. However, naked DNA vaccine inherently has low efficiency to generate Th2 response and immune memory, and requires higher DNA dose due to the low abundance of antigen presenting cells (APCs) at the injection site. This study will directly address these challenges by combining two novel approaches to engineer a more potent ZIKV DNA vaccine: (1) a chimeric DNA construct encoding ZIKA envelope protein sequences and lysosomal-associated membrane protein-1 (LAMP), which can direct the expressed antigen to MHC Class II-rich compartment, thus skewing ZIKV antigen presentation towards a strongly Th2-biased response and generation of immune memory; and (2) flash nanocomplexation (FNC)-produced small (~40 nm) ZIKV/LAMP DNA nanoparticles to enable drainage to the dLNs following s.c. injection, thus enhancing the immune response and allowing for substantial reduction in vaccination dose. We will first engineer the DNA nanoparticles with different sizes and narrow distribution using the FNC method, and characterize their stability, surface properties, and ability to transfect DCs; then determine the effect of NP size on LN-draining efficiency and gene expression in DCs in different dLNs; and lastly demonstrate the robust immune responses elicited by LN-targeting NPs with LAMP/ZIKV DNA vaccine, and characterize the subtypes of anti-ZIKV response. This timely study will not only develop a more effective ZIKV vaccine and support further testing of the next generation of Zika vaccine, but also provide a potent DNA vaccine platform against other emerging pathogens.

项目摘要 本项目的目标是开发DNA纳米颗粒，并将其有效地输送到局部引流淋巴 淋巴结（dLN）经皮下（s.c.）施用以促进树突状细胞（DC）转染，抗原 淋巴结中的T细胞活化，并引发稳健的抗体滴度和免疫学效应。 寨卡病毒是什么？随着寨卡病毒（ZIKV）在巴西的流行在南部和北部蔓延， 在美洲、欧洲和亚洲，世界各地的团队一直在竞相开发ZIKV疫苗。DNA 疫苗在易于生产，具有长保存期的优异稳定性， 多价能力和快速的开发周期。第一代候选疫苗是用 来自巴西ZIKV毒株的前膜和包膜序列作为优势免疫原。他们有 作为裸质粒DNA疫苗通过肌内（i.m.）注入，并生成 在小鼠和非人灵长类动物中有效的中和抗体。然而，裸DNA疫苗本身具有 低效率产生Th 2应答和免疫记忆，并且由于低效率需要更高的DNA剂量， 注射部位抗原呈递细胞（APC）的丰度。这项研究将直接解决这些问题。 通过组合两种新方法来工程化更有效的ZIKV DNA疫苗的挑战：（1）嵌合的 编码ZIKA包膜蛋白序列和溶酶体相关膜蛋白-1的DNA构建体 （LAMP），其可以将表达的抗原引导至富含MHC II类的区室，从而使ZIKV抗原 向强Th 2偏向性应答的呈递和免疫记忆的产生;和（2）闪光 纳米复合（FNC）-产生的小（~40 nm）ZIKV/LAMP DNA纳米颗粒，以使得能够引流到 皮下注射后dLN注射，从而增强免疫应答，并允许显著减少 接种剂量。我们将首先设计具有不同尺寸和窄分布的DNA纳米颗粒 使用FNC方法，并表征它们的稳定性、表面性质和抑制DC的能力;然后 确定NP大小对不同dLN中DC中LN排出效率和基因表达的影响;和 最后证明了由具有LAMP/ZIKV DNA疫苗的靶向LN的NP引起的稳健的免疫应答， 并表征抗ZIKV应答的亚型。这一及时的研究不仅将制定一个更有效的 ZIKV疫苗和支持下一代寨卡疫苗的进一步测试，也提供了一个有效的DNA 针对其他新兴病原体的疫苗平台。