Leveraging cytoplasmic transcription to develop self-amplifying DNA vaccines

利用细胞质转录开发自我扩增 DNA 疫苗

• 批准号: 10579667
• 负责人: Jean M Peccoud
• 金额: $ 20.77万
• 依托单位: COLORADO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-07-07 至 2025-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10579667
• 关键词: 2019-nCoV; Address; Animals; Antigens; Architecture; Bacteriophage T7; Biological Assay; Biology; Biomanufacturing; COVID-19; COVID-19 pandemic; COVID-19 vaccine; Cell Culture Techniques; Cell Nucleus; Cell Survival; Cells; Clinical; Code; Cold Chains; Complex; Country; Cytoplasm; DNA; DNA Vaccines; DNA amplification; DNA cassette; DNA delivery; Data; Development; Emergency Situation; Emerging Communicable Diseases; Ensure; Enzymes; Event; Feedback; Flow Cytometry; Fluorescent in Situ Hybridization; Freezing; Frequencies; Future; Gene Amplification; Gene Expression; Generations; Genes; Genetic; Genetic Transcription; Genomics; Healthcare Systems; Hour; Human; Immune response; Incentives; Industry; Inferior; Infrastructure; Investigation; Learning; Life; Location; Measures; Messenger RNA; Methods; Nucleic Acid Vaccines; Plasmids; Process; Production; Program Development; Property; RNA; RNA amplification; RNA vaccine; Research Personnel; Safety; Speed; Structure; System; T7 RNA polymerase; Technology; Temperature; Testing; Time; Transcription Initiation; Transfection; Translations; Vaccine Design; Vaccines; Variant; Viral Genes; Virus; design; design and construction; design,build,test; dosage; experimental study; expression vector; feasibility testing; future epidemic; immunogenic; immunogenicity; improved; innovation; interest; mRNA capping; manufacture; manufacturing facility; manufacturing process; next generation; novel; novel vaccines; plasmid DNA; preservation; prevent; promoter; scaffold; single molecule; socioeconomics; success; synthetic biology; trend; vaccine access; vaccine candidate; vaccine development; vaccine platform; vector; vector vaccine

Project Summary The unprecedented speed of COVID-19 vaccine development has demonstrated the value of vaccine platforms. In particular, mRNA vaccines proved surprisingly successful at eliciting a strong immune response while having a remarkable safety profile considering the novelty of this system. Just as important are the remarkable speed and scale of their production. Despite their spectacular success, mRNA vaccines suffer from major limitations. mRNA is an inherently unstable molecule. One consequence of this property is that mRNA vaccines need to be stored in freezers and their shelf-life is measured in hours after they have been thawed. These storage requirements are considered difficult to ensure even in countries with developed healthcare systems and are extremely problematic in many other parts of the world. The second limitation of mRNA vaccines is that their production is unlike any other biomanufacturing process. As a result, it is limited by a critical lack of infrastructure and expertise. The COVID-19 mRNA vaccines provided an incentive to imagine the next generation of nucleic acid vaccines that would be easier to manufacture at scale and distribute to healthcare systems throughout the world. This proposal hypothesizes that a DNA-based vaccine could enable the design and deployment of safe and effective vaccines that would be faster, easier, and cheaper to manufacture at scale. The production of clinical- grade DNA relies on biomanufacturing processes that are some of the simplest, fastest, and most inexpensive processes in the industry. However, DNA vaccines have failed to elicit a protective immune response so far because only a small fraction of the DNA molecules entering a cell are transported to the nucleus where they can be transcribed. In this R21, researchers will test the feasibility of developing a new generation of DNA vaccines by applying methods from synthetic biology to introduce genetic circuits allowing the expression of the antigen to take place in the cytoplasm. Self-amplifying DNA vaccines will include several genes of viral origins that will transcribe the antigenic sequences from plasmids located in the cell cytoplasm. In addition, these vectors will include additional enzymes to modify mRNA molecules to increase their stability and translation efficiency. By introducing several levels of amplification, the expression of the antigen is expected to be several orders of magnitude higher than what can be achieved with traditional DNA vaccines. The project will proceed through eight iterations of the design-build-test-learn cycle to rationally improve vaccine designs using gene expression data in cell culture. If successful, future studies will test the platform compatibility with a broad range of antigens, optimize the delivery of DNA-based vaccines, and analyze the safety and efficacy of candidate vaccines in animal studies.

项目摘要 COVID-19疫苗开发的前所未有的速度证明了疫苗平台的价值。 特别是，事实证明，mRNA疫苗在引起强烈的免疫反应方面取得了惊人的成功 考虑到该系统的新颖性，一个显着的安全性。同样重要的速度也很重要 和他们的生产规模。尽管取得了惊人的成功，但mRNA疫苗仍受到重大局限性。 mRNA是一种固有的不稳定分子。该特性的结果之一是mRNA疫苗需要为 存放在冰柜中，其保质期在解冻后数小时在数小时内测量。这些存储 即使在具有发达医疗保健系统的国家，也很难确保要求 在世界许多其他地区都非常有问题。 mRNA疫苗的第二个局限性是它们 生产与任何其他生物制造过程不同。结果，它受到基础架构的严重缺乏的限制 和专业知识。 COVID-19 mRNA疫苗提供了一种想象下一代核的动力 酸性疫苗会更容易按比例制造并在整个整个整个医疗保健系统中分发 世界。 该提案假设基于DNA的疫苗可以使安全和部署 有效的疫苗将更快，更容易，更便宜地制造。临床的产生 级DNA依赖于最简单，最快，最便宜的生物制造过程 行业的过程。但是，DNA疫苗迄今未能引起保护性免疫反应 因为只有一小部分进入细胞的DNA分子被转运到核中 可以转录。 在此R21中，研究人员将通过应用来测试开发新一代DNA疫苗的可行性 合成生物学的方法引入遗传回路，允许进行抗原的表达 在细胞质中。自我放大的DNA疫苗将包括几种病毒起源的基因，这些基因将转录 位于细胞质中的质粒的抗原序列。此外，这些向量将包括其他 酶修饰mRNA分子以提高其稳定性和翻译效率。通过介绍几个 放大水平，抗原的表达预计将高几个数量级高于 传统的DNA疫苗可以实现的目标。该项目将进行八个迭代 设计建造测试的研究周期使用细胞培养中的基因表达数据合理地改善疫苗设计。 如果成功，未来的研究将测试平台与广泛抗原的兼容性，优化 递送基于DNA的疫苗，并分析候选疫苗在动物研究中的安全性和功效。