Automated Patient-Specific Dendritic Cell Generation for Transcriptomics-Driven Vaccinology

用于转录组驱动的疫苗学的自动患者特异性树突状细胞生成

• 批准号: 9093709
• 负责人: Shashi Murthy
• 金额: $ 39.23万
• 依托单位: NORTHEASTERN UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-06-20 至 2019-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9093709
• 关键词: Address; Arts; Autoimmunity; Autologous; Autologous Dendritic Cells; Basic Science; Benchmarking; Biological Assay; Biomedical Engineering; Biomedical Technology; Blood; Blood Cells; Blood Volume; Blood specimen; CD14 gene; CD4 Positive T Lymphocytes; Calmette-Guerin Bacillus; Cell Culture System; Cell Therapy; Cells; Clinical Research; Coculture Techniques; Communicable Diseases; Culture Media; Data; Dendritic Cell Vaccine; Dendritic Cells; Development; Dinoprostone; Disease; Elements; Environment; Epitopes; Flow Cytometry; Generations; Goals; Graft Rejection; Granulocyte-Macrophage Colony-Stimulating Factor; Health; Homing; Hour; Human; Human Resources; IL4 gene; IL6 gene; Immune; Immunity; Immunology; Individual; Infection; Interleukin-1; Knowledge; Lymphocyte; Malignant Neoplasms; Manuals; Memory; Metabolic Pathway; Methods; Microfluidics; Patients; Pattern; Performance; Perfusion; Peripheral Blood Mononuclear Cell; Phase; Phenotype; Prevention; Process; Protocols documentation; Regenerative Medicine; Regimen; Reproducibility; Research; Resources; Role; Sampling; Savings; Series; Standardization; System; T cell response; T cell therapy; T memory cell; T-Lymphocyte; T-Lymphocyte Subsets; TNF gene; Techniques; Technology; Testing; Time; Tube; Vaccination; Vaccine Clinical Trial; Vaccines; Whole Blood; blood perfusion; cohort; commercialization; cost; cost effective; cytokine; design; drug discovery; experience; genome-wide; head-to-head comparison; improved; individual patient; infectious disease treatment; insight; memory CD4 T lymphocyte; monocyte; new technology; novel vaccines; pathogen; programs; prototype; research study; response; self-renewal; tool; transcriptomics; vaccine candidate; vaccine development; vaccine efficacy; vaccine trial; vaccinology; whole genome

 DESCRIPTION (provided by applicant): Dendritic cells (DCs) are an indispensable part of studying human responses that are important for protective immunity against cancer and infectious diseases as well as prevention of autoimmunity and transplant rejection. These cells are also key elements of personalized vaccines which are a major research focus in cancer and infectious diseases. Despite the vital role of DCs in both clinical and basic research contexts, methods for obtaining these cells from individuals remains a comparatively under-developed and inefficient process. Because DCs are present in very low concentrations (<1%) in blood, these cells must be generated from monocytes and the state of the art in such generation involves a laborious process of static culture and stimulation with cytokines contained in culture medium. Numerous manual steps are required to go from a sample of patient-derived blood or peripheral blood mononuclear cells (PBMCs) to sufficient numbers of DCs that can be utilized for vaccine development, T cell therapy, or mechanistic studies. When scaled even to the level of tens of samples for a study involving one or two conditions or separate blood draws, the resource requirement in terms of personnel hours and number of manual steps becomes significant. Considering the existing and projected use of these cells at much larger scale, such as in Phase II or III clinical trials of vaccines and personalized cell therapy regimens, the curret approach to DC generation poses an unusually large burden, most significantly in terms of cost, but also in terms of the time required to perform comprehensive studies and trials. This proposal aims to address the unmet need for effective DC generation technologies by designing of a fully-automated microfluidic system (microDEN) that accepts a blood or PBMC sample and directly delivers DCs following a period of perfusion with cytokines. This system will combine monocyte isolation from blood and perfusion culture into individual, patient-specific chips. The elimination of manual steps associated with monocyte purification and culturing in two different media types containing cytokines will, in itself, represent a major advance relative to start of th art DC generation. We further hypothesize that the perfusion technique employed in our microfluidic method will allow reduction in the time required for DC generation (currently ~ 6 days), thereby offering significant additional savings in cost and resources. The proposed microDEN system will be built and rigorously benchmarked against the conventional DC generation technique using a range of functional assays, the most critical of which will be transcriptomal profiling of CD4+ T cells stimulated with autologous, BCG-infected DCs.

 描述（由申请人提供）：树突状细胞（DC）是研究人类反应不可或缺的一部分，这对于针对癌症和传染病的保护性免疫以及预防自身免疫和移植排斥非常重要。这些细胞也是个性化疫苗的关键要素，而个性化疫苗是癌症和传染病的主要研究重点。尽管树突状细胞在临床和基础研究中都发挥着重要作用，但从个体中获取这些细胞的方法仍然相对不发达且效率低下。由于 DC 在血液中的浓度非常低 (<1%)，因此这些细胞必须由单核细胞产生，而这种产生的现有技术涉及静态培养和用培养基中所含细胞因子刺激的费力过程。从患者血液或外周血单核细胞 (PBMC) 样本到足够数量的可用于疫苗开发、T 细胞治疗或机制研究的 DC 需要许多手动步骤。当涉及一种或两种情况或单独抽血的研究扩展到数十个样本的水平时，人员时间和手动步骤数量方面的资源需求变得非常大。考虑到这些细胞的现有和预计的更大规模的使用，例如在疫苗和个性化细胞治疗方案的 II 期或 III 期临床试验中，当前的 DC 生成方法造成了异常巨大的负担，最重要的是在成本方面，而且在进行全面研究和试验所需的时间方面。该提案旨在通过设计全自动微流体系统 (microDEN) 来解决对有效 DC 生成技术的未满足需求，该系统接受血液或 PBMC 样本，并在细胞因子灌注一段时间后直接输送 DC。该系统将从血液中分离的单核细胞和灌注培养物结合到个体的、患者特异性的芯片中。与单核细胞纯化和在含有细胞因子的两种不同培养基类型中培养相关的手动步骤的消除本身将代表相对于现有DC生成的开始的重大进步。我们进一步假设，我们的微流体方法中采用的灌注技术将减少 DC 生成所需的时间（目前约为 6 天），从而显着节省成本和资源。所提出的 microDEN 系统将使用一系列功能测定来构建并与传统 DC 生成技术进行严格的基准测试，其中最关键的是用自体、BCG 感染的 DC 刺激的 CD4+ T 细胞的转录组分析。