Transcriptional program of ex vivo expanded T cells

离体扩增T细胞的转录程序

• 批准号: 6690368
• 负责人: Eleftherios T Papoutsakis
• 金额: $ 36.41万
• 依托单位: NORTHWESTERN UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-01-10 至 2005-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6690368
• 关键词: Transcriptional; program; ex; vivo; expanded

DESCRIPTION (provided by applicant): Cellular immunotherapies with ex vivo expanded T cells (with or without genetic modifications) are promising tools for the treatment of otherwise untreatable cancers and viral infections, including iatrogenic ones (such as Epstein-Barr virus or Cytomegalovirus infections). In all cases, immune cells are isolated from the patient or an allogeneic donor, genetically modified if necessary, and expanded ex vivo before a large number of cells (10e9 -10e11) are infused into the patient. Thus, optimization of culture parameters with respect to cell proliferation and function is critical for successful immunotherapy. The majority of current protocols culture T cells in 5% C02 and ambient air (~20% 02 tension, p02). However, in vivo the mean pO2 in hematopoietic and lymphoid-organ tissues is closer to 40 mmHg (or 5% of the 02 in the gas atmosphere). We have shown that pO2 affects T-cell proliferation (by 4.8 to 13-fold), receptor expression and density, apoptosis, and metabolic rates. Similarly, we found that autologous plasma has a profound effect (43 to 74-fold over non-plasma cultures) on total cellular expansion. However, little is known about the underlying molecular biology of these profound effects, and specifically, about the large-scale gene expression patterns (transcriptome) during the activation and expansion process of T cells ex vivo. Thus, we propose to examine the large-scale transcriptome of ex vivo expanded human primary T cells using DNA arrays, whereby up to 8,500 genes will be examined for differential expression. Both T-cell selected and unselected peripheral blood mononuclear cells (PBMC) will be used to examine: (1) the temporal differential transcriptional program of T cells cultured under 5% or 20% pO2, and with or without autologous plasma; (2) the "relative" differential transcriptional program of cells cultured under 5% versus 20% pO2, and with versus without autologous plasma; (3) the differential transcriptional program of T cells cultured in the presence versus absence of an antioxidant; and (4) the differential transcriptional program of selected T cells vs. unselected cells under both low and high pO2. Analysis of DNA-array data will be used to test the following hypotheses as to whether these effects result from large transcriptional effects on: (i) The cell-cycle machinery; (ii) the apoptosis machinery; (iii) signal transduction/transcription factor networks; (iv) glycolysis/TCA cycle; and (v) stress and oxidative damage responses. In order to achieve these objectives, we will purse advanced clustering methods and new bioinformatic approaches that may allow us to examine if the collected DNA array expression data are consistent with existing pathway and regulatory networks models.

描述（由申请人提供）：离体细胞免疫疗法 扩增的 T 细胞（有或没有基因修饰）是有前途的工具 用于治疗其他无法治疗的癌症和病毒感染， 包括医源性病毒（例如 Epstein-Barr 病毒或巨细胞病毒） 感染）。在所有情况下，免疫细胞都是从患者或 同种异体供体，必要时进行基因改造，并进行离体扩增 在将大量细胞（10e9 -10e11）输注到患者体内之前。 因此，优化细胞增殖和培养参数 功能对于成功的免疫治疗至关重要。目前大多数 方案在5% CO 2 和环境空气（~20% O 2 张力，pO 2 ）中培养T细胞。 然而，体内造血和淋巴器官组织中的平均pO 2 接近40mmHg（或气氛中O 2 的5%）。我们已经证明 pO2 影响 T 细胞增殖（4.8 至 13 倍）、受体表达和 密度、细胞凋亡和代谢率。同样，我们发现自体 血浆对总培养物具有深远的影响（是非血浆培养物的 43 至 74 倍） 细胞扩张。然而，人们对潜在的分子机制知之甚少。 这些深远影响的生物学，特别是关于大规模基因 激活和扩展过程中的表达模式（转录组） T 细胞离体。因此，我们建议检查大规模转录组 使用 DNA 阵列离体扩增人类原代 T 细胞，其中多达 8,500 个 将检查基因的差异表达。 T 细胞都被选择并且 未选择的外周血单核细胞 (PBMC) 将用于检查： (1)培养下T细胞的时间差异转录程序 5% 或 20% pO2，有或没有自体血浆； (2)“亲戚” 在 5% 与 20% pO2 下培养的细胞的差异转录程序， 以及使用与不使用自体血浆的比较； (3)差异转录 在存在抗氧化剂和不存在抗氧化剂的情况下培养的 T 细胞程序； (4) 选定 T 细胞与非 T 细胞的差异转录程序。 在低和高 pO2 下未选择的细胞。 DNA 阵列数据分析将 用于检验以下假设是否会产生这些影响 来自对以下方面的大转录效应：（i）细胞周期机制； (二) 细胞凋亡机制； (iii) 信号转导/转录因子网络； (iv)糖酵解/TCA循环； (v) 应激和氧化损伤反应。在 为了实现这些目标，我们将寻求先进的聚类方法 新的生物信息学方法可以让我们检查收集到的数据是否 DNA 阵列表达数据与现有途径和监管一致 网络模型。

项目成果

Comparative analysis of transcriptional profiling of CD3+, CD4+ and CD8+ T cells identifies novel immune response players in T-cell activation.

• DOI: 10.1186/1471-2164-9-225
• 发表时间: 2008-05-16
• 期刊: BMC GENOMICS
• 影响因子: 4.4
• 作者: Wang, Min;Windgassen, Dirk;Papoutsakis, Eleftherios T
• 通讯作者: Papoutsakis, Eleftherios T

A model-based optimization framework for the inference of regulatory interactions using time-course DNA microarray expression data.

• DOI: 10.1186/1471-2105-8-228
• 发表时间: 2007-06-29
• 期刊: BMC BIOINFORMATICS
• 影响因子: 3
• 作者: Thomas, Reuben;Paredes, Carlos J;Mehrotra, Sanjay;Hatzimanikatis, Vassily;Papoutsakis, Eleftherios T
• 通讯作者: Papoutsakis, Eleftherios T