背景:2019年冠状病毒病(COVID-19)是由严重急性呼吸道综合征冠状病毒2(SARS-CoV-2)引起的。SARS-CoV-2病毒特异性细胞毒性T细胞淋巴细胞(vCTL)可以为COVID-19治疗提供有希望的方式。我们的目标是筛选,制造,并使用CliniMACS®细胞因子捕获系统(CCS)表征从恢复期COVID-19供体产生的SARS-CoV-2-vCTL。
方法:通过用病毒肽刺激恢复期COVID-19供者外周血单个核细胞,并使用流式细胞术鉴定IFN-γ+CD4和CD8T细胞进行供者筛选。使用CliniMACS®CCS制造临床级SARS-CoV-2-vCTL。富集的SARS-CoV-2-vCTLs通过T细胞受体测序来表征,质量细胞计数,和转录组分析。
结果:93%的康复期献血者血液样本通过了临床生产的筛选标准。三次验证运行产生富集的T细胞,其为79%±21%IFN-γ+T细胞。SARS-CoV-2-vCTL表现出高度多样化的TCR库,增强了记忆CD8和CD4T细胞,特别是在CD8TEM中,CD4TCM和CD4TEMRA细胞亚群。SARS-CoV-2-vCTLs是多功能的,在T细胞功能中基因表达增加,白细胞介素,病原体防御,和肿瘤坏死因子超家族通路。
结论:高功能SARS-CoV-2-vCTLs可以通过直接细胞因子富集(12小时)从康复供体中快速产生。
UNASSIGNED:NCT04896606、NCT03266627、NCT03266640、NCT03266653、NCT04197596。
Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). SARS-CoV-2 virus-specific cytotoxic T-cell lymphocytes (vCTLs) could provide a promising modality in COVID-19 treatment. We aimed to screen, manufacture, and characterize SARS-CoV-2-vCTLs generated from convalescent COVID-19 donors using the CliniMACS Cytokine Capture System (CCS).
Donor screening was done by stimulation of convalescent COVID-19 donor peripheral blood mononuclear cells with viral peptides and identification of interferonγ (IFN-γ)+ CD4 and CD8 T cells using flow cytometry. Clinical-grade SARS-CoV-2-vCTLs were manufactured using the CliniMACS CCS. The enriched SARS-CoV-2-vCTLs were characterized by T-cell receptor sequencing, mass cytometry, and transcriptome analysis.
Of the convalescent donor blood samples, 93% passed the screening criteria for clinical manufacture. Three validation runs resulted in enriched T cells that were 79% (standard error of the mean 21%) IFN-γ+ T cells. SARS-CoV-2-vCTLs displayed a highly diverse T-cell receptor repertoire with enhancement of both memory CD8 and CD4 T cells, especially in CD8 TEM, CD4 TCM, and CD4 TEMRA cell subsets. SARS-CoV-2-vCTLs were polyfunctional with increased gene expression in T-cell function, interleukin, pathogen defense, and tumor necrosis factor superfamily pathways.
Highly functional SARS-CoV-2-vCTLs can be rapidly generated by direct cytokine enrichment (12 hours) from convalescent donors.
NCT04896606.