Thammanichanond Duangtawan, Moneer Sarah, Yotnda Patricia, Aitken Campbell, Earnest-Silveira Linda, Jackson David, Hellard Margaret, McCluskey James, Torresi Joseph, Bharadwaj Mandvi
Department of Microbiology and Immunology, University of Melbourne, Parkville, Victoria 3010, Australia.
Clin Immunol. 2008 Sep;128(3):329-39. doi: 10.1016/j.clim.2008.04.002. Epub 2008 Jun 3.
Hepatitis C virus (HCV)-specific cytotoxic T lymphocytes (CTLs) play an important role in HCV clearance. The frequency of HCV-specific T(CD8) in peripheral blood of HCV-infected donors is very low and HCV cannot be cultivated for reinfection of antigen presenting cells, making it difficult to detect T(CD8) of broad HCV specificities from peripheral blood mononuclear cells (PBMCs). We have developed a recombinant adenoviral system that efficiently reactivates and expands HCV-specific CTLs from PBMCs of HCV-infected donors. Replication-incompetent adenoviruses expressing individual HCV proteins (core and NS3) were produced and PBMCs from HCV-infected donors were transduced with these recombinant adeno-HCV constructs to stimulate HCV-specific CTL populations. T cells expanded from adeno-HCV stimulated cultures were potent producers of HCV-specific IFN-gamma and TNF-alpha and efficiently lysed target cells pulsed with HCV peptides. These constructs could stimulate T(CD8) directed towards multiple HCV peptides while preserving the determinant hierarchy. This approach therefore overcomes some of the shortcomings of the selective expansion of CTLs with peptide-based vaccine strategies. These findings provide an effective approach for the expansion of HCV-specific CTLs from PBMCs of HCV-infected patients and have potential for immunotherapeutic/vaccine development.
丙型肝炎病毒（HCV）特异性细胞毒性T淋巴细胞（CTL）在HCV清除中发挥重要作用。HCV感染供体外周血中HCV特异性T（CD8）细胞的频率非常低，且HCV无法培养以重新感染抗原呈递细胞，因此难以从外周血单核细胞（PBMC）中检测到具有广泛HCV特异性的T（CD8）细胞。我们开发了一种重组腺病毒系统，可有效激活并扩增来自HCV感染供体PBMC的HCV特异性CTL。制备了表达单个HCV蛋白（核心蛋白和NS3蛋白）的无复制能力腺病毒，并用这些重组腺病毒-HCV构建体转导HCV感染供体的PBMC，以刺激HCV特异性CTL群体。从腺病毒-HCV刺激培养物中扩增的T细胞是HCV特异性干扰素-γ和肿瘤坏死因子-α的有效产生者，并能有效裂解用HCV肽脉冲处理的靶细胞。这些构建体可刺激针对多种HCV肽的T（CD8）细胞，同时保留决定簇层次结构。因此，该方法克服了基于肽的疫苗策略在选择性扩增CTL方面的一些缺点。这些发现为从HCV感染患者的PBMC中扩增HCV特异性CTL提供了一种有效方法，并具有免疫治疗/疫苗开发的潜力。
