Generation of multivirus-specific T cells by a single stimulation of peripheral blood mononuclear cells with a peptide mixture using serum-free medium
Generation of multivirus-specific T cells by a single stimulation of peripheral blood mononuclear cells with a peptide mixture using serum-free medium
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DOI：
10.1016/j.jcyt.2018.05.009
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发表时间：
2018-09-01
期刊：
影响因子：
4.5
通讯作者：
Takahashi, Satoshi
中科院分区：
文献类型：
--
作者：
Nishiyama-Fujita, Yuriko;Kawana-Tachikawa, Ai;Takahashi, Satoshi
背景：通过病毒特异性T细胞（VSTs）恢复病毒特异性免疫提供了一种有吸引力的替代传统药物，并且对免疫功能低下的患者（包括造血干细胞移植（HSCT）接受者）非常有效。然而，传统的VSTs制造需要制备专门的抗原提呈细胞（APCs），在含血清的培养基中长时间体外培养，用病毒或病毒载体再刺激抗原，以提供抗原在APCs上提呈。方法：为了简化这一复杂的过程，我们开发了一种在无血清培养基中直接刺激具有重叠肽库的外周血单个核细胞（PBMCs）产生多个vst的方法。结果：我们在一个细胞系中生成了针对巨细胞病毒（CMV）、eb病毒（EBV）、腺病毒（AdV）、人疱疹病毒6 （HHV-6）、BK病毒（BKV）、JC病毒（JCV）和水痘带状疱疹病毒（VZV） 7种病毒的VSTs。无血清培养基中产生的多个VSTs的表型、生长和特异性与常规含血清培养基中产生的VSTs相当。讨论：无血清培养基的使用使该方法易于引入临床实践，成本更低，由于血清中不存在批次间的可变性，因此可重复性更高，并且无需担心所用血清中的感染性原。这种简化的方法现在将在人类白细胞抗原（HLA）匹配的同胞HSCT受体中进行测试。
Background: Restoration of virus-specific immunity by virus specific T cells (VSTs) offers an attractive alternative to conventional drugs, and can be highly effective in immunocompromised patients, including hematopoietic stem cell transplant (HSCT) recipients. However, conventional VSTs manufacture requires preparation of specialized antigen-presenting cells (APCs), prolonged ex vivo culture in serum-containing medium and antigen re-stimulation with viruses or viral vectors to provide viral antigens for presentation on APCs. Methods: To simplify this complex process, we developed a method to generate multiple VSTs by direct stimulation of peripheral blood mononuclear cells (PBMCs) with overlapping peptide libraries in serum-free medium. Results: We generated VSTs that targeted seven viruses (cytomegalovirus [CMV], Epstein-Barr virus [EBV], adenovirus [AdV], human herpesvirus 6 [HHV-6], BK virus [BKV], JC virus [JCV] and Varicella Zoster virus [VZV]) in a single line. The phenotype, growth and specificity of multiple VSTs produced in serum-free medium were equivalent to those generated in conventional serum-containing medium. Discussion: The use of serum-free medium allows this approach to be readily introduced to clinical practice with lower cost, greater reproducibility due to the absence of batch-to batch variability in serum and without concerns for infectious agents in the serum used. This simplified approach will now be tested in recipients of Human Leukocyte Antigen (HLA) matched sibling HSCT.