Division of Molecular Therapy, Advanced Clinical Research Center, The Institute of Medical Science, University of Tokyo, Tokyo, Japan.
AIDS Research Center, National Institute of Infectious Diseases, Tokyo, Japan.
Cytotherapy. 2018 Sep;20(9):1182-1190. doi: 10.1016/j.jcyt.2018.05.009. Epub 2018 Aug 17.
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.
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.
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.
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.
通过病毒特异性 T 细胞（VST）恢复病毒特异性免疫为传统药物提供了一种有吸引力的替代方案，并且在包括造血干细胞移植（HSCT）受者在内的免疫功能低下的患者中可能非常有效。然而，常规 VST 的制造需要制备专门的抗原呈递细胞（APC），在含血清的培养基中进行长时间的体外培养，并使用病毒或病毒载体重新刺激以提供 APC 上呈现的病毒抗原。
为了简化这个复杂的过程，我们开发了一种通过在无血清培养基中直接用重叠肽文库刺激外周血单核细胞（PBMC）来产生多种 VST 的方法。
我们生成了针对七种病毒（巨细胞病毒[CMV]、EB 病毒[EBV]、腺病毒[AdV]、人类疱疹病毒 6[HHV-6]、BK 病毒[BKV]、JC 病毒[JCV]和水痘带状疱疹病毒[VZV]）的 VST。无血清培养基中产生的 VST 的表型、生长和特异性与在常规含血清培养基中产生的 VST 相当。
无血清培养基的使用允许以更低的成本、更高的重现性（由于血清批次间变异性的消除）并且无需担心血清中存在的传染性病原体，将这种方法很容易引入临床实践。这种简化的方法现在将在 HLA 匹配的同胞 HSCT 受者中进行测试。
