The potential of adoptive cell therapy with regulatory T cells (Tregs) to promote transplant tolerance is under active exploration. However, the impact of specific transplant settings and protocols on Treg manufacturing is not well-delineated. Here, we compared the use of peripheral blood mononuclear cells (PBMCs) from patients before or after liver transplantation to the use of healthy control PBMCs to determine their suitability for Treg manufacture using ex vivo costimulatory blockade with belatacept. Despite liver failure or immunosuppressive therapy, the capacity for Treg expansion during the manufacturing process was preserved. These experiments did not identify performance or quality issues that disqualified the use of posttransplant PBMCs—the currently favored protocol design. However, as Treg input correlated with output, significant CD4-lymphopenia in both pre- and posttransplant patients limited Treg yield. We therefore turned to leukapheresis posttransplant to improve absolute yield. To make deceased donor use feasible, we also developed protocols to substitute splenocytes for PBMCs as allostimulators. In addition to demonstrating that this Treg expansion strategy works in a liver transplant context, this preclinical study illustrates how characterizing cellular input populations and their performance can both inform and respond to clinical trial design and Treg manufacturing requirements.

中文翻译：

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使用共刺激阻断从肝移植受者体外产生调节性 T 细胞

正在积极探索使用调节性 T 细胞 (Treg) 促进移植耐受的过继细胞疗法的潜力。然而，特定移植设置和协议对 Treg 制造的影响尚未明确描述。在这里，我们将肝移植前后患者的外周血单核细胞 (PBMC) 的使用与健康对照 PBMC 的使用进行了比较，以确定它们是否适合使用贝拉西普离体共刺激阻断来制造 Treg。尽管进行了肝功能衰竭或免疫抑制治疗，但在制造过程中 Treg 扩增的能力得以保留。这些实验没有发现不符合移植后 PBMC 使用资格的性能或质量问题——目前最受青睐的方案设计。然而，由于 Treg 输入与输出相关，移植前和移植后患者显着的 CD4 淋巴细胞减少限制了 Treg 产量。因此，我们转向移植后白细胞分离术以提高绝对产量。为了使已故供体的使用变得可行，我们还开发了用脾细胞代替 PBMC 作为同种异体刺激剂的方案。除了证明这种 Treg 扩增策略在肝移植环境中有效之外，这项临床前研究还说明了表征细胞输入群体及其性能如何为临床试验设计和 Treg 制造要求提供信息并做出响应。