细胞因子和刺激条件影响靶向肿瘤GD2的三代嵌合抗原受体修饰的T细胞的增殖以及免疫表型

技术资料下载地址: http://www.ncbi.nlm.nih.gov/pubmed/25573334
文献来源: Cytotherapy.
Different cytokine and stimulation conditions influence the expansion and immune phenotype of third-generation chimeric antigen receptor T cells specific for tumor antigen GD2.

BACKGROUND AIMS:
Chimeric antigen receptor (CAR) T cells are a novel immunotherapy for cancer. To achieve anti-tumor efficacy, these cells must survive, expand, and persist after infusion into patients, functions that are reportedly best achieved by cells with a stem or central-memory rather than effector-memory phenotype. We have developed third-generation CAR T cells specific for the tumor-associated antigen GD2 for use in a phase I clinical trial. We investigated the optimal cell culture conditions for CAR T-cell production, and here we describe the relative effects of 3 activation and cytokine conditions on CAR T-cell expansion, effector function and phenotype.
METHODS:
Peripheral blood mononuclear cells were activated by anti-CD3 and anti-CD28 or anti-CD3 and Retronectin. Activated cells were transduced with the CAR-encoding retroviral vector and expanded in either interleukin (IL)-2 or IL-7 and IL-15. Immune phenotype and expansion were tracked throughout the culture, and transduction efficiency, and subsequent GD2-specific effector functions were evaluated by flow cytometry and cytotoxic T lymphocytes assay.
RESULTS:
CD3/Retronectin stimulation with IL-2 resulted in poorer activation, expansion and Th1 cytokine secretion of CAR T cells than CD3/CD28 stimulation with either IL-2 or IL-7 and IL-15. However, CAR T cells cultured in CD3/CD28/IL7/IL-15 and CD3/Retronectin/IL-2 had superior cytotoxic T lymphocyte activity and a more stem-like phenotype.
DISCUSSION:
The combination of CD3 and CD28 with IL-7 and IL-15 gave the best balance of CAR T-cell expansion and potent GD2-specific effector functions while retaining a stem/memory phenotype, and these growth conditions will therefore be used to manufacture CAR T cells for our phase I clinical trial.

不同的细胞因子和刺激条件影响靶向肿瘤GD2的三代嵌合抗原受体修饰的T细胞的增殖以及免疫表型
背景目标
CAR-T疗法是一种对抗癌症的新型免疫治疗方案。为了实现抗肿瘤效应，这些细胞必须存活、然后扩增并且注射入病人体内后需要持续存活。我们已经发展了一种三代CAR-T细胞来对抗GD2阳性肿瘤细胞，并运用在1期临床试验中。我们研究了最优的CAR-T培养的环境，在这里，我们描述了活化和细胞因子对CAR-T细胞扩增、效应功能和表型的影响。
方法：
通过靶向CD3和靶向CD28或者靶向CD3和Retronectin来活化外周血单个核细胞。活化的细胞利用CAR编码的逆转录病毒载体进行转导，并在白细胞介素(IL)-2 或白细胞介素-7 和白细胞介素-15作用下扩增。免疫表型和扩增通过培养来跟踪、转导效率和GD-2特异性效应功能通过流式细胞仪和细胞毒性T淋巴细胞测定。
结果：
CD3/Retronectin和IL-2刺激CAR-T，与利用IL-2 或者 IL-7 和 IL-15.刺激CD3/CD28 CAR-T细胞相比，会有较差的活性、扩增和Th1细胞因子分泌。然而，在CD3/CD28/IL7/IL-15和CD3/Retronectin/IL-2中培养的CAR-T会有一个更好的细胞毒性活性和干细胞表型。
讨论：
运用 IL-7、IL-15和CD3、 CD28的组合给CAR-T细胞的扩增和GD2靶向抗肿瘤能力提供最好的平衡，并且维持干细胞/记忆细胞的表型，因此这种生长条件可以用于我们1期临床试验的CAR-T细胞培养。

出自爱康得生物技术