Dmitry Rogachev National Medical Research Center of Pediatric Hematology, Oncology, and Immunology, Moscow, Russia.
Dmitry Rogachev National Medical Research Center of Pediatric Hematology, Oncology, and Immunology, Moscow, Russia.
Cytotherapy. 2024 Jun;26(6):567-578. doi: 10.1016/j.jcyt.2024.02.025. Epub 2024 Mar 2.
BACKGROUND AIMS: The CliniMACS Prodigy closed system is widely used for the manufacturing of chimeric antigen receptor T cells (CAR-T cells). Our study presents an extensive immunophenotypic and functional characterization and comparison of the properties of anti-CD19 CAR-T cell products obtained during long (11 days) and short (7 days) manufacturing cycles using the CliniMACS Prodigy system, as well as cell products manufactured from different donor sources of T lymphocytes: from patients, from patients who underwent HSCT, and from haploidentical donors. We also present the possibility of assessing the efficiency of transduction by an indirect method. METHODS: Seventy-six CD19 CAR-T cell products were manufactured using the CliniMACS Prodigy automated system. Immunophenotypic properties, markers of cell activation and exhaustion, antitumor, anti-CD19 specific activity in vitro of the manufactured cell products were evaluated. As an indirect method for assessing the efficiency of transduction, we used the method of functional assessment of cytokine secretion and expression of the CD107a marker after incubation of CAR-T cells with tumor targets. RESULTS: The CliniMACS Prodigy platform can produce a product of CD19 CAR-T cells with sufficient cell expansion (4.6 × 10 cells-median for long process [LP] and 1.6 × 10-for short process [SP]), transduction efficiency (43.5%-median for LP and 41.0%-for SP), represented mainly by T central memory cell population, with low expression of exhaustion markers, and with high specific antitumor activity in vitro. We did not find significant differences in the properties of the products obtained during the 7- and 11-day manufacturing cycles, which is in favor of reducing the duration of production to 7 days, which may accelerate CAR-T therapy. We have shown that donor sources for CAR-T manufacturing do not significantly affect the composition and functional properties of the cell product. CONCLUSIONS: This study demonstrates the possibility of using the CliniMACS Prodigy system with a shortened 7-day production cycle to produce sufficient amount of functional CAR-T cells. CAR transduction efficiency can be measured indirectly via functional assays.
背景目的：CliniMACS Prodigy 封闭式系统广泛用于嵌合抗原受体 T 细胞（CAR-T 细胞）的生产。我们的研究对使用 CliniMACS Prodigy 系统在较长（11 天）和较短（7 天）制造周期中获得的抗 CD19 CAR-T 细胞产品的特性进行了广泛的免疫表型和功能表征和比较，以及由不同供体来源的 T 淋巴细胞制造的细胞产品：来自患者、接受 HSCT 的患者和单倍体供体。我们还提出了通过间接方法评估转导效率的可能性。 方法：使用 CliniMACS Prodigy 自动化系统制造了 76 种 CD19 CAR-T 细胞产品。评估了制造的细胞产品的免疫表型特性、细胞激活和衰竭标志物、抗肿瘤、体外抗 CD19 特异性活性。作为评估转导效率的间接方法，我们使用了在 CAR-T 细胞与肿瘤靶标孵育后评估细胞因子分泌和 CD107a 标志物表达的功能评估方法。 结果：CliniMACS Prodigy 平台能够生产具有足够细胞扩增（长过程[LP]中位数为 4.6×10 细胞，短过程[SP]中位数为 1.6×10）、转导效率（LP 中位数为 43.5%，SP 中位数为 41.0%）的 CD19 CAR-T 细胞产品，主要由 T 中央记忆细胞群体组成，衰竭标志物表达水平低，体外具有高抗肿瘤特异性活性。我们没有发现 7 天和 11 天制造周期获得的产品性质存在显著差异，这有利于将生产时间缩短至 7 天，从而加速 CAR-T 治疗。我们已经表明，用于制造 CAR-T 的供体来源不会显著影响细胞产品的组成和功能特性。 结论：本研究表明，使用 CliniMACS Prodigy 系统缩短 7 天生产周期生产足够数量的功能性 CAR-T 细胞是可行的。CAR 转导效率可以通过功能测定间接测量。
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