Automated generation of gene-edited CAR T cells at clinical scale.
Automated generation of gene-edited CAR T cells at clinical scale.
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在临床规模上自动生成基因编辑的CAR T细胞。
DOI：
10.1016/j.omtm.2020.12.008
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发表时间：
2021-03-12
期刊：
影响因子：
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通讯作者：
Cornu TI
中科院分区：
文献类型：
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作者：
Alzubi J;Lock D;Rhiel M;Schmitz S;Wild S;Mussolino C;Hildenbeutel M;Brandes C;Rositzka J;Lennartz S;Haas SA;Chmielewski KO;Schaser T;Kaiser A;Cathomen T;Cornu TI
当与基因组编辑相结合时，采用细胞疗法的潜力可以扩大。然而，原材料质量的差异和不同的制造步骤与生产失败和产品污染有关。在这里，我们提出了一种自动化的T细胞工程过程，在包含在线电穿孔单元的扩展CliniMACS Prodigy平台上生产现成的嵌合抗原受体(CAR)T细胞。该装置用于将慢病毒传递CD19靶向CAR与转移编码TRAC位点靶向转录激活物样效应核酸酶(TALEN)的mRNA相结合。在临床规模的三次运行中，编码TRAC基因的T细胞受体(TCR)阿尔法链在35%的细胞活力高(90%)的细胞中被破坏，并且没有检测到脱靶活性。最后的阴性选择步骤允许产生纯度为99.5%的不含TcRα/β的CAR T细胞。这些CAR T细胞增殖良好，保持T细胞记忆表型，消除CD19阳性肿瘤细胞，并在接触B细胞白血病细胞时释放预期的细胞因子。总之，我们建立了一种自动化的、符合GMP的工艺，该工艺将慢病毒转导与TALEN mRNA的电穿孔相结合，以生产临床规模的功能性TCRCAR19T细胞。CAR T细胞免疫疗法的潜力很可能在与基因组编辑相结合时得到延长。Katomen和他的同事开发了一种符合GMP的自动化流程，将慢病毒车递送与临床规模的TRAC编辑相结合。这些方案将促进现成产品的生产及其临床应用的成功。
The potential of adoptive cell therapy can be extended when combined with genome editing. However, variation in the quality of the starting material and the different manufacturing steps are associated with production failure and product contamination. Here, we present an automated T cell engineering process to produce off-the-shelf chimeric antigen receptor (CAR) T cells on an extended CliniMACS Prodigy platform containing an in-line electroporation unit. This setup was used to combine lentiviral delivery of a CD19-targeting CAR with transfer of mRNA encoding a TRAC locus-targeting transcription activator-like effector nuclease (TALEN). In three runs at clinical scale, the T cell receptor (TCR) alpha chain encoding TRAC locus was disrupted in >35% of cells with high cell viability (>90%) and no detectable off-target activity. A final negative selection step allowed the generation of TCRα/β-free CAR T cells with >99.5% purity. These CAR T cells proliferated well, maintained a T cell memory phenotype, eliminated CD19-positive tumor cells, and released the expected cytokines when exposed to B cell leukemia cells. In conclusion, we established an automated, good manufacturing practice (GMP)-compliant process that integrates lentiviral transduction with electroporation of TALEN mRNA to produce functional TCRα/β-free CAR19 T cells at clinical scale. The potential of CAR T cell immunotherapy can likely be extended when combined with genome editing. Cathomen and colleagues developed a GMP-compliant, automated process that integrates lentiviral CAR delivery with TRAC editing at clinical scale. Such protocols will facilitate the production of off-the-shelf products and their successful clinical application.
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影响因子：
16.6
作者：
Sachdeva, Mohit;Busser, Brian W.;Valton, Julien
通讯作者：
Valton, Julien
影响因子：
64.8
作者：
Eyquem J;Mansilla-Soto J;Giavridis T;van der Stegen SJ;Hamieh M;Cunanan KM;Odak A;Gönen M;Sadelain M
通讯作者：
Sadelain M
影响因子：
11.4
作者：
Cooper ML;Choi J;Staser K;Ritchey JK;Devenport JM;Eckardt K;Rettig MP;Wang B;Eissenberg LG;Ghobadi A;Gehrs LN;Prior JL;Achilefu S;Miller CA;Fronick CC;O'Neal J;Gao F;Weinstock DM;Gutierrez A;Fulton RS;DiPersio JF
通讯作者：
DiPersio JF
影响因子：
8
作者：
Chen, Pei-Hsuan;Lipschitz, Mikel;Rodig, Scott J.
通讯作者：
Rodig, Scott J.
DOI：
10.1038/mtna.2014.64
发表时间：
2014-12-02
期刊：
Molecular therapy. Nucleic acids
影响因子：
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作者：
通讯作者：
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