State Key Laboratory of Experimental Hematology, National Clinical Research Center for Blood Diseases, Haihe Laboratory of Cell Ecosystem, Institute of Hematology and Blood Diseases Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Tianjin, China.
Tianjin Key Laboratory of Cell Therapy for Blood Diseases, Institute of Hematology and Blood Diseases Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Tianjin, China.
Hum Gene Ther. 2023 Dec;34(23-24):1257-1272. doi: 10.1089/hum.2023.029. Epub 2023 Nov 24.
Chimeric antigen receptor T (CAR-T) cell therapy initiates new methods and turns the scale of clinical treatment on relapsed/refractory acute T lymphoblastic leukemia (T-ALL). In this study, we generated the second-generation CD7-targeting CAR-T cells with a new antigen-binding single-chain variable fragment sequence and made it universal via CRISPR-based knockout of and genes (termed UCAR-T). The CD7 UCAR-T cells can efficiently proliferate and lyse T-ALL tumor cell , along with prominent proinflammatory cytokines secretion. A Jurkat-based xenograft mouse model further verified the superior cytotoxicity of the UCAR-T cells . During the UCAR-T construction, we observed a CD4/CD8 ratio shift among CD7 T/CAR-T cells, which motivated us to further analyze the effects of CD7 antigen on T/CAR-T cells. We sorted out CD7 T or anti-CD19 CAR-T cells after partially CD7 knockout and performed functional, phenotypic detection, as well as translational analysis. CD7 CAR-T cells tended to be CD8 negative and showed slightly better cytotoxicity at long-term assay. RNA-seq further confirmed an elevation of activated CD4 memory cell subpopulation. However, limited distinction on crucial regulatory genes and pathways was revealed, suggesting the safety and feasibility of UCAR-T application as well as the potential translational rather than transcriptional regulation of CD7 antigen.
嵌合抗原受体 T (CAR-T) 细胞疗法开创了新方法，并在复发/难治性急性 T 淋巴细胞白血病 (T-ALL) 的临床治疗中取得了重大进展。在这项研究中，我们利用一种新型抗原结合单链可变片段序列，生成了第二代靶向 CD7 的 CAR-T 细胞，并通过基于 CRISPR 的 和 基因敲除使其具有通用性（称为 UCAR-T）。CD7 UCAR-T 细胞可以有效地增殖和裂解 T-ALL 肿瘤细胞，并伴随着明显的促炎细胞因子分泌。基于 Jurkat 的异种移植小鼠模型进一步验证了 UCAR-T 细胞的优越细胞毒性。在 UCAR-T 构建过程中，我们观察到 CD7 T/CAR-T 细胞中的 CD4/CD8 比值发生了变化，这促使我们进一步分析 CD7 抗原对 T/CAR-T 细胞的影响。我们对部分 CD7 敲除后的 CD7 T 或抗 CD19 CAR-T 细胞进行了分选，并进行了功能、表型检测和转化分析。CD7 CAR-T 细胞倾向于 CD8 阴性，并且在长期检测中表现出稍好的细胞毒性。RNA-seq 进一步证实了激活的 CD4 记忆细胞亚群的增加。然而，关键调控基因和通路的差异有限，这表明 UCAR-T 应用的安全性和可行性，以及 CD7 抗原的潜在转化而非转录调控。
