Inserting EF1α-driven CD7-specific CAR at CD7 locus reduces fratricide and enhances tumor rejection
Inserting EF1α-driven CD7-specific CAR at CD7 locus reduces fratricide and enhances tumor rejection
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DOI：
10.1038/s41375-023-01948-3
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发表时间：
2023-06
期刊：
影响因子：
11.4
通讯作者：
Jie Jiang;Jiangqing Chen;C. Liao;Y. Duan;Yajie Wang;Kai Shang;Yanjie Huang;Yongming Tang;Xiaofei Gao;Ying Gu;Jie Sun
中科院分区：
文献类型：
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作者：
Jie Jiang;Jiangqing Chen;C. Liao;Y. Duan;Yajie Wang;Kai Shang;Yanjie Huang;Yongming Tang;Xiaofei Gao;Ying Gu;Jie Sun
治疗T细胞恶性肿瘤的CAR-T疗法面临着独特的障碍。正常和恶性T细胞通常表达相同的CAR靶标，导致自相残杀。靶向在各种恶性T细胞中表达的CD 7的CAR-T细胞由于自相残杀而具有有限的扩增。使用CRISPR/Cas9敲除CD 7可以减少自相残杀。在这里，我们开发了一种二合一策略，将EF 1 α驱动的CD 7特异性CAR插入到破坏的CD 7位点，并将其与其他两种已知策略进行了比较：一种是通过逆转录病毒随机整合CAR，另一种是在T细胞受体α恒定（TRAC）位点的位点特异性整合，两者都是在CD 7破坏的背景下。所有三种类型的具有减少的杀兄弟作用的CD 7 CAR-T细胞都可以很好地扩增，并对CD 7+肿瘤细胞系和患者来源的原发性肿瘤显示出强效的细胞毒性。此外，在CD 7位点表达的EF 1 α驱动的CAR增强了T细胞急性淋巴细胞白血病（T-ALL）小鼠异种移植模型的肿瘤排斥反应，表明了巨大的临床应用潜力。此外，采用这种二合一策略来产生CD 7特异性CAR-NK细胞，因为NK也表达CD 7，这将防止恶性细胞的污染。因此，我们的同步抗原敲除CAR敲入策略可以减少自相残杀并增强抗肿瘤活性，推进T细胞恶性肿瘤的临床CAR-T治疗。
CAR-T therapies to treat T-cell malignancies face unique hurdles. Normal and malignant T cells usually express the same target for CAR, leading to fratricide. CAR-T cells targeting CD7, which is expressed in various malignant T cells, have limited expansion due to fratricide. Using CRISPR/Cas9 to knockout CD7 can reduce the fratricide. Here we developed a 2-in-1 strategy to insert EF1α-driven CD7-specific CAR at the disruptedCD7locus and compared it to two other known strategies: one was random integration of CAR by a retrovirus and the other was site-specific integration at T-cell receptor alpha constant (TRAC)locus, both in the context of CD7 disruption. All three types of CD7 CAR-T cells with reduced fratricide could expand well and displayed potent cytotoxicity to both CD7+tumor cell lines and patient-derived primary tumors. Moreover, EF1α-driven CAR expressed at theCD7locus enhances tumor rejection in a mouse xenograft model of T-cell acute lymphoblastic leukemia (T-ALL), suggesting great clinical application potential. Additionally, this 2-in-1 strategy was adopted to generate CD7-specific CAR-NK cells as NK also expresses CD7, which would prevent contamination from malignant cells. Thus, our synchronized antigen-knockout CAR-knockin strategy could reduce the fratricide and enhance anti-tumor activity, advancing clinical CAR-T treatment of T-cell malignancies.