Bister Arthur, Ibach Tabea, Haist Corinna, Smorra Denise, Roellecke Katharina, Wagenmann Martin, Scheckenbach Kathrin, Gattermann Norbert, Wiek Constanze, Hanenberg Helmut
Department of Otorhinolaryngology, Head & Neck Surgery, Heinrich Heine University, 40225 Düsseldorf, Germany.
Department of Pediatrics III, University Children's Hospital, University of Duisburg-Essen, Hufelandstrasse 55, 45147 Essen, Germany.
Mol Ther Oncolytics. 2021 Nov 11;23:534-546. doi: 10.1016/j.omto.2021.11.003. eCollection 2021 Dec 17.
Immunotherapy including chimeric antigen receptor (CAR) T cell therapy has revolutionized modern cancer therapy and has achieved remarkable remission and survival rates for several malignancies with historically dismal outcomes. The hinge of the CAR connects the antigen binding to the transmembrane domain and can be exploited to confer features to CAR T cells including additional stimulation, targeted elimination or detection and enrichment of the genetically modified cells. For establishing a novel hinge derived from human CD34, we systematically tested CD34 fragments of different lengths, all containing the binding site of the QBend-10 monoclonal antibody, in a FMC63-based CD19 CAR lentiviral construct. A final construct of 99 amino acids called C6 proved to be the best candidate for flow cytometry-based detection of CAR T cells and >95% enrichment of genetically modified T cells on MACS columns. The C6 hinge was functionally indistinguishable from the commonly used CD8α hinge as well as in experiments in NSG mice. We also showed that the C6 hinge can be used for a variety of different CARs and mediates high killing efficacy without unspecific activation by target antigen-negative cells, thus making C6 ideally suited as a universal hinge for CARs for clinical applications.
免疫疗法,包括嵌合抗原受体(CAR)T细胞疗法,已经彻底改变了现代癌症治疗,并在几种以往预后不佳的恶性肿瘤中取得了显著的缓解率和生存率。CAR的铰链区将抗原结合与跨膜结构域相连,可用于赋予CAR T细胞一些特性,包括额外的刺激、靶向清除或对基因改造细胞的检测和富集。为了构建一种源自人CD34的新型铰链区,我们在基于FMC63的CD19 CAR慢病毒构建体中系统地测试了不同长度的CD34片段,所有片段都包含QBend-10单克隆抗体的结合位点。一种名为C6的99个氨基酸的最终构建体被证明是基于流式细胞术检测CAR T细胞以及在MACS柱上对基因改造T细胞进行>95%富集的最佳候选者。在NSG小鼠的实验中,C6铰链区在功能上与常用的CD8α铰链区没有区别。我们还表明,C6铰链区可用于多种不同的CAR,并介导高杀伤效力,而不会被靶抗原阴性细胞非特异性激活,因此C6非常适合作为临床应用中CAR的通用铰链区。
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