Abstract：
CD28 and 4-1BB costimulatory endodomains included in chimeric antigen receptor (CAR) molecules play a critical role in promoting sustained antitumor activity of CAR-T cells. However, the molecular events associated with the ectopic and constitutive display of either CD28 or 4-1BB in CAR-T cells have been only partially explored. In the current study, we demonstrated that 4-1BB incorporated within the CAR leads to cell cluster formation and cell death in the forms of both apoptosis and necroptosis in the absence of CAR tonic signaling. Mechanistic studies illustrate that 4-1BB sequesters A20 to the cell membrane in a TRAF-dependent manner causing A20 functional deficiency that in turn leads to NF-κB hyperactivity, cell aggregation via ICAM-1 overexpression, and cell death including necroptosis via RIPK1/RIPK3/MLKL pathway. Genetic modulations obtained by either overexpressing A20 or releasing A20 from 4-1BB by deleting the TRAF-binding motifs of 4-1BB rescue cell cluster formation and cell death and enhance the antitumor ability of 4-1BB-costimulated CAR-T cells.
摘要：
包含在嵌合抗原受体(CAR)分子中的CD28和4-1BB共刺激内域在促进CAR-T细胞的持续抗肿瘤活性中起关键作用。然而,与CAR-T细胞中CD28或4-1BB的异位和组成性展示相关的分子事件仅得到部分研究.在目前的研究中,我们证明,在不存在CAR强直信号的情况下,4-1BB掺入CAR导致细胞簇形成和细胞死亡,表现为细胞凋亡和坏死.机制研究表明，4-1BB以TRAF依赖性方式将A20隔离到细胞膜上，导致A20功能缺乏，进而导致NF-κB过度活跃。通过ICAM-1过表达的细胞聚集，和细胞死亡，包括通过RIPK1/RIPK3/MLKL途径的坏死。通过过表达A20或通过删除4-1BB拯救细胞簇形成和细胞死亡的TRAF结合基序从4-1BB释放A20而获得的遗传调制，并增强4-1BB共刺激的CAR-T细胞的抗肿瘤能力。
