Evaluation of Intracellular Signaling Downstream Chimeric Antigen Receptors.
Evaluation of Intracellular Signaling Downstream Chimeric Antigen Receptors.
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DOI：
10.1371/journal.pone.0144787
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发表时间：
2015
期刊：
影响因子：
3.7
通讯作者：
Loskog A
中科院分区：
文献类型：
--
作者：
Karlsson H;Svensson E;Gigg C;Jarvius M;Olsson-Strömberg U;Savoldo B;Dotti G;Loskog A
靶向CD 19的CAR T细胞在靶向B细胞白血病的临床试验中显示出效力。尽管已经在患者中主要研究了携带CD 28或4-1BB的第二代（2G）汽车，但临床前研究表明具有CD 28和4-1BB的第三代（3G）汽车具有增强的能力。然而，关于汽车下游的细胞内信号传导途径知之甚少。在本工作中，我们分析了2G和3G汽车中抗原刺激后的信号传导能力。在用IL-2和自体B细胞重复刺激后，3G CAR T细胞比2G CAR T细胞更好地扩增。抗原刺激后CAR+细胞的抗原驱动的积累是明显的。通过重复刺激维持2G和3G CAR T细胞的细胞毒性。抗原刺激后细胞内信号传导蛋白的磷酸化状态显示3G CAR T细胞具有比2G更高的活化状态。参与TCR下游信号传导的几种蛋白质被激活，参与细胞周期、细胞粘附和胞吐的蛋白质也被激活。总之，3G CAR T细胞具有比2G汽车更高程度的细胞内信号传导活性，这可以解释在3G CAR T细胞中观察到的增加的增殖能力。该研究还表明，在设计或评估新一代汽车时，可能需要考虑其他信号通路。
CD19-targeting CAR T cells have shown potency in clinical trials targeting B cell leukemia. Although mainly second generation (2G) CARs carrying CD28 or 4-1BB have been investigated in patients, preclinical studies suggest that third generation (3G) CARs with both CD28 and 4-1BB have enhanced capacity. However, little is known about the intracellular signaling pathways downstream of CARs. In the present work, we have analyzed the signaling capacity post antigen stimulation in both 2G and 3G CARs. 3G CAR T cells expanded better than 2G CAR T cells upon repeated stimulation with IL-2 and autologous B cells. An antigen-driven accumulation of CAR+ cells was evident post antigen stimulation. The cytotoxicity of both 2G and 3G CAR T cells was maintained by repeated stimulation. The phosphorylation status of intracellular signaling proteins post antigen stimulation showed that 3G CAR T cells had a higher activation status than 2G. Several proteins involved in signaling downstream the TCR were activated, as were proteins involved in the cell cycle, cell adhesion and exocytosis. In conclusion, 3G CAR T cells had a higher degree of intracellular signaling activity than 2G CARs which may explain the increased proliferative capacity seen in 3G CAR T cells. The study also indicates that there may be other signaling pathways to consider when designing or evaluating new generations of CARs.