抑制VEGF信号传导可通过Wnt/β-连环蛋白途径防止CAR-T细胞耗竭并增强其抗白血病疗效。
Inhibition of VEGF signaling prevents exhaustion and enhances anti-leukemia efficacy of CAR-T cells via Wnt/β-catenin pathway.
作者信息
Wu Suwan, Wei Yiyi, Qiu Yingqi, Ai Kexin, Chen Mu, Wang Hao, Zhang Honghao, Cen Qingyan, Liao Peiyun, Ding Xiangyang, Xie Xiaoling, Li Yuhua
机构信息
Department of Hematology, Zhujiang Hospital, Southern Medical University, Guangzhou, China.
Bioland laboratory (Guangzhou Regenerative Medicine and Health Guangdong Laboratory), Guangzhou, China.
出版信息
J Transl Med. 2025 Apr 30;23(1):494. doi: 10.1186/s12967-024-05907-z.
BACKGROUND
Current challenges in Chimeric Antigen Receptor (CAR) -T cell therapy for hematological cancers include T cell exhaustion and limited persistence, which contribute to cancer relapse.
METHODS
The effects of Axitinib, a VEGFR inhibitor, on the biological functions of CAR-T cells in vitro and in vivo were investigated by comparing CAR-T cells pre-treated ex vivo with Axitinib, as well as utilizing a B-ALL mouse model. Real-time quantitative PCR and Western blotting were employed to detect the expression of molecules related to differentiation, exhaustion, and the Wnt pathway in CAR-T cells. Flow cytometry was used to assess changes in CAR-T cell differentiation, exhaustion, activation, apoptosis, proliferation, and cytokine secretion. Western blotting and flow cytometry were used to assess changes in VEGFR expression. Bioluminescence imaging, flow cytometry, and immunohistochemistry (IHC) analysis were used to evaluate changes in tumor burden in mice receiving different treatments, while hematoxylin and eosin (H&E) staining were used to monitor histological changes in the liver and spleen of mice.
RESULTS
Axitinib treatment notably reduced CAR-T cell exhaustion and terminal differentiation both under tonic signaling and tumor antigen exposure scenarios. Furthermore, CAR-T cells pretreated with Axitinib demonstrated enhanced anti-tumor efficacy and prolonged survival in vivo. Mechanistically, Axitinib treatment upregulated the Wnt/β-catenin signaling pathway in CAR-T cells. Using agonists/inhibitors of the Wnt/β-catenin pathway could respectively mimic or counteract the effects of Axitinib on CAR-T cell exhaustion and differentiation. CAR-T cells treated with Axitinib can inhibit the VEGFR2 pathway. CAR-T cells treated with anti-VEGFR2 antibody can activate the Wnt/β-catenin pathway and prevent CAR-T cell exhaustion.
CONCLUSION
Axitinib confers resistance to exhaustion in CAR-T cells by modulating the Wnt/β-catenin signaling pathway.
背景
嵌合抗原受体（CAR）-T细胞疗法在血液系统癌症治疗中的当前挑战包括T细胞耗竭和持久性有限，这会导致癌症复发。
方法
通过将体外预先用阿昔替尼处理的CAR-T细胞进行比较，并利用B-ALL小鼠模型，研究VEGFR抑制剂阿昔替尼对CAR-T细胞体外和体内生物学功能的影响。采用实时定量PCR和蛋白质印迹法检测CAR-T细胞中与分化、耗竭和Wnt通路相关分子的表达。流式细胞术用于评估CAR-T细胞分化、耗竭、激活、凋亡、增殖和细胞因子分泌的变化。蛋白质印迹法和流式细胞术用于评估VEGFR表达的变化。生物发光成像、流式细胞术和免疫组织化学（IHC）分析用于评估接受不同治疗的小鼠的肿瘤负荷变化，而苏木精和伊红（H&E）染色用于监测小鼠肝脏和脾脏的组织学变化。
结果
在静息信号和肿瘤抗原暴露情况下，阿昔替尼治疗均显著降低了CAR-T细胞的耗竭和终末分化。此外，用阿昔替尼预处理的CAR-T细胞在体内表现出增强的抗肿瘤疗效和延长的生存期。机制上，阿昔替尼治疗上调了CAR-T细胞中的Wnt/β-连环蛋白信号通路。使用Wnt/β-连环蛋白通路的激动剂/抑制剂可分别模拟或抵消阿昔替尼对CAR-T细胞耗竭和分化的影响。用阿昔替尼处理的CAR-T细胞可抑制VEGFR2通路。用抗VEGFR2抗体处理的CAR-T细胞可激活Wnt/β-连环蛋白通路并防止CAR-T细胞耗竭。
结论
阿昔替尼通过调节Wnt/β-连环蛋白信号通路赋予CAR-T细胞抗耗竭能力。
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