Zhang Yuanyuan, Huang Meijuan, Zhang Shujia, Liu Tianjiao, Ye Shanwei, Cheng Yuhang, Cao Yang, Chen Liting, Zhu Li, Sun Xueyan, Shen Kefeng, Xu Qian, Li Tongjuan, Li Dengju, Huang Liang, Mu Wei, Zhao Lei, Wang Jue
Department of Hematology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China.
Immunotherapy Research Center for Hematologic Diseases of Hubei Province, Wuhan, China.
Exp Hematol Oncol. 2025 Aug 25;14(1):110. doi: 10.1186/s40164-025-00701-z.
Chimeric antigen receptor (CAR)-T-cell therapy has achieved remarkable clinical success in the treatment of B-cell malignancies; however, its efficacy can be limited by poor T-cell persistence and insufficient antitumor activity in certain cases. Moreover, interleukin-12 (IL-12) is a prominent agent in cancer immunotherapy, but its clinical application is constrained by severe toxicity associated with systemic exposure. In this study, we developed a novel cytokine delivery platform based on CAR target-modified cell-derived extracellular vesicles (EVs) that preferentially bind CAR-T cells to improve CAR-T-cell function. EVs with surface-displayed CD19 and/or IL-12 were successfully generated from HEK-293T cells. Compared with an equivalent concentration of rhIL-12, IL-12 EVs significantly enhanced the effector function of anti-CD19 CAR-T cells in vitro, resulting in increased Interferon-γ (IFN-γ) and TNF-α secretion, cytolytic activity, and T-cell expansion. Additionally, compared with EVs expressing IL-12 alone, EVs co-expressing IL-12 and CD19 (CD19/IL-12 EVs) exhibited superior binding efficiency to CAR-T cells but not to T cells, as indicated by flow cytometry. In xenograft model mice bearing CD19 + Raji tumors, intratumoral injection of CD19/IL-12 EVs resulted in durable antitumor responses and enhanced the in vivo expansion of CAR-T cells, outperforming CD19 EVs, IL-12 EVs and control EVs, without causing systemic toxicity. RNA sequencing (RNA-seq) analysis of CAR-T cells stimulated with EVs suggested that the increased efficacy was driven by IL-12 signaling. These data demonstrate that CAR-targeted modified EVs may serve as targeted cytokine delivery systems for CAR-T cells, offering a safe and effective strategy to augment CAR-T-cell function.
嵌合抗原受体（CAR）-T细胞疗法在治疗B细胞恶性肿瘤方面取得了显著的临床成功；然而，在某些情况下，其疗效可能会受到T细胞持久性差和抗肿瘤活性不足的限制。此外，白细胞介素-12（IL-12）是癌症免疫治疗中的一种重要药物，但其临床应用受到与全身暴露相关的严重毒性的限制。在本研究中，我们开发了一种基于CAR靶点修饰的细胞衍生细胞外囊泡（EVs）的新型细胞因子递送平台，该平台优先结合CAR-T细胞以改善CAR-T细胞功能。成功从人胚肾293T细胞（HEK-293T细胞）中产生了表面展示CD19和/或IL-12的EVs。与同等浓度的重组人IL-12（rhIL-12）相比，IL-12 EVs在体外显著增强了抗CD19 CAR-T细胞的效应功能，导致干扰素-γ（IFN-γ）和肿瘤坏死因子-α（TNF-α）分泌增加、细胞溶解活性增强以及T细胞扩增。此外，流式细胞术结果表明，与单独表达IL-12的EVs相比，共表达IL-12和CD19的EVs（CD19/IL-12 EVs）对CAR-T细胞表现出更高的结合效率，但对T细胞则不然。在携带CD19 + 拉吉（Raji）肿瘤的异种移植模型小鼠中，瘤内注射CD19/IL-12 EVs产生了持久的抗肿瘤反应，并增强了CAR-T细胞在体内的扩增，其效果优于CD19 EVs、IL-12 EVs和对照EVs，且未引起全身毒性。对用EVs刺激的CAR-T细胞进行的RNA测序（RNA-seq）分析表明，疗效的提高是由IL-12信号传导驱动的。这些数据表明，CAR靶向修饰的EVs可作为CAR-T细胞的靶向细胞因子递送系统，为增强CAR-T细胞功能提供了一种安全有效的策略。
