利用 CD133 特异性嵌合抗原受体 T 细胞通过非病毒 Sleeping Beauty 转座实现 PD-1 阻断 scFv 的靶向递送，显著提高晚期肝细胞癌的抗肿瘤疗效。
Targeted delivery of a PD-1-blocking scFv by CD133-specific CAR-T cells using nonviral Sleeping Beauty transposition shows enhanced antitumour efficacy for advanced hepatocellular carcinoma.
机构信息
Department of Experimental Research, State Key Laboratory of Oncology in South China, Collaborative Innovation Center for Cancer Medicine, Sun Yat-Sen University Cancer Center, Guangzhou, Guangdong, 510060, People's Republic of China.
Department of Biotherapy, Sun Yat-Sen University Cancer Center, Guangzhou, Guangdong, 510060, People's Republic of China.
出版信息
BMC Med. 2023 Aug 28;21(1):327. doi: 10.1186/s12916-023-03016-0.
BACKGROUND
CD133 is considered a marker for cancer stem cells (CSCs) in several types of tumours, including hepatocellular carcinoma (HCC). Chimeric antigen receptor-specific T (CAR-T) cells targeting CD133-positive CSCs have emerged as a tool for the clinical treatment of HCC, but immunogenicity, the high cost of clinical-grade recombinant viral vectors and potential insertional mutagenesis limit their clinical application.
METHODS
CD133-specific CAR-T cells secreting PD-1 blocking scFv (CD133 CAR-T and PD-1 s cells) were constructed using a sleeping beauty transposon system from minicircle technology, and the antitumour efficacy of CD133 CAR-T and PD-1 s cells was analysed in vitro and in vivo.
RESULTS
A univariate analysis showed that CD133 expression in male patients at the late stage (II and III) was significantly associated with worse progression-free survival (PFS) (P = 0.0057) and overall survival (OS) (P = 0.015), and a multivariate analysis showed a trend toward worse OS (P = 0.041). Male patients with advanced HCC exhibited an approximately 20-fold higher PD-L1 combined positive score (CPS) compared with those with HCC at an early stage. We successfully generated CD133 CAR-T and PD-1 s cells that could secrete PD-1 blocking scFv based on a sleeping beauty system involving minicircle vectors. CD133 CAR-T and PD-1 s cells exhibited significant antitumour activity against HCC in vitro and in xenograft mouse models. Thus, CD133 CAR-T and PD-1 s cells may be a therapeutically tractable strategy for targeting CD133-positive CSCs in male patients with advanced HCC.
CONCLUSIONS
Our study provides a nonviral strategy for constructing CAR-T cells that could also secrete checkpoint blockade inhibitors based on a Sleeping Beauty system from minicircle vectors and revealed a potential benefit of this strategy for male patients with advanced HCC and high CD133 expression (median immunohistochemistry score > 2.284).
背景
CD133 被认为是多种肿瘤包括肝细胞癌（HCC）中的癌症干细胞（CSC）的标志物。针对 CD133 阳性 CSCs 的嵌合抗原受体特异性 T（CAR-T）细胞已成为 HCC 临床治疗的一种工具，但免疫原性、临床级重组病毒载体的高成本和潜在的插入突变限制了其临床应用。
方法
使用源自微小环技术的睡眠美人转座子系统构建了分泌 PD-1 阻断 scFv 的 CD133 特异性 CAR-T 细胞（CD133 CAR-T 和 PD-1 s 细胞），并在体外和体内分析了 CD133 CAR-T 和 PD-1 s 细胞的抗肿瘤功效。
结果
单因素分析显示，晚期（II 和 III 期）男性患者的 CD133 表达与无进展生存期（PFS）（P=0.0057）和总生存期（OS）（P=0.015）的恶化显著相关，多因素分析显示 OS 恶化的趋势（P=0.041）。晚期 HCC 男性患者的 PD-L1 联合阳性评分（CPS）比早期 HCC 患者高约 20 倍。我们成功地基于涉及微小环载体的睡眠美人系统生成了能够分泌 PD-1 阻断 scFv 的 CD133 CAR-T 和 PD-1 s 细胞。CD133 CAR-T 和 PD-1 s 细胞在体外和异种移植小鼠模型中对 HCC 具有显著的抗肿瘤活性。因此，CD133 CAR-T 和 PD-1 s 细胞可能是针对晚期 HCC 男性患者中 CD133 阳性 CSCs 的一种有治疗前途的策略。
结论
我们的研究提供了一种非病毒策略，用于构建基于微小环载体的睡眠美人系统的 CAR-T 细胞，该系统还可以分泌检查点抑制剂阻断剂，并揭示了该策略对高 CD133 表达（中位数免疫组织化学评分>2.284）的晚期 HCC 男性患者的潜在益处。
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