Pancreatic ductal adenocarcinoma (PDAC) is one of the most malignant tumors. Macrophages are abundant in the tumor microenvironment, making them an attractive target for therapeutic intervention. While current immunotherapies, including immune checkpoint inhibition (ICI) and chimeric antigen receptor T (CAR-T) cells, have shown limited efficacy in pancreatic cancer, a novel approach involving chimeric antigen receptor macrophages (CAR-M) has, although promising, not been explored in pancreatic cancer. In this study, we first investigated the role of CAR-M cells targeting c-MET in pancreatic cancer. The effectiveness and rationality of c-MET as a target for CAR-M in pancreatic cancer were validated through bioinformatic analyses and immunohistochemical staining of samples from pancreatic cancer patients. We utilized flow cytometry and bioluminescence detection methods to demonstrate the specific binding and phagocytic killing effect of CAR-M on pancreatic cancer cells. Additionally, we observed the process of CAR-M engulfing pancreatic cancer cells using confocal microscopy and a long-term fluorescence live cell imaging system. In an in situ tumor model transplanted into NOD/SCID mice, we administered intraperitoneal injections of CAR-M to confirm its inhibitory function on pancreatic cancer. Furthermore, we validated these findings in human monocyte-derived macrophages (hMDM). Bioinformatics and tumor tissue microarray analyses revealed significantly higher expression levels of c-MET in tumor tissues, compared to the paired peritumoral tissues, and higher c-MET expression correlated with worse patient survival. CAR-M cells were engineered using human monocytic THP-1 cell line and hMDM targeting c-MET (CAR-M-c-MET). The CAR-M-c-MET cells demonstrated highly specific binding to pancreatic cancer cells and exhibited more phagocytosis and killing abilities than the pro-inflammatory polarized control macrophages. In addition, CAR-M-c-MET cells synergized with various cytotoxic chemotherapeutic drugs. In a NOD/SCID murine model, intraperitoneally injected CAR-M-c-MET cells rapidly migrated to tumor tissue and substantially inhibited tumor growth, which did not lead to obvious side effects. Cytokine arrays and mRNA sequencing showed that CAR-M-c-MET produced higher levels of immune activators than control macrophages. This study provides compelling evidence for the safety and efficacy of CAR-M therapy in treating pancreatic cancer. The results demonstrate that CAR-M-c-MET significantly suppresses pancreatic cancer progression and enhances the effectiveness of cytotoxic chemotherapy. Remarkably, no discernible side effects occur. Further clinical trials are warranted in human pancreatic cancer patients.

中文翻译：

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靶向 c-MET （CAR-M-C-MET） 的嵌合抗原受体巨噬细胞抑制胰腺癌进展并提高细胞毒性化疗疗效


胰腺导管腺癌 （PDAC） 是最恶性的肿瘤之一。巨噬细胞在肿瘤微环境中丰富，使其成为治疗干预的有吸引力的靶点。虽然目前的免疫疗法，包括免疫检查点抑制 （ICI） 和嵌合抗原受体 T （CAR-T） 细胞，在胰腺癌中显示出有限的疗效，但一种涉及嵌合抗原受体巨噬细胞 （CAR-M） 的新方法虽然很有希望，但尚未在胰腺癌中探索。在这项研究中，我们首先研究了靶向 c-MET 的 CAR-M 细胞在胰腺癌中的作用。通过对胰腺癌患者样本进行生物信息学分析和免疫组化染色，验证了 c-MET 作为胰腺癌 CAR-M 靶点的有效性和合理性。我们利用流式细胞术和生物发光检测方法来证明 CAR-M 对胰腺癌细胞的特异性结合和吞噬杀伤作用。此外，我们使用共聚焦显微镜和长期荧光活细胞成像系统观察了 CAR-M 吞噬胰腺癌细胞的过程。在移植到 NOD/SCID 小鼠体内的原位肿瘤模型中，我们腹膜内注射 CAR-M 以确认其对胰腺癌的抑制功能。此外，我们在人单核细胞衍生的巨噬细胞 （hMDM） 中验证了这些发现。生物信息学和肿瘤组织微阵列分析显示，与成对的瘤周组织相比，肿瘤组织中 c-MET 的表达水平显著升高，并且较高的 c-MET 表达与较差的患者生存率相关。CAR-M 细胞使用人单核细胞 THP-1 细胞系和靶向 c-MET 的 hMDM （CAR-M-c-MET） 进行工程改造。 CAR-M-c-MET 细胞表现出与胰腺癌细胞的高度特异性结合，并且比促炎极化对照巨噬细胞表现出更多的吞噬作用和杀伤能力。此外，CAR-M-c-MET 细胞与各种细胞毒性化疗药物协同作用。在 NOD/SCID 小鼠模型中，腹膜内注射的 CAR-M-c-MET 细胞迅速迁移到肿瘤组织并显着抑制肿瘤生长，未导致明显的副作用。细胞因子阵列和 mRNA 测序显示，CAR-M-c-MET 产生的免疫激活剂水平高于对照巨噬细胞。本研究为 CAR-M 疗法治疗胰腺癌的安全性和有效性提供了令人信服的证据。结果表明，CAR-M-c-MET 显着抑制胰腺癌进展并增强细胞毒性化疗的有效性。值得注意的是，没有明显的副作用发生。有必要在人类胰腺癌患者中进行进一步的临床试验。