背景:放射抗性和免疫逃逸是胶质母细胞瘤(GBM)治疗效果不理想的关键原因。尽管在骨髓细胞-2(TREM2)上表达的触发受体参与形成免疫抑制微环境,但其介导癌症放射抗性的潜在机制及其作用仍不清楚,此外,靶向TREM2的药物向GBM的有效递送面临严峻挑战.因此,本研究旨在阐明靶向TREM2沉默逆转谷胱甘肽反应性仿生纳米粒子(NP)平台辅助下GBM的放射抗性和免疫逃逸的作用和机制.
方法:首先建立抗辐射GBM细胞系和TREM2稳定敲低GBM细胞系。RNA测序,集落形成试验,westernblot,采用酶联免疫吸附法和免疫共沉淀法检测TREM2调节GBM放射抗性和免疫逃逸的分子机制。谷胱甘肽响应的仿生NP,然后构建血管肽-2(A2)-细胞膜(CM)-NP/siTREM2/spam1,以三重并靶向抑制TREM2用于体内研究。建立原位GBM小鼠模型以评估TREM2抑制的抗GBM作用。采用多重免疫荧光法检测免疫细胞的浸润情况。
结果:TREM2通过参与DNA损伤修复并与高迁移率族蛋白盒1(HMGB1)形成正反馈回路,级联激活Toll样受体4(TLR4)/蛋白激酶B(Akt)信号,从而加速GBM的放射抗性和免疫逃逸。成功合成了A2-CM-NP/siTREM2/spam1,具有良好的被动靶向性,主动靶向和同源靶向,通过促进1型辅助性T细胞和CD8+T细胞的浸润,减少2型辅助性T细胞和调节性T细胞的浸润,将巨噬细胞重新极化为M1型,并减少促肿瘤和免疫抑制细胞因子的分泌。
结论:靶向TREM2治疗是优化放疗和免疫治疗以改善GBM患者预后的有希望的途径。
BACKGROUND: Radioresistance and immune escape are crucial reasons for unsatisfactory therapeutic effects of glioblastoma (GBM). Although triggering receptor expressed on myeloid cells-2 (TREM2) involved in forming immunosuppressive microenvironment, but the underlying mechanism and its roles in mediating cancer
radioresistance remain unclear, moreover, the efficient delivery of drugs targeting TREM2 to GBM encounters serious challenges. Hence, this study aimed to elucidate the effect and mechanisms of targeted TREM2 silencing on reversing the
radioresistance and immune escape of GBM aided by a glutathione-responsive biomimetic nanoparticle (NP) platform.
METHODS: Radioresistant GBM cell lines and TREM2 stable knockdown GBM cell lines were firstly established. RNA sequencing, colony formation assay, western blot, enzyme-linked immunosorbent assay and co-immunoprecipitation assay were used to detect the molecular mechanisms of TREM2 in regulating the
radioresistance and immune escape of GBM. The glutathione-responsive biomimetic NP, angiopep-2 (A2)- cell membrane (CM)-NP/siTREM2/spam1, was then constructed to triply and targeted inhibit TREM2 for in vivo study. Orthotopic GBM-bearing mouse models were established to evaluate the anti-GBM effect of TREM2 inhibition, multiplex immunofluorescence assay was conducted to detect the infiltration of immune cells.
RESULTS: TREM2 was a regulator in accelerating the
radioresistance and immune escape of GBM through participating in DNA damage repair and forming a positive feedback loop with high mobility group box 1 (HMGB1) to cascade the activation of Toll-like receptor 4 (TLR4)/protein kinase B (Akt) signaling. A2-CM-NP/siTREM2/spam1 was successfully synthesized with excellent passive targeting, active targeting and homologous targeting, and the in vivo results exhibited its remarkable anti-GBM therapeutic effect through promoting the infiltration of type 1 helper T cells and CD8+T cells, reducing the infiltration of type 2 helper T cells and regulatory T cells, repolarizing macrophages to M1-type, and decreasing the secretion of pro-tumor and immunosuppressive cytokines.
CONCLUSIONS: Targeting TREM2 therapy is a promising avenue for optimizing radiotherapy and immunotherapy to improve the prognosis of GBM patients.