Wang Siqi, Yu Yawen, Wang Aiping, Duan Xinliu, Sun Yuchen, Wang Liangxiao, Chu Liuxiang, Lv Yanan, Cui Nan, Fan Xuesong, Sha Chunjie, Xu Lixiao, Sun Kaoxiang
School of Pharmacy, Key Laboratory of Molecular Pharmacology and Drug Evaluation (Yantai University), Ministry of Education, Collaborative Innovation Center of Advanced Drug Delivery System and Biotech Drugs in Universities of Shandong, Yantai University, Yantai, China.
State Key Laboratory of Long-Acting and Targeting Drug Delivery System, Luye Pharmaceutical Co, Ltd, Yantai, China.
Front Pharmacol. 2022 Aug 17;13:965789. doi: 10.3389/fphar.2022.965789. eCollection 2022.
Temozolomide (TMZ) is the first-line drug for glioblastoma (GBM), but it is limited in clinical use due to the drug resistance, poor brain targeting, and side effects. Temozolomide hexadecyl ester (TMZ16e), a TMZ derivative with high lipophilicity, membrane permeability, and high anti-glioma properties, has the potential to reverse drug resistance. In this study, anti-ephrin type-A receptor 3 (EphA3) modified TMZ16e loaded nanoparticles (NPs) were prepared for targeted GBM therapy via intranasal administration to deliver TMZ16e to the brain, treat drug-resistant glioma effectively, and reduce peripheral toxicity. TMZ16e loaded NPs were prepared by emulsion solvent evaporation method followed by modified with anti-EphA3 (anti-EphA3-TMZ16e-NPs). evaluations were performed by an MTT assay and flow cytometry analysis. The orthotopic nude mice models were used to evaluate the anti-glioma effect . Additionally, we investigated the anti-drug resistant mechanism by western blot analysis. The particle size of the prepared NPs was less than 200 nm, and the zeta potential of TMZ16e-NPs and anti-EphA3-TMZ16e-NPs were -23.05 ± 1.48 mV and -28.65 ± 1.20mV, respectively, which is suitable for nasal delivery. studies have shown that anti-EphA3 modification increased the cellular uptake of nanoparticles in T98G cells. The cytotoxicity in the anti-EphA3-TMZ16e-NPs treated group was significantly higher than that of the TMZ16e-NPs, TMZ16e, and TMZ groups ( < 0.01), and the cell cycle was blocked. Western blotting analysis showed that the TMZ16e-loaded NPs were able to effectively downregulate the expression level of O6-methylguanine-deoxyribonucleic acid-methyltransferase (MGMT) protein in T98G cells and reverse drug resistance. studies showed that the median survival time of tumor-bearing nude mice in the anti-EphA3-TMZ16e-NPs group was extended to 41 days, which was 1.71-fold higher than that of the saline group and the TUNEL staining results of the brain tissue section indicated that the TMZ16e-loaded NPs could elevate apoptosis in T98G cells. In conclusion, the TMZ16e-loaded NPs can be effectively delivered to the brain and targeted to gliomas, exhibiting better anti-glioma activity, indicating they possess great potential in the treatment of drug-resistant glioma.
替莫唑胺（TMZ）是胶质母细胞瘤（GBM）的一线用药，但由于耐药性、脑靶向性差和副作用等问题，其临床应用受到限制。替莫唑胺十六烷基酯（TMZ16e）是一种具有高亲脂性、膜通透性和高抗胶质瘤特性的TMZ衍生物，具有逆转耐药性的潜力。在本研究中，制备了抗EphA3型受体3（EphA3）修饰的负载TMZ16e的纳米颗粒（NPs），通过鼻腔给药用于GBM的靶向治疗，将TMZ16e输送到脑部，有效治疗耐药性胶质瘤并降低外周毒性。采用乳液溶剂蒸发法制备负载TMZ16e的NPs，然后用抗EphA3进行修饰（抗EphA3-TMZ16e-NPs）。通过MTT法和流式细胞术分析进行评估。利用原位裸鼠模型评估抗胶质瘤效果。此外，我们通过蛋白质印迹分析研究了抗耐药机制。所制备的NPs粒径小于200nm，TMZ16e-NPs和抗EphA3-TMZ16e-NPs的zeta电位分别为-23.05±1.48mV和-28.65±1.20mV，适合鼻腔给药。研究表明，抗EphA3修饰增加了纳米颗粒在T98G细胞中的细胞摄取。抗EphA3-TMZ16e-NPs处理组的细胞毒性显著高于TMZ16e-NPs、TMZ16e和TMZ组（P＜0.01），且细胞周期被阻断。蛋白质印迹分析表明，负载TMZ16e的NPs能够有效下调T98G细胞中O6-甲基鸟嘌呤-脱氧核糖核酸甲基转移酶（MGMT）蛋白的表达水平并逆转耐药性。研究表明，抗EphA3-TMZ16e-NPs组荷瘤裸鼠的中位生存时间延长至41天，比生理盐水组高1.71倍，脑组织切片的TUNEL染色结果表明，负载TMZ16e的NPs可提高T98G细胞的凋亡率。总之，负载TMZ16e的NPs可有效输送至脑部并靶向胶质瘤，表现出更好的抗胶质瘤活性，表明它们在治疗耐药性胶质瘤方面具有巨大潜力。
