Department of Clinical Biochemistry, School of Medicine, Tehran University of Medical Sciences, Tehran, Iran.
Department of Clinical Biochemistry, School of Medicine, Tehran University of Medical Sciences, Tehran, Iran.
Life Sci. 2020 Nov 15;261:118361. doi: 10.1016/j.lfs.2020.118361. Epub 2020 Aug 28.
AIM: Antibody-conjugated nanoparticles have attracted much attention in the field of cancer treatment due to the enhancement of the tumor cell response to anticancer drugs as well as reducing the side effects of chemotherapeutic agents on healthy tissues. However, most studies in this field generally mentioned the specific cellular uptake of conjugated nanoparticles. In this study, we loaded doxorubicin (DXR: as an effective antineoplastic agent) in PLGA-PEG (D,L-lactic-co-glycolic acid)-(polyethylene glycol) biocompatible polymeric nanoparticles (NPs) and then conjugated with anti-EGFRvIII antibody. The resulting nanoparticles had remarkable sensitivity to pH decrease and were capable of targeting specific cells. MATERIALS AND METHODS: To this aim, PLGA-PEG-COOH was used for the synthesis of nanoparticles and stabilized by polyvinyl alcohol (PVA) according to the nanoprecipitation method. The carboxylic groups on the surface of PLGA-PEG NPs were activated by EDC/NHS and covalently conjugated to amino groups of the monoclonal antibody. The prepared NPs were characterized by Zetasizer and transmission electron microscopy (TEM). The resulting NPs were evaluated in terms of entrapment efficiency (EE), drug loading efficiency (DLE), drug-release profile, and cell internalization. Intrinsic cytotoxicity was assessed by the MTT, apoptosis (Annexin V-PI) and cell cycle assays. KEY FINDINGS: The in vitro drug release assessment of conjugated particles (MAb-DXR-PLGA NPs) showed a slow sustained DXR release in physiological pH (7.4) values, while the initial drug release was markedly higher (the 1.9 fold) in acidic pH (6.5) ranges. The selectivity for cellular internalization of MAb-DXR-PLGA NPs into U87MG vIII cells (overexpressing EGFRvIII) in comparison with U87MG cells (lacking EGFRvIII expression) was also confirmed. The MTT assay demonstrated that the cytotoxicity of MAb-DXR-PLGA NPs against U87MG vIII cells was more pronounced when compared with BSA-DXR-PLGA NPs. The results of the MTT assay were also confirmed by apoptosis and cell cycle assays. SIGNIFICANCE: Our findings suggest that the designed anti-EGFRvIII MAb-DXR-PLGA NPs could be considered as a proper option for targeted drug delivery systems due to pH sensitivity and specific cellular internalization.
目的：由于抗体偶联纳米颗粒增强了肿瘤细胞对抗癌药物的反应，同时减少了化疗药物对健康组织的副作用，因此在癌症治疗领域引起了广泛关注。然而，该领域的大多数研究通常仅提到了共轭纳米颗粒的特定细胞摄取。在这项研究中，我们将阿霉素（DXR：一种有效的抗肿瘤药物）载入 PLGA-PEG（D，L-乳酸-乙醇酸）-（聚乙二醇）生物相容性聚合物纳米颗粒（NPs）中，然后与抗 EGFRvIII 抗体结合。所得纳米颗粒对 pH 值降低具有显著的敏感性，并且能够靶向特定细胞。
材料与方法：为此，使用 PLGA-PEG-COOH 合成纳米颗粒，并根据纳米沉淀法用聚乙烯醇（PVA）稳定。PLGA-PEG NPs 表面的羧酸基团通过 EDC/NHS 活化，并与单克隆抗体的氨基共价结合。用 Zetasizer 和透射电子显微镜（TEM）对制备的 NPs 进行了表征。通过 MTT、凋亡（Annexin V-PI）和细胞周期测定评估了所得 NPs 的包封效率（EE）、载药量（DLE）、药物释放曲线和细胞内化。通过 MTT、细胞凋亡（Annexin V-PI）和细胞周期测定评估了内在细胞毒性。
主要发现：偶联颗粒（MAb-DXR-PLGA NPs）的体外药物释放评估表明，在生理 pH 值（7.4）范围内，DXR 的释放缓慢且持续，而在酸性 pH 值（6.5）范围内，初始药物释放明显更高（1.9 倍）。还证实了 MAb-DXR-PLGA NPs 对 U87MG vIII 细胞（过表达 EGFRvIII）的细胞内化选择性高于 U87MG 细胞（缺乏 EGFRvIII 表达）。MTT 测定表明，与 BSA-DXR-PLGA NPs 相比，MAb-DXR-PLGA NPs 对 U87MG vIII 细胞的细胞毒性更为明显。MTT 测定的结果也通过细胞凋亡和细胞周期测定得到了证实。
意义：我们的研究结果表明，由于具有 pH 敏感性和特定的细胞内化作用，设计的抗 EGFRvIII MAb-DXR-PLGA NPs 可被视为靶向药物递送系统的一种合适选择。
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