Departamento de Química Orgánica, Facultad de Química y de Farmacia, Pontificia Universidad Católica de Chile, Av. Vicuña McKenna 4860, 7820436, Macul, Santiago, Chile; Department of Pharmaceutical Sciences, Irma Lerma Rangel College of Pharmacy, Texas A&M University, USA.
Chemical Biology & Drug Discovery Lab, Facultad de Medicina y Ciencia, Universidad San Sebastián, Campus Los Leones, Lota 2465 Providencia 7510157, Santiago, Chile.
Biomed Pharmacother. 2022 Sep;153:113428. doi: 10.1016/j.biopha.2022.113428. Epub 2022 Jul 15.
Ligand-based targeting of the receptors that are overexpressed explicitly on cancer cells represents an effective drug delivery approach to enhance the chemotherapeutic efficacy. Proprotein convertase subtilisin/kexin type 9 (PCSK9) which is a serine protease enzyme primarily produced by the liver cells, can potentially be used as a targeting ligand. PCSK9 binds to the LDL-r on hepatocytes' surface, leading to endocytosis and endosomal degradation. High LDL-r expression, which is believed to meet the higher demand of the cholesterol and phospholipids to build proliferating cancer cell membrane, ensures selective uptake of the PCSK9 conjugated liposomes. In the present work, the PCSK9 conjugated liposomal system was developed to deliver paclitaxel (PTX) to cancer cells. The protein was conjugated by EDC and NHS in a two-step coupling reaction to the liposomes containing COOH-PEG-COOH lipid. Conjugation was confirmed by NMR, and liposomes were further characterized by SEM and zeta sizer. PCSK9-conjugated liposomes showed high encapsulation efficiency of 69.1% with a diameter of 90.0 ± 4.9 nm. Long-term stability (30 days) study (Zeta potential: -9.88) confirmed excellent constancy and significant drug retention (58.2%). Invitro cytotoxicity and targeting efficiency was explored using MTS assay in human embryonic kidney cells (HEK293), liver hepatocellular cells (HEPG2), and a human colon cancer cell line (HCT116) for 24 h. PCSK9 conjugated liposomes exhibited significantly higher growth inhibition than the unconjugated (control) liposomes in HCT116 cell line (p < 0.001). The novel PCSK9 conjugated liposomes presented potent and precise in vitro anticancer activity and, therefore, are suggested for the first time as a promising targeted delivery system for cancer treatment.
基于配体的靶向治疗方法针对的是明确过度表达于癌细胞表面的受体，是一种增强化疗疗效的有效药物输送方法。组织蛋白酶 K 前体转化酶亚基 9（PCSK9）是一种主要由肝细胞产生的丝氨酸蛋白酶，它有可能被用作靶向配体。PCSK9 与肝细胞表面的 LDL-r 结合，导致内吞作用和内体降解。高 LDL-r 表达水平，据信可以满足胆固醇和磷脂构建增殖癌细胞膜的更高需求，确保了 PCSK9 缀合脂质体的选择性摄取。在本工作中，开发了 PCSK9 缀合脂质体系统将紫杉醇（PTX）递送到癌细胞中。该蛋白通过 EDC 和 NHS 在两步偶联反应中与含有 COOH-PEG-COOH 脂质的脂质体结合。通过 NMR 确认了缀合，并且通过 SEM 和 zeta 粒度仪进一步对脂质体进行了表征。PCSK9 缀合脂质体显示出 69.1%的高包封效率，直径为 90.0±4.9nm。30 天的长期稳定性（Zeta 电位：-9.88）研究证实了其优异的稳定性和显著的药物保留率（58.2%）。通过 MTS 测定法在人胚肾细胞（HEK293）、肝肝细胞（HEPG2）和人结肠癌细胞系（HCT116）中进行了 24 小时的体外细胞毒性和靶向效率研究。PCSK9 缀合脂质体在 HCT116 细胞系中表现出比未缀合（对照）脂质体更高的生长抑制作用（p<0.001）。新型 PCSK9 缀合脂质体表现出强大而精确的体外抗癌活性，因此首次被提议作为癌症治疗的有前途的靶向递药系统。
