CNC - Center for Neuroscience and Cell Biology (CNC), University of Coimbra, Coimbra, Portugal.
Institute for Interdisciplinary Research (IIIUC), University of Coimbra, Coimbra, Portugal.
Int J Nanomedicine. 2021 May 18;16:3385-3405. doi: 10.2147/IJN.S302288. eCollection 2021.
Hepatocellular carcinoma (HCC) is one of the main causes of cancer-related death. Sorafenib, which is the first-line therapy for this disease, is associated with reduced therapeutic efficacy that could potentially be overcome by combination with selumetinib. In this context, the main goal of this work was to develop a new nanosystem, composed of a polymeric core coated by a lipid bilayer containing the targeting ligand GalNAc, to specifically and efficiently co-deliver both drugs into HCC cells, in order to significantly increase their therapeutic efficacy.
The physicochemical characterization of hybrid nanosystems (HNP) and their components was performed by dynamic light scattering, zeta potential, matrix-assisted laser desorption ionization - time of flight mass spectroscopy, and transmission electron microscopy. Cellular binding, uptake and specificity of HNP were evaluated through flow cytometry and confocal microscopy. The therapeutic activity was evaluated namely through: cell viability by the Alamar Blue assay; cell death by flow cytometry using FITC-Annexin V; caspases activity by luminescence; mitochondrial membrane potential by flow cytometry; and molecular target levels by Western blot.
The obtained data show that these hybrid nanosystems present high stability and loading capacity of both drugs, and suitable physicochemical properties, namely in terms of size and surface charge. Moreover, the generated formulation allows to circumvent drug resistance and presents high specificity, promoting great cell death levels in HCC cells, but not in non-tumor cells. This potentiation of the antitumor effect of co-loaded drugs was carried out by an increased programmed cell death, being associated with a strong reduction in the mitochondrial membrane potential, a significant increase in the activity of caspases 3/7 and caspase 9, and much greater number of annexin V-positive cells.
The developed formulation resulted in a high and synergistic antitumor effect, revealing a translational potential to improve therapeutic approaches against HCC.
肝细胞癌（HCC）是癌症相关死亡的主要原因之一。索拉非尼是治疗这种疾病的一线药物，但疗效降低，与 selumetinib 联合使用可能会克服这种情况。在这种情况下，这项工作的主要目标是开发一种新的纳米系统，该系统由聚合物核组成，表面覆盖有脂质双层，其中包含靶向配体 GalNAc，以将两种药物特异性和有效地递送到 HCC 细胞中，从而显著提高其治疗效果。
通过动态光散射、zeta 电位、基质辅助激光解吸电离 - 飞行时间质谱和透射电子显微镜对杂交纳米系统（HNP）及其成分进行了理化特性表征。通过流式细胞术和共聚焦显微镜评估了 HNP 的细胞结合、摄取和特异性。通过以下方法评估治疗活性：通过 Alamar Blue 测定评估细胞活力；通过流式细胞术使用 FITC-Annexin V 评估细胞死亡；通过发光评估半胱天冬酶活性；通过流式细胞术评估线粒体膜电位；通过 Western blot 评估分子靶标水平。
获得的数据表明，这些杂交纳米系统具有高稳定性和两种药物的高载药量，以及合适的理化性质，特别是在粒径和表面电荷方面。此外，所产生的制剂可以规避药物耐药性，并具有高特异性，可促进 HCC 细胞中高水平的细胞死亡，但对非肿瘤细胞没有影响。这种共载药物的抗肿瘤作用增强是通过程序性细胞死亡的增加来实现的，与线粒体膜电位的强烈降低、半胱天冬酶 3/7 和半胱天冬酶 9 的活性显著增加以及 Annexin V 阳性细胞数量的增加有关。
所开发的制剂具有高协同抗肿瘤作用，显示出改善 HCC 治疗方法的转化潜力。
