Department of Pharmacy, DFM Scarl - University of Naples "Federico II", 80134 Naples, Italy.
Université François-Rabelais, EA 6295 «Nanomédicaments et Nanosondes», Tours F-37200, France.
Int J Pharm. 2015 Jul 5;488(1-2):59-66. doi: 10.1016/j.ijpharm.2015.04.039. Epub 2015 Apr 17.
The present work investigates in vitro the delivery of the anticancer drug mitoxantrone (MTX) to HeLa cancer cells by means of polyethylene glycol (PEG) liposomes functionalized with the novel cell penetrating peptide gH625. This hydrophobic peptide enhances the delivery of doxorubicin (Doxo) to the cytoplasm of cancer cells, while the mechanism of this enhancement has not yet been understood. Here, in order to get a better insight into the role of gH625 on the mechanism of liposome-mediated drug delivery, we treated HeLa cells with liposomes functionalized with gH625 and loaded with MTX; functionalized and not liposome were characterized in terms of their physico-chemical properties and drug release kinetics. To quantify the MTX uptake and to study the subcellular drug distribution and interaction, we took advantage of the intrinsic fluorescence of MTX and of the fluorescence-based techniques like fluorescence-activated cell sorting (FACS) and confocal spectral imaging (CSI). FACS data confirmed that gH625 increases the total intracellular MTX content. CSI data indicated that when liposomes are decorated with gH625 an enhanced staining of the internalized drug is observed mainly in hydrophobic regions of the cytoplasm, where the increased presence of an oxidative metabolite of the drug is observed. The cytotoxicity on HeLa cell line was higher for functionalized liposomes within 4-6h of treatment. To summarise, the MTX delivery with gH625-decorated nanoliposomes enhances the quantity of both the intracellular drug and of its oxidative metabolite and contributes to higher anticancer efficacy of the drug at the delay of 4-6h.
本研究旨在通过聚乙二醇（PEG）脂质体传递抗癌药物米托蒽醌（MTX），并利用新型穿透肽 gH625 对其进行功能化，以研究其对 HeLa 癌细胞的体外递送。该疏水肽可增强阿霉素（Doxo）向癌细胞细胞质的递送，但其增强机制尚不清楚。为了更好地了解 gH625 在脂质体介导的药物递送机制中的作用，我们用 gH625 功能化的载有 MTX 的脂质体处理 HeLa 细胞；对功能化和非脂质体的性质及其药物释放动力学进行了表征。为了定量摄取 MTX，并研究细胞内药物分布和相互作用，我们利用了 MTX 的固有荧光和基于荧光的技术，如荧光激活细胞分选（FACS）和共聚焦光谱成像（CSI）。FACS 数据证实，gH625 增加了总细胞内 MTX 含量。CSI 数据表明，当脂质体被 gH625 修饰时，可观察到内化药物的荧光强度增强，主要集中在细胞质的疏水区，在这些区域中观察到药物的氧化代谢物的含量增加。在 4-6 小时的治疗期间，功能化脂质体对 HeLa 细胞系的细胞毒性更高。总之，用 gH625 修饰的纳米脂质体递送 MTX 可增加细胞内药物及其氧化代谢物的数量，并有助于在 4-6 小时的延迟时间内提高药物的抗癌疗效。
