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Lipoprotein-Inspired Nanocarrier Composed of Folic Acid-Modified Protein and Lipids: Preparation and Evaluation of Tumor-Targeting Effect

Authors Han M, Ji X, Li J, Ge Z, Luo B, Zhou K, Wang Q, Sun X, Zhang W, Li J

Received 7 December 2019

Accepted for publication 15 April 2020

Published 14 May 2020 Volume 2020:15 Pages 3433—3445


Checked for plagiarism Yes

Review by Single anonymous peer review

Peer reviewer comments 2

Editor who approved publication: Dr Mian Wang

Mengmeng Han,1,2,* Xiaoman Ji,1,2,* Jianfei Li,1,2,* Zhiming Ge,1,2 Bin Luo,1,2 Kai Zhou,1,2 Qianqian Wang,1,2 Xin Sun,1,2 Wei Zhang,1,2 Jin Li1,2

1Department of Pharmacy, Xuzhou Medical University, Xuzhou 221004, Jiangsu, People’s Republic of China; 2Jiangsu Key Laboratory of New Drug Research and Clinical Pharmacy, Xuzhou Medical University, Xuzhou 221004, Jiangsu, People’s Republic of China

*These authors contributed equally to this work

Correspondence: Jin Li
Department of Pharmacy, Xuzhou Medical University, 209 Tongshan Road, Xuzhou 221004, People’s Republic of China
Tel/Fax +86 25 83271293

Background: Reconstituted lipoproteins (rLips) based on endogenous lipid nanostructures has been increasingly regarded as an excellent and promising antitumor drug delivery. However, some problems relating to the main component, apolipoprotein, for instance, rare source, unaffordable price, and low specificity of relevant receptor expression, become chief obstacles to its broad development and application.
Purpose: The primary aim of this study is to develop biomimetic rLips by utilizing folic acid (FA)-modified bovine serum albumin (BSA) as a replacement for apolipoprotein and demonstrate its tumor targeting and antitumor efficacy.
Methods: The amino groups of BSA were covalently conjugated with FA through the amide reaction. PTX-loaded nanostructured lipid carrier (termed as P-NLC) consisting of phospholipid, cholesteryl ester, triglyceride and cholesterol was prepared by the emulsification–evaporation method and utilized as the lipid core. FA-modified BSA (FA-BSA) was characterized for the protein substitute degree and attached with NLC by incubation-insert method to form the lipoprotein-mimic nanocomplex (termed as PFB-rLips). The morphology of nanoparticles was observed under transmission electron microscopy (TEM), and the particle size and zeta potential were determined using dynamic light scattering. In vitro release behavior of PTX from PFB-rLips was investigated with the dialysis method. Hemolysis tests were conducted to evaluate the biosecurity of PFB-rLips. Cell uptake and cytotoxicity assays were performed on human hepatocytes (LO2) and human hepatoma cells (HepG2). Tumor targeting was assessed using in vivo imaging system in H22 tumor-bearing mice model. Antitumor efficacy in vivo was investigated and compared between Taxol® (paclitaxel) formulation and PTX-incorporated nanoparticles in the same tumor model.
Results: A fixed molar ratio 50:1 of FA to BSA was chosen as the optimal input ratio based on the balance between appropriate degree of protein substitution and amphiphilicity of FA-BSA. The morphology of FB-rLips exhibited as a homogeneous spherical structure featured by lipid cores surrounded with a cloudy protein shell observed under TEM. The particle size, zeta potential and encapsulation efficiency were 174.6± 3.2 nm, − 17.26± 0.9 mV and 82.2± 2.4%, respectively. In vitro release behavior of PTX from PFB-rLips was slow and sustained. The uptake of FB-rLips was much higher in HepG2 cells than in LO2 cells. Furthermore, the uptake of FB-rLips was significantly higher than that of rLips without FA involved (termed as B-rLips) and NLC in HepG2 cells. Hemolysis and cytotoxicity assays showed good biocompatibility of FB-rLips. The internalization mechanism of FB-rLips mainly depended on clathrin-mediated and caveolin-mediated endocytosis coupling with energy consumption, and FA receptors expressed on tumor cells played a critical role in cellular uptake process. CCK-8 studies demonstrated that PFB-rLips exhibited significantly better tumor killing ability than Taxol® (paclitaxel) formulation in vitro. Moreover, FB-rLips produced more excellent tumor-targeting properties than NLC through in vivo imaging assays. On the basis of this, PTX-loaded FB-rLips also performed more remarkable anticancer activity than other therapy groups in H22 tumor-bearing mice.
Conclusion: FB-rLips would serve as a potential nanocarrier for improving tumor-targeting and therapeutic efficacy while reducing the side effects on normal tissues and organs.

Keywords: lipoprotein-inspired nanocarrier, BSA, folic acid, paclitaxel, tumor targeting

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