Single peptide ligand-functionalized uniform hollow mesoporous silica nanoparticles achieving dual-targeting drug delivery to tumor cells and angiogenic blood vessel cells

Abstract

Background

The purpose of this study was to construct hollow mesoporous silica nanoparticles (HMSN) decorated with tLyp-1 peptide (tHMSN) for dual-targeting drug delivery to tumor cells and angiogenic blood vessel cells.

Methods

HMSN were synthesized de novo using a novel cationic surfactant-assisted selective etching strategy and were then modified with tLyp-1. Multiple methods, including transmission electron microscopy, X-ray photoelectron spectroscopy, thermogravimetric analysis, bicinchoninic acid assay, and nitrogen adsorption and desorption isotherms, were used to characterize the tHMSN. Doxorubicin were chosen as the model cargo, and the uptake of doxorubicin-loaded tHMSN into MDA-MB-231 cells and human umbilical vein endothelial cells (HUVECs), as models of tumor cells and tumor neovascular endothelial cells, respectively, were observed and detected by confocal laser scanning microscopy and flow cytometry. An in vitro pharmacodynamic study and a study of the mechanism via which the nanoparticles were endocytosed were also performed.

Results

HMSN with a highly uniform size and well oriented mesopores were synthesized. After tHMSN were characterized, enhanced uptake of the cargo carried by tHMSN into MDA-MB-231 cells and HUVECs compared with that of their unmodified counterparts was validated by confocal laser scanning microscopy and flow cytometry at the qualitative and quantitative levels, respectively. Further, the pharmacodynamic study suggested that, compared with their unmodified counterparts, doxorubicin-loaded tHMSN had an enhanced inhibitory effect on MDA-MB-231 cells and HUVECs in vitro. Finally, a preliminary study on the mechanism by which the nanoparticles were endocytosed indicated that the clathrin-mediated endocytosis pathway has a primary role in the transport of tHMSN into the cytoplasm.

Conclusion

tHMSN might serve as an effective active targeting nanocarrier strategy for anti-mammary cancer drug delivery.

Keywords:

• hollow mesoporous silica nanoparticles
• tLyp-1 peptide
• drug delivery
• anti-mammary cancer

Supplementary material

Detailed description of methods used to characterize the nanoparticles

The particle hydrodynamic size and zeta potential of hollow mesoporous silica nanoparticles (HMSN), tLyp-1 and polyethylene glycol co-modified HMSN (tHMSN), and polyethylene glycol-modified HMSN (pHMSN) were determined by dynamic light scattering analysis using a zeta potential/particle sizer (Nicomp 380, Particle Sizing Systems, Horiba Scientific, Edison, NJ, USA). Morphological examination of HMSN was performed using a transmission electron microscope (Tecnai G-20, FEI, Hillsboro, NJ, USA). Nitrogen adsorption–desorption isotherms were measured at 77 K using a surface area and porosity analyzer (ASAP2020, Micromeritics Instrument Corporation, Norcross, GA, USA). The specific surface area was calculated by the Brunauer–Emmett–Teller method and the pore size distribution was calculated using the Barrett–Joyner–Halenda model.

Bicinchoninic acid assay (BCA), X-ray photoelectron spectroscopy, and thermogravimetric analysis were used to confirm conjugation of tLyp-1 and polyethylene glycol on the surface of the tHMSN. Qualitative assessment of tLyp-1 bound on tHMSN was determined by BCA. Briefly, the BCA working solution was prepared by mixing reagent A with reagent B (reagent A:B 50:1, v/v) for further use. Next, 20 μL of tHMSN (HMSN and pHMSN acted as control) were added into a 96-well plate, followed by incubation with 200 μL of BCA working solution. After 30 minutes at 60°C, the assay was carried out at 570 nm using a microplate reader (Synergy TM2, Bio-Tek Instruments Inc, Winooski, VT, USA). The nanoparticle samples were also lyophilized (0.180 mbar vacuum, −50°C) using a freeze dryer (Alpha 2–4, Labconco, Kansas City, MO, USA) and then analyzed by X-ray photoelectron spectroscopy (Escalab 250Xi, Thermo Fisher Scientific, Waltham, MA, USA) to determine the surface composition. Further, the samples were subjected to thermogravimetric analysis (SDT-2960, TA Instruments, New Castle, DE, USA) in a controlled atmosphere of nitrogen from 90°C up to 590°C at a constant heating rate of 10°C per minute.

Acknowledgments

We acknowledge the National Natural Science Foundation of China (No 81341134) for financial support.

Disclosure

The authors report no conflicts of interest in this work.