Theranostics 2018; 8(7):1892-1910. doi:10.7150/thno.22386

Research Paper

Cell-penetrating Peptide-modified Targeted Drug-loaded Phase-transformation Lipid Nanoparticles Combined with Low-intensity Focused Ultrasound for Precision Theranostics against Hepatocellular Carcinoma

Hongyun Zhao1,2*, Meng Wu3*, Leilei Zhu2, Yi Tian4, Mingxing Wu5, Yizhen Li2, Liming Deng2, Wei Jiang2,6, Wei Shen1, Zhigang Wang2, Zhechuan Mei1,2, Pan Li2,7, Haitao Ran2,7, Zhiyi Zhou2,8✉, Jianli Ren2,7✉

1. Department of Gastroenterology, The Second Affiliated Hospital of Chongqing Medical University, Chongqing400010, P.R. China.
2. Chongqing Key Laboratory of Ultrasound Molecular Imaging, The Second Affiliated Hospital of Chongqing Medical University, Chongqing400016, P.R. China.
3. Department of Ultrasound, Zhongnan Hospital of Wuhan University, Wuhan, Hubei 430071, China.
4. Department of Plastic Surgery, The Second Affiliated Hospital of Chongqing Medical University, Chongqing400010, P.R. China.
5. Department of Ophthalmology, The Second Affiliated Hospital of Chongqing Medical University, Chongqing400010, P.R. China.
6. Department of Radiology, The Second Affiliated Hospital of Chongqing Medical University, Chongqing 400010, China.
7. Department of Ultrasound, The Second Affiliated Hospital of Chongqing Medical University, Chongqing 400010, China.
8. Department of Geriatrics, Chongqing General Hospital, Chongqing 400014, China.
* Co-first Authors

Abstract

Objective: Prepare a multifunctional ultrasound molecular probe, hyaluronic acid-mediated cell-penetrating peptide-modified 10-hydroxycamptothecin-loaded phase-transformation lipid nanoparticles (HA/CPPs-10-HCPT-NPs), and to combine HA/CPPs-10-HCPT-NPs with low-intensity focused ultrasound (LIFU) for precision theranostics against hepatocellular carcinoma (HCC).

Methods: HA/CPPs-10-HCPT-NPs were prepared using thin-film dispersion, ultrasound emulsification, and electrostatic effects. HA/CPPs-10-HCPT-NPs were characterized for particle size, zeta potential, encapsulation efficiency and drug-loading efficiency. In vitro, HA/CPPs-10-HCPT-NPs were tested for acoustic droplet vaporization (ADV) at different time points/acoustic intensities; the ability of HA/CPPs-10-HCPT-NPs to target SMMC-7721 cells was detected by confocal laser scanning microscopy (CLSM); the penetrating ability of CG-TAT-GC-modified NPs was verified by CLSM in a 3D multicellular tumor spheroid (MCTS) model; the effect of HA/CPPs-10-HCPT-NPs combined with LIFU on killing SMMC-7721 cells was measured by CCK-8 and flow cytometry. In vivo, the tumor-target efficiency of HA/CPPs-10-HCPT-NPs was evaluated by a small-animal fluorescence imaging system and CLSM; the enhanced ultrasound imaging efficiency of HA/CPPs-10-HCPT-NPs combined with LIFU was measured by an ultrasound imaging analyzer; the therapeutic effect of HA/CPPs-10-HCPT-NPs combined with LIFU was evaluated by tumor volume, tumor inhibition rate, and staining (hematoxylin and eosin (H & E), proliferating cell nuclear antigen (PCNA) and TUNEL).

Results: Mean particle size and mean zeta potential of HA/CPPs-10-HCPT-NPs were 284.2±13.3 nm and - 16.55±1.50 mV, respectively. HA/CPPs-10-HCPT-NPs could bind to SMMC-7721 cells more readily than CPPs-10-HCPT-NPs. Penetration depth into 3D MCTS of HA/CPPs-10-HCPT-NPs was 2.76-fold larger than that of NPs without CG-TAT-GC. HA/CPPs-10-HCPT-NPs could enhance ultrasound imaging by undergoing ADV triggered by LIFU. HA/CPPs-10-HCPT-NPs+LIFU group demonstrated significantly higher efficiency of anti-proliferation and apoptosis percentage than all other groups. In mouse liver tumor xenografts, HA/CPPs-10-HCPT-NPs could target tumor sites and enhance ultrasound imaging under LIFU. HA/CPPs-10-HCPT-NPs+LIFU group had a significantly smaller tumor volume, lower proliferative index (PI), and higher tumor inhibition and apoptotic index (AI) than all other groups.

Conclusions: Combined application of HA/CPPs-10-HCPT-NPs and LIFU should be a valuable and promising strategy for precise HCC theranostics.

Keywords: Hepatocellular carcinoma, cell-penetrating peptide, phase-transformation, low-intensity focus ultrasound, ultrasound molecular imaging, theranostics

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How to cite this article:
Zhao H, Wu M, Zhu L, Tian Y, Wu M, Li Y, Deng L, Jiang W, Shen W, Wang Z, Mei Z, Li P, Ran H, Zhou Z, Ren J. Cell-penetrating Peptide-modified Targeted Drug-loaded Phase-transformation Lipid Nanoparticles Combined with Low-intensity Focused Ultrasound for Precision Theranostics against Hepatocellular Carcinoma. Theranostics 2018; 8(7):1892-1910. doi:10.7150/thno.22386. Available from http://www.thno.org/v08p1892.htm