Drug Release from Phase-Changeable Nanodroplets Triggered by Low-Intensity Focused Ultrasound

Abstract

Background: As one of the most effective triggers with high tissue-penetrating capability and non-invasive feature, ultrasound shows great potential for controlling the drug release and enhancing the chemotherapeutic efficacy. In this study, we report, for the first time, construction of a phase-changeable drug-delivery nanosystem with programmable low-intensity focused ultrasound (LIFU) that could trigger drug-release and significantly enhance anticancer drug delivery. Methods: Liquid-gas phase-changeable perfluorocarbon (perfluoropentane) and an anticancer drug (doxorubicin) were simultaneously encapsulated in two kinds of nanodroplets. By triggering LIFU, the nanodroplets could be converted into microbubbles locally in tumor tissues for acoustic imaging and the loaded anticancer drug (doxorubicin) was released after the microbubble collapse. Based on the acoustic property of shell materials, such as shell stiffness, two types of nanodroplets (lipid-based nanodroplets and PLGA-based nanodroplets) were activated by different acoustic pressure levels. Ultrasound irradiation duration and power of LIFU were tested and selected to monitor and control the drug release from nanodroplets. Various ultrasound energies were introduced to induce the phase transition and microbubble collapse of nanodroplets in vitro (3 W/3 min for lipid nanodroplets; 8 W/3 min for PLGA nanodroplets). Results: We detected three steps in the drug-releasing profiles exhibiting the programmable patterns. Importantly, the intratumoral accumulation and distribution of the drug with LIFU exposure were significantly enhanced, and tumor proliferation was substantially inhibited. Co-delivery of two drug-loaded nanodroplets could overcome the physical barriers of tumor tissues during chemotherapy. Conclusion: Our study provides a new strategy for the efficient ultrasound-triggered chemotherapy by nanocarriers with programmable LIFU capable of achieving the on-demand drug release.

Keywords: Low-intensity focused ultrasound (LIFU); Perfluorocarbon nanodroplets; Programmable drug release; Ultrasound imaging.

Figure 1

Optical images of PLGA nanodroplets (A1) and lipid nanodroplets (B1) (presented as mean ± standard deviations (scale bar=10 μm); TEM image of PLGA nanodroplets (A2) and lipid nanodroplets (B2) (scale bar=200 nm); Size distribution of PLGA nanodroplets (A3) and lipid nanodroplets (B3).

Figure 2

Harmonic images (A), average ultrasound intensity values (B), average diameters (C) and optical images (D) of two types of nanodroplets after LIFU irradiation. (The LN 8 W, 3 min group: scale bar=50 μm; The other groups: scale bar=200 μm). Control: nanodroplet without ultrasound exposure.

Figure 3

A) Drug-releasing profiles triggered by LIFU under different conditions (black arrow: ultrasound administration); B) Cell uptake observed by confocal fluorescent microscopy (Dio: green, DOX: red, DAPI: blue). Scale bar=10 μm; C) Cell viability of various therapeutic agents (*p<0.05 vs the control group; ***p<0.001 vs the control group; #p<0.05 vs the LN-PN group; p<0.05 vs the LN-PN LIFU1 group; n=3); D) Cell apoptosis in different groups by LIFU treatment.

Figure 4

Ultrasound images (A) and average ultrasound intensity values (B) in tumors. Scale bar=0.3 cm. (C) in vivo distribution of accumulated DOX per gram tissue after intravenous injection of different formulations for 24 h. Distribution of nanodroplets labeled by DiR in vivo (D) and in different organs (E) (Control: no ultrasound radiation; 1: before ultrasound exposure; 2: the1^st treatment; 3: the 2^nd treatment) Scale bar=2.5 cm.

Figure 5

Schematic diagram of the release of nanodroplets complex triggered by LIFU radiation at different stages.

Figure 6

A) Tumor-volume curves after the injection of LN-PN followed by LIFU treatment in vivo; B) Cumulative survival of animals after intravenous administration of various formulations and LIFU irradiations; C) Histological characteristics of the MDA-MB 231 tumor tissues in different therapeutic groups evaluated by H&E staining, PCNA, TUNEL and CD31 staining. (×400) Scale bar=10 μm; D) Typical photographs of tumor-bearing nude mice and excised tumors from mice at the end of treatment (1: Control; 2: LN-PN; 3: LN-PN LIFU1; 4: LN-PN LIFU2) Scale bar=10 mm.