In this study, multifunctional theranostic agents for photoacoustic (PA), ultrasound (US), fluorescent imaging, and for therapeutic drug delivery were developed and tested. These agents consisted of a shell made from a biodegradable Poly(lactide-co-glycolic acid) (PLGA) polymer, loaded with perfluorohexane (PFH) liquid and gold nanoparticles (GNPs) in the core, and lipophilic carbocyanines fluorescent dye DiD and therapeutic drug Paclitaxel (PAC) in the shell. Their multifunctional capacity was investigated in an in vitro study. The PLGA/PFH/DiD-GNPs particles were synthesized by a double emulsion technique. The average PLGA particle diameter was 560 nm, with 50 nm diameter silica-coated gold nano-spheres in the shell. MCF7 human breast cancer cells were incubated with PLGA/PFH/DiDGNPs for 24 hours. Fluorescent and PA images were recorded using a fluorescent/PA microscope using a 1000 MHz transducer and a 532 nm pulsed laser. For the particle vaporization and drug delivery test, MCF7 cells were incubated with the PLGA/PFH-GNPs-PAC or PLGA/PFH-GNPs particles for 6, 12 and 24 hours. The effects of particle vaporization and drug delivery inside the cells were examined by irradiating the cells with a laser fluence of 100 mJ/cm2, and cell viability quantified using the MTT assay. The PA images of MCF7 cells containing PLGA/PFH/DiD-GNPs were spatially coincident with the fluorescent images, and confirmed particle uptake. After exposure to the PLGA/PFHGNP- PAC for 6, 12 and 24 hours, the cell survival rate was 43%, 38%, and 36% respectively compared with the control group, confirming drug delivery and release inside the cells. Upon vaporization, cell viability decreased to 20%. The particles show potential as imaging agents and drug delivery vehicles.
There is currently interest in the development of nanoemulsions as imaging and therapeutic agents, particularly perfluorohexane (PFH) droplets, whose amphiphilic shell protects drugs against physico-chemical and enzymatic degradation. When delivered to their target sites, these perfluorocarbon (PFC) droplets can vaporize upon laser excitation, efficiently releasing their drug payload and/or imaging tracers. Due to the optical properties of gold, coupling PFC droplets with gold nanoparticles significantly reduces the energy required for vaporization. In this work, nanoemulsions with a PFC core and Zonyl FSP surfactant shell were produced using sonication. Droplets were characterized in terms of size and morphology using high resolution fluorescence microscopy (i.e. total internal reflection fluorescence microscopy, TIRFM), fluorescence correlation spectroscopy (FCS), transmission electron microscopy (TEM), and light scattering techniques (i.e. dynamic light scattering, DLS). The ability of PFC droplets to vaporize was demonstrated using optical light microscopy.
Phase-change contrast agents consisting of a perfluorocarbon (PFC) liquid core stabilized by a lipid, protein, or polymer shell have been proposed for a variety of clinical applications. Previous work has demonstrated that vaporization can be induced by laser irradiation through optical absorbers incorporated inside the droplet. In this study, Poly-lactide-coglycolic acid (PLGA) particles loaded with PFC liquid and silica-coated gold nanoparticles (GNPs) were developed and characterized using photoacoustic (PA) methods. Microsized PLGA particles were loaded with PFC liquid and GNPs (14, 35, 55nm each with a 20nm silica shell) using a double emulsion method. The PA signal intensity and optical vaporization threshold were investigated using a 375 MHz transducer and a focused 532-nm laser (up to 450-nJ per pulse). The laser-induced vaporization threshold energy decreased with increasing GNP size. The vaporization threshold was 850, 690 and 420 mJ/cm2 for 5μm-sized PLGA particles loaded with 14, 35 and 55 nm GNPs, respectively. The PA signal intensity increased as the laser fluence increased prior to the vaporization event. This trend was observed for all particles sizes. PLGA particles were then incubated with MDA-MB-231 breast cancer cells for 6 hours to investigate passive targeting, and the vaporization of the PLGA particles that were internalized within cells. The PLGA particles passively internalized by MDA cells were visualized via confocal fluorescence imaging. Upon PLGA particle vaporization, bubbles formed inside the cells resulting in cell destruction. This work demonstrates that GNPs-loaded PLGA/PFC particles have potential as PA theranostic agents in PA imaging and optically-triggered drug delivery systems.