Ghaleb A. Husseini

Ultrasonic drug delivery – a general review

Ultrasound has an ever-increasing role in the delivery of therapeutic agents, including genetic material, protein and chemotherapeutic agents. Cavitating gas bodies, such as microbubbles, are the mediators through which the energy of relatively non-interactive pressure waves is concentrated to produce forces that permeabilise cell membranes and disrupt the vesicles that carry drugs. Thus, the presence of microbubbles enormously enhances ultrasonic delivery of genetic material, proteins and smaller chemical agents. Numerous reports show that the most efficient delivery of genetic material occurs in the presence of cavitating microbubbles. Attaching the DNA directly to the microbubbles, or to gas-containing liposomes, enhances gene uptake even further. Ultrasonic-enhanced gene delivery has been studied in various tissues, including cardiac, vascular, skeletal muscle, tumour and even fetal tissue. Ultrasonic-assisted delivery of proteins has found most application in transdermal transport of insulin. Cavitation events reversibly disrupt the structure of the stratus corneum to allow transport of these large molecules. Other hormones and small proteins could also be delivered transdermally. Small chemotherapeutic molecules are delivered in research settings from micelles and liposomes exposed to ultrasound. Cavitation appears to play two roles: it disrupts the structure of the carrier vesicle and releases the drug; and makes cell membranes and capillaries more permeable to drugs. There remains a need to better understand the physics of cavitation of microbubbles and the impact that such cavitation has on cells and drug-carrying vesicles.

Micelles and Nanoparticles for Ultrasonic Drug and Gene Delivery

Drug delivery research employing micelles and nanoparticles has expanded in recent years. Of particular interest is the use of these nanovehicles that deliver high concentrations of cytotoxic drugs to diseased tissues selectively, thus reducing the agents side effects on the rest of the body. Ultrasound, traditionally used in diagnostic medicine, is finding a place in drug delivery in connection with these nanoparticles. In addition to their non-invasive nature and the fact that they can be focused on targeted tissues, acoustic waves have been credited with releasing pharmacological agents from nanocarriers, as well as rendering cell membranes more permeable. In this article, we summarize new technologies that combine the use of nanoparticles with acoustic power both in drug and gene delivery. Ultrasonic drug delivery from micelles usually employs polyether block copolymers and has been found effective in vivo for treating tumors. Ultrasound releases drug from micelles, most probably via shear stress and shock waves from the collapse of cavitation bubbles. Liquid emulsions and solid nanoparticles are used with ultrasound to deliver genes in vitro and in vivo. The small packaging allows nanoparticles to extravasate into tumor tissues. Ultrasonic drug and gene delivery from nanocarriers has tremendous potential because of the wide variety of drugs and genes that could be delivered to targeted tissues by fairly non-invasive means.

Drug delivery in pluronic micelles: effect of high-frequency ultrasound on drug release from micelles and intracellular uptake

The effect of high-frequency ultrasound on doxorubicin (DOX) release from Pluronic micelles and intracellular DOX uptake was studied for promyelocytic leukemia HL-60 cells, ovarian carcinoma drug-sensitive and multidrug-resistant (MDR) cells (A2780 and A2780/ADR, respectively), and breast cancer MCF-7 cells. Cavitation events initiated by high-frequency ultrasound were recorded by radical trapping. The onset of transient cavitation and DOX release from micelles were observed at much higher power densities than at low-frequency ultrasound (20-100 kHz). Even a short (15-30 s) exposure to high-frequency ultrasound significantly enhanced the intracellular DOX uptake from PBS, RPMI 1640, and Pluronic micelles. The mechanisms of the observed effects are discussed.

Investigating the mechanism of acoustically activated uptake of drugs from Pluronic micelles

BackgroundThis paper examines the mechanism of ultrasonic enhanced drug delivery from Pluronic micelles. In previous publications by our group, fluorescently labeled Pluronic was shown to penetrate HL-60 cells with and without the action of ultrasound, while drug uptake was increased with the application of ultrasound.MethodsIn this study, the amount of uptake of two fluorescent probes, Lysosensor Green (a pH-sensitive probe) and Cell Tracker Orange CMTMR (a pH-independent probe), was measured in HL-60 and HeLa cells.ResultsThe results of our experiments show that the increase in drug accumulation in the cells as a result of ultrasonication is not due to an increase in endocytosis due to ultrasonication.ConclusionsWe hypothesize that sonoporation plays an important role in the acoustically activated drug delivery of chemotherapy drugs delivered from Pluronic micelles.

Kinetics of ultrasonic release of doxorubicin from pluronic P105 micelles

The aim of this research was to measure and model the kinetics of acoustic release and subsequent re-encapsulation of Doxorubicin (DOX) from Pluronic P105 micelles. A fluorescence detection ultrasound exposure chamber was used. Experimental data showed that no significant release was observed when DOX loaded in Pluronic P105 micelles was exposed to ultrasound for less than 0.1 s at a power density of 58 mW/cm2 and a frequency of 20 kHz. Above this threshold, the amount of release was shown to increase as the pulse length increased up to 0.6 s. The same experiments showed that it requires at least 0.1 s of no ultrasound for measurable re-encapsulation to occur. Release and re-encapsulation are completed within about 0.6 s of the beginning of the ON and OFF phases of pulsed ultrasound. Several physical models and their corresponding mathematical solutions were analyzed to see which most closely fit the data. The model of zero-order release with first-order re-encapsulation appears to represent data from this polymeric system better than other models. This technique has possible applications in site-specific chemotherapy.

Ultrasonic release of doxorubicin from Pluronic P105 micelles stabilized with an interpenetrating network of N,N-diethylacrylamide.

Pluronic P105 micelles sequester hydrophobic drugs and release them upon insonation with low frequency ultrasound; however these micelles dissolve relatively quickly upon dilution. The objective of this research was to determine whether stabilization of these micelles would compromise their ability to sequester and release drug. P105 micelles were stabilized with an interpenetrating network of poly (N,N-diethylacrylamide), and ultrasonically-activated release of doxorubicin (Dox) was measured by a fluorescence technique. Results showed that stabilized micelles sequestered the Dox and released it upon insonation at 70 kHz. The amount released was not significantly different from that released from P105 micelles (P=0.481), and the drug re-encapsulation upon cessation of insonation was complete. This system has potential for controlled drug delivery to insonated tissues in vivo.

DNA damage induced by micellar-delivered doxorubicin and ultrasound: comet assay study

To minimize adverse side effects of chemotherapy, we have developed a micellar drug carrier that retains hydrophobic drugs, and then releases the drug by ultrasonic stimulation. This study investigated the DNA damage induced by doxorubicin (DOX) delivered to human leukemia (HL-60) cells from pluronic P-105 micelles with and without the application of ultrasound. The comet assay was used to quantify the amount of DNA damage. No significant DNA damage was observed when the cells were treated with 0.1, 1 and 10 wt% P-105 with or without ultrasound (70 kHz, 1.3 W/cm(2)) for 1 h or for up to 3 h in 10 wt% P-105. However, when cells were incubated with 10 microg/ml free DOX for up to 9 h, DNA damage increased with incubation time (P=0.0011). Exposure of cells to the same concentration of DOX in the presence of 10-wt% P-105 showed no significant DNA damage for up to 9 h of incubation. However, when ultrasound was applied, a rapid and significant increase in DNA damage was observed (P=0.0001). The application of ultrasound causes the release of DOX from micelles or causes the HL-60 cells to take up the micelle encapsulated DOX. Our experiments indicated that the combination of DOX, ultrasound and pluronic P105 causes the largest DNA damage to HL-60 cells. We believe that this technique can be used for controlled drug delivery.

Ultrasonic-Activated Micellar Drug Delivery for Cancer Treatment

The use of nanoparticles and ultrasound in medicine continues to evolve. Great strides have been made in the areas of producing micelles, nanoemulsions, and solid nanoparticles that can be used in drug delivery. An effective nanocarrier allows for the delivery of a high concentration of potent medications to targeted tissue while minimizing the side effect of the agent to the rest of the body. Polymeric micelles have been shown to encapsulate therapeutic agents and maintain their structural integrity at lower concentrations. Ultrasound is currently being used in drug delivery as well as diagnostics, and has many advantages that elevate its importance in drug delivery. The technique is noninvasive, thus no surgery is needed; the ultrasonic waves can be easily controlled by advanced electronic technology so that they can be focused on the desired target volume. Additionally, the physics of ultrasound are widely used and well understood; thus ultrasonic application can be tailored towards a particular drug delivery system. In this article, we review the recent progress made in research that utilizes both polymeric micelles and ultrasonic power in drug delivery.

Intracellular uptake of Pluronic copolymer: effects of the aggregation state

The effect of the Pluronic P-105 aggregation state on its uptake by HL-60 cells was studied by flow cytometry, fluorescence spectroscopy, and confocal and fluorescence microscopy using a fluorescently labeled Pluronic P105. In the low concentration region below the critical micelle concentration (CMC), Pluronic uptake was proportional to the concentration in the incubation medium. The proportionality broke sharply above the CMC, revealing a less efficient intracellular uptake of Pluronic micelles than that of unimers. The data suggested that Pluronic micelles were internalized via fluid-phase endocytosis while unimers were internalized via diffusion through plasma membranes. Based on the above findings, the shielding effect of Pluronic micelles on drug intracellular uptake was explained.

Phase transitions of nanoemulsions using ultrasound: experimental observations.

The ultrasound-induced transformation of perfluorocarbon liquids to gases is of interest in the area of drug and gene delivery. In this study, three independent parameters (temperature, size, and perfluorocarbon species) were selected to investigate the effects of 476-kHz and 20-kHz ultrasound on nanoemulsion phase transition. Two levels of each factor (low and high) were considered at each frequency. The acoustic intensities at gas bubble formation and at the onset of inertial cavitation were recorded and subsequently correlated with the acoustic parameters. Experimental data showed that low frequencies are more effective in forming and collapsing a bubble. Additionally, as the size of the emulsion droplet increased, the intensity required for bubble formation decreased. As expected, perfluorohexane emulsions require greater intensity to form cavitating bubbles than perfluoropentane emulsions.