The role of ultrasound-driven microbubble dynamics in drug delivery : from microbubble fundamentals to clinical translation

In the last couple of decades, ultrasound-driven microbubbles have proven excellent candidates for local drug delivery applications. Besides being useful drug carriers, microbubbles have demonstrated the ability to enhance cell and tissue permeability and, as a consequence, drug uptake herein. Notwithstanding the large amount of evidence for their therapeutic efficacy, open issues remain. Because of the vast number of ultrasound- and microbubble-related parameters that can be altered and the variability in different models, the translation from basic research to (pre)clinical studies has been hindered. This review aims at connecting the knowledge gained from fundamental microbubble studies to the therapeutic efficacy seen in in vitro and in vivo studies, with an emphasis on a better understanding of the response of a microbubble upon exposure to ultrasound and its interaction with cells and tissues. More specifically, we address the acoustic settings and microbubble-related parameters (i.e., bubble size and physicochemistry of the bubble shell) that play a key role in microbubble cell interactions and in the associated therapeutic outcome. Additionally, new techniques that may provide additional control over the treatment, such as monodisperse microbubble formulations, tunable ultrasound scanners, and cavitation detection techniques, are discussed. An in-depth understanding of the aspects presented in this work could eventually lead the way to more efficient and tailored microbubble-assisted ultrasound therapy in the future

Sonoprinting of nanoparticle-loaded microbubbles : unraveling the multi-timescale mechanism

Ultrasound-triggered microbubble-assisted drug delivery is a promising tool for localized therapy. Several studies have shown the potential of nanoparticle-loaded microbubbles to effectively enhance the delivery of therapeutic agents to target tissue. We recently discovered that nanoparticle-carrying microbubbles can deposit the nanoparticles in patches onto cell membranes, a process which we termed 'sonoprinting'. However, the biophysical mechanisms behind sonoprinting are not entirely clear. In addition, the question remains how the ultrasound parameters, such as acoustic pressure and pulse duration, influence sonoprinting. Aiming for a better understanding of sonoprinting, this report investigates the behavior of nanoparticle-loaded microbubbles under ultrasound exposure, making use of three advanced optical imaging techniques with frame rates ranging from 5 frames per second to 10 million frames per second, to capture the biophysical cell-bubble interactions that occur on a multitude of timescales. We observed that non-spherically oscillating microbubbles release their nano particle payload in the first few cycles of ultrasound insonation. At low acoustic pressures, the released nano particles are transported away from the cells by microstreaming, which does not favor uptake of the nano particles by the cells. However, higher acoustic pressures ( > 300 kPa) and longer ultrasound pulses ( > 100 cycles) lead to rapid translation of the microbubbles, due to acoustic radiation forces. As a result, the released nanoparticles are transported along in the wake of the microbubbles, which eventually leads to the deposition of nanoparticles in elongated patches on the cell membrane, i.e. sonoprinting. We conclude that a sufficiently high acoustic pressure and long pulses are needed for sonoprinting of nanoparticles on cells

Sonoprinting liposomes on tumor spheroids by microbubbles and ultrasound

Ultrasound-triggered drug-loaded microbubbles have great potential for drug delivery due to their ability to locally release drugs and simultaneously enhance their delivery into the target tissue. We have recently shown that upon applying ultrasound, nanoparticle-loaded microbubbles can deposit nanoparticles onto cells grown in 2D monolayers, through a process that we termed "sonoprinting". However, the rigid surfaces on which cell monolayers are typically growing might be a source of acoustic reflections and aspherical microbubble oscillations, which can influence microbubble-cell interactions. In the present study, we aim to reveal whether sonoprinting can also occur in more complex and physiologically relevant tissues, by using free-floating 3D tumor spheroids as a tissue model. We show that both monospheroids (consisting of tumor cells alone) and cospheroids (consisting of tumor cells and fibroblasts, which produce an extracellular matrix) can be sonoprinted. Using doxorubicin-liposome-loaded microbubbles, we show that sonoprinting allows to deposit large amounts of doxorubicin-containing liposomes to the outer cell layers of the spheroids, followed by doxorubicin release into the deeper layers of the spheroids, resulting in a significant reduction in cell viability. Sonoprinting may become an attractive approach to deposit drug patches at the surface of tissues, thereby promoting the delivery of drugs into target tissues

Ultrasound-Responsive Cavitation Nuclei for Therapy and Drug Delivery

Therapeutic ultrasound strategies that harness the mechanical activity of cavitation nuclei for beneficial tissue bio-effects are actively under development. The mechanical oscillations of circulating microbubbles, the most widely investigated cavitation nuclei, which may also encapsulate or shield a therapeutic agent in the bloodstream, trigger and promote localized uptake. Oscillating microbubbles can create stresses either on nearby tissue or in surrounding fluid to enhance drug penetration and efficacy in the brain, spinal cord, vasculature, immune system, biofilm or tumors. This review summarizes recent investigations that have elucidated interactions of ultrasound and cavitation nuclei with cells, the treatment of tumors, immunotherapy, the blood–brain and blood–spinal cord barriers, sonothrombolysis, cardiovascular drug delivery and sonobactericide. In particular, an overview of salient ultrasound features, drug delivery vehicles, therapeutic transport routes and pre-clinical and clinical studies is provided. Successful implementation of ultrasound and cavitation nuclei-mediated drug delivery has the potential to change the way drugs are administered systemically, resulting in more effective therapeutics and less-invasive treatments

Temperature-induced self-assembly of the group B Streptococcus (GBS) fusion antigen GBS-NN

Group B Streptococcus (GBS) is a leading cause of serious bacterial neonatal infections worldwide, which provides an unmet medical need for a globally effective vaccine. The recombinant GBS fusion antigen GBS-NN contains the N-terminal regions of the GBS Rib and Alpha C proteins. It shows promising immunogenicity eliciting protective immunity in mice and encouraging results in early human clinical trials. Understanding the physical stability of GBS-NN containing conformational B-cell epitopes is crucial to ensure optimal vaccine stability, efficacy, and safety. We initially discovered that GBS-NN is prone to form higher-order structures at elevated temperatures. We therefore investigated the self-assembly behavior of GBS-NN and characterized the higher-order conformational structures as a function of temperature. In the native state, GBS-NN exists as a monomer and has a secondary structure containing alpha-helix and beta-sheet. Langmuir studies demonstrated that the native protein is highly surface-active and forms a monolayer film at the air-water interface because of its amphipathic properties. The conformational stability of GBS-NN was measured as a function of temperature. GBS-NN has an unusual thermal behavior with a phase transition of approximately 61 degrees C, which is not accompanied by any major changes in the secondary structure. However, the antigen showed irreversible self-assembly as a function of temperature into higher-order structures with a hydrodynamic diameter of approximately 100 nm. Cryo-transmission electron microscopy analyses demonstrated that these self-assemblies consist of vesicular, ring-like structures with a hollow aqueous interior. Therefore, GBS-NN is a physically stable monomeric protein but is prone to temperature-induced self-assembly above 61 degrees C

Ultrasound-driven microbubbles for local drug delivery

International audienceIn ultrasound-mediated drug delivery, bubbles decorated with a functional payload are convenient transport vehicles and offer highly localized release. These bubbles are traceable using ultrasound, can be activated on demand and protect their cargo from the natural clearing mechanisms of the body. In addition, microbubbles offer an active way to mechanically overcome biological barriers. Such microbubbles are therefore being extensively investigated for single cell, gene and cancer therapy. Experimental investigation of microbubble interactions with cell monolayers has led to the description of a phenomenon termed ?sonoprinting?: confocal imaging shows how exposure of the microbubbles to ultrasound can result in the deposition of their payload in patches onto the cell membrane. In parallel, investigation of single microbubbles with a combination of ultra-high-speed imaging at 10 million frames per second, and high-speed fluorescence imaging at 50,000 frames per second demonstrates how non-spherical bubble oscillations induce both, the release of the drug payload and its transport by microbubble-induced streaming. These observations are validated with a simple theoretical model based on the Rayleigh-Plesset equation. In order to reconcile these observations, we combine three optical imaging techniques to capture the bubble-cell interactions on timescales ranging from sub-microseconds to several seconds. We observe that non-spherically oscillating microbubbles release their nanoparticle payload in the first few ultrasound cycles. At low pressures, the released nanoparticles are transported away by microstreaming, as observed for single microbubbles. This configuration, however, does not favor of the nanoparticles uptake. Higher pressures (> 300 kPa) and longer ultrasound pulses (>100 cycles) lead to rapid translation of the microbubbles and transport of the released nanoparticles in the microbubbles wake, eventually leading to the deposition of nanoparticles in elongated patches onto the cell membrane

Evaluation of liposome-loaded microbubbles as a theranostic tool in a murine collagen-induced arthritis model

Rheumatoid arthritis (RA) is an autoimmune disease characterized by severe inflammation of the synovial tissue. Here, we assess the feasibility of liposome-loaded microbubbles as theranostic agents in a murine arthritis model. First, contrast-enhanced ultrasound (CEUS) was used to quantify neovascularization in this model since CEUS is well-established for RA diagnosis in humans. Next, the potential of liposome-loaded microbubbles and ultrasound (US) to selectively enhance liposome delivery to the synovium was evaluated with in vivo fluorescence imaging. This procedure is made very challenging by the presence of hard joints and by the limited lifetime of the microbubbles. The inflamed knee joints were exposed to therapeutic US after intravenous injection of liposome-loaded microbubbles. Loaded microbubbles were found to be quickly captured by the liver. This resulted in fast clearance of attached liposomes while free and long-circulating liposomes were able to accumulate over time in the inflamed joints. Our observations show that murine arthritis models are not well-suited for evaluating the potential of microbubble-mediated drug delivery in joints given: (i) restricted microbubble passage in murine synovial vasculature and (ii) limited control over the exact ultrasound conditions in situ given the much shorter length scale of the murine joints as compared to the therapeutic wavelength

Ultrasound-Responsive Cavitation Nuclei for Therapy and Drug Delivery

Therapeutic ultrasound strategies that harness the mechanical activity of cavitation nuclei for beneficial tissue bio-effects are actively under development. The mechanical oscillations of circulating microbubbles, the most widely investigated cavitation nuclei, which may also encapsulate or shield a therapeutic agent in the bloodstream, trigger and promote localized uptake. Oscillating microbubbles can create stresses either on nearby tissue or in surrounding fluid to enhance drug penetration and efficacy in the brain, spinal cord, vasculature, immune system, biofilm or tumors. This review summarizes recent investigations that have elucidated interactions of ultrasound and cavitation nuclei with cells, the treatment of tumors, immunotherapy, the blood–brain and blood–spinal cord barriers, sonothrombolysis, cardiovascular drug delivery and sonobactericide. In particular, an overview of salient ultrasound features, drug delivery vehicles, therapeutic transport routes and pre-clinical and clinical studies is provided. Successful implementation of ultrasound and cavitation nuclei-mediated drug delivery has the potential to change the way drugs are administered systemically, resulting in more effective therapeutics and less-invasive treatments.ImPhys/Medical Imagin