Current Pharmaceutical Design - Volume 18, Issue 15, 2012

Microbubbles have driven the field of diagnostic ultrasound to the cusp of major change. In the last decade, drug companies, ultrasound scanner manufacturers, and academic centers have invested manpower and funding in developing efficacious ultrasound contrast agents and new contrast-specific imaging modalities. In addition, new generation of targeted microbubbles have not only enhanced diagnostic accuracy but also therapeutic effectiveness by providing a new “vehicle” of gene and drug delivery. This special issue of Current Pharmaceutical Design aims to present the engineering and clinical aspects of microbubbles, highlighting the advances and perspectives in their diagnostic and therapeutic applications. Microbubbles have been the subject of numerous experiments and theoretical analysis over the past 100 years. From the theories on acoustic waves and the discovery of ultrasound, to the genesis of microbubbles around 1968, the magnificent story of these “magical” bubbles is briefly traced through the years by the article of Karamanou et al. [1]. The key physical principles which determine how microbubbles and ultrasound interact are thoroughly discussed by Azmin et al. [2]. Furthermore, the implications for microbubbles' design, preparation and exploitation are presented [2]. Further to the understanding of the physical processes underlying the microbubbles' stability and acoustic behavior, efforts are also now put into the study of chemical and biological features of multifunctional lipid, protein, or polymer shelled microbubbles. A number of methods of preparation of “smart” microbubbles for ultrasound image-guided therapy have been recently developed. Cavalieri et al., in their review article [3], provide an in-depth discussion of different approaches utilized in preparing multifunctional microbubbles. In vitro and in vivo assessment of microbubbles' stability and activity is discussed, with a particular emphasis on the emerging applications of microbubbles for the multiple imaging modalities, the effective opening of blood brain barrier, and for the therapeutic treatment of antimicrobial films [3]. Advances in microbubbles engineering led to a new generation of contrast agents; the “Phasechange contrast agents” (PCCAs). PCCAs for ultrasound-based applications can provide novel ways of approaching diagnostic and therapeutic techniques, beyond what is possible with traditional microbubbles and liquid emulsions. The history of PCCAs, their basic physical properties as well as factors that influence their design are discussed by Sheeran and Dayton [4]. Microbubbles are now used in several diagnostic and therapeutic interventions. In this issue, the application of microbubbles in cardiovascular, cancer, cerebrovascular and liver diseases is discussed by experts in each field....

Microbubbles were the subject of numerous experiments and theoretical analysis over the past 100 years. Linked with the discovery and evolution of ultrasound, they were considered to be excellent echoenhancers. From the theories on acoustic waves in antiquity and the discovery of ultrasound, to the genesis of microbubbles around 1968, in our paper we trace the magnificent story of these “magical” bubbles.

Ultrasound contrast agents consisting of gas microbubbles stabilised by a polymer or surfactant coating have been in clinical use for several decades. Research into the biomedical uses of microbubbles, however, remains a highly active and growing field. This is largely due to their considerable versatility and the wide range of applications for which they have demonstrated potential benefits. In addition to contrast enhancement, diagnostic applications include: perfusion mapping and quantification and molecular imaging. In drug and gene therapy microbubbles can be used as vehicles which are inherently traceable in vivo and can provide both targeted and controlled release. In addition, the dynamic behaviour of the microbubbles in response to ultrasound excitation contributes to the therapeutic process. At low intensities microbubbles have been shown to mediate reversible enhancement of cell and endothelial permeability, including temporary opening of the blood brain barrier. At higher intensities they have been used as means of increasing the efficiency of thrombolysis, high-intensity focused ultrasound (HIFU) surgery and lithotripsy. The aim of this review is to describe the key physical principles which determine how microbubbles and ultrasound interact and the implications for their design, preparation and exploitation in diagnostic and therapeutic applications.

Microbubbles (MBs) are ultrasound responsive colloidal particles with a strong potential to become theranostic agents, combining the contrast agent activity with therapeutic functionality. In the last decades, MBs have played a significant role as ultrasound contrast agents in diagnostic imaging. MBs have also shown great potential in applications such as molecular imaging, drug delivery, gene therapy and sonothrombolysis. A full understanding of all physical processes underlying the MBs' stability and acoustic behavior is available in the literature. Efforts have been now addressed to the study of chemical and biological features of multifunctional lipid, protein, or polymer shelled MBs. A number of methods of preparation of “smart” MBs for ultrasound image-guided therapy have been recently developed. In this review, different approaches utilized in preparing multifunctional MBs are discussed with specific attention to the current strategies adopted to design MBs with specialized functions. In vitro / in vivo assessment of MBs' stability and activity will be discussed with a particular emphasis on the emerging applications of MBs for the multiple imaging modalities, the effective opening of blood brain barrier, BBB, and for the therapeutic treatment of antimicrobial films.

Phase-change contrast agents (PCCAs) for ultrasound-based applications have resulted in novel ways of approaching diagnostic and therapeutic techniques beyond what is possible with microbubble contrast agents and liquid emulsions. When subjected to sufficient pressures delivered by an ultrasound transducer, stabilized droplets undergo a phase-transition to the gaseous state and a volumetric expansion occurs. This phenomenon, termed acoustic droplet vaporization, has been proposed as a means to address a number of in vivo applications at the microscale and nanoscale. In this review, the history of PCCAs, physical mechanisms involved, and proposed applications are discussed with a summary of studies demonstrated in vivo. Factors that influence the design of PCCAs are discussed, as well as the need for future studies to characterize potential bioeffects for administration in humans and optimization of ultrasound parameters.

Ultrasound targeted microbubble destruction (UTMD) has evolved as a novel system for non-invasive, organ- and tissuespecific drug and gene delivery. Initially developed as ultrasound contrast agents, microbubbles (MBs) have increasingly gained attention for their ability to directly deliver different classes of bioactive substances (e.g. genes, drugs, proteins, gene silencing constructs) to various organ systems and tumors. Bioactive substances can be attached to or incorporated in the microbubble shells. Applying ultrasound at their resonance frequency, microbubbles oscillate. When using higher ultrasound energies, oscillation amplitudes increase, finally resulting in microbubble destruction. This leads to increased capillary and cell membrane permeability in the immediate vicinity of the ruptured MBs, thus facilitating tissue and cell penetration of co-administered or loaded bioactive substances. Numerous proof of principle studies have been performed, demonstrating the broad potential of UTMD as a site-specific, non-invasive therapeutic tool, delivering microbubble payload to various target tissues and organ systems or facilitating uptake of bioactive substances into tissues or cells. This review focuses on current in vivo studies and therapeutic approaches of UTMD. Promising results give hope for future clinical applications of this novel non-viral vector system. Nevertheless, several limitations remain, which will also be discussed in this review article.

Ultrasound is one of the workhorses in clinical cancer diagnosis. In particular, it is routinely used to characterize lesions in liver, urogenital tract, head and neck and soft tissues. During the last years image quality steadily improved, which, among others, can be attributed to the development of harmonic image analysis. Microbubbles were introduced as intravascular contrast agents and can be detected with superb sensitivity and specificity using contrast specific imaging modes. By aid of these unspecific contrast agents tissues can be characterised regarding their vascularity. Antibodies, peptides and other targeting moieties were bound to microbubbles to target sites of angiogenesis and inflammation intending to get more disease-specific information. Indeed, many preclinical studies proved the high potential of targeted ultrasound imaging to better characterize tumors and to more sensitively monitor therapy response. Recently, first targeted microbubbles had been developed that meet the pharmacological demands of a clinical contrast agent. This review articles gives an overview on the history and current status of targeted ultrasound imaging of cancer. Different imaging concepts and contrast agent designs are introduced ranging from the use of experimental nanodroplets to agents undergoing clinical evaluation. Although it is clear that targeted ultrasound imaging works reliably, its broad acceptance is hindered by the user dependency of ultrasound imaging in general. Automated 3D-scanning techniques – like being used for breast diagnosis - and novel 3D transducers will help to make this fascinating method clinical reality.

Contrast echocardiography is considered by many as the Holy Grail for the assessment of heart disease, including coronary artery disease, structural abnormalities and cardiomyopathies. Although contrast agents have been severely criticized at times, they are now officially implemented into an extended variety of clinical applications, as illustrated by the American and European echocardiographic societies published consensus statements. The current review highlights contrast major clinical applications under the light of current literature data.

Atherosclerotic cardiovascular disease (CVD), primarily manifested as heart attacks and strokes, remains the main cause of death in the developed countries and is rapidly increasing in the developing world. Early detection and aggressive treatment of hidden (asymptomatic) atherosclerotic plaques that cause heart attack or stroke are most needed. However, existing clinical tools are not sufficient to address this need. Intravascular ultrasound (IVUS) is a catheter-based medical imaging tool that is capable of providing cross-sectional images of arteries. It is by far the most powerful clinical tool available for characterization of atherosclerotic plaques. However, existing IVUS is unable to detect plaque inflammation which is a key factor in complications of atherosclerotic plaques. Contrast enhanced IVUS (CE-IVUS) for detection of Vasa Vasorum (VV), microvessles that feed the vessel wall, can indirectly image plaque inflammation and thereby significantly increase the diagnostic power of IVUS. Several studies have shown that the density of VV in the atherosclerotic plaques is strongly correlated with the intensity of plaque inflammation and related processes which lead to plaque destabilization and rupture (the Vulnerable Plaque). Therefore the detection and measurement of VV in plaque, and leakage of blood from VV into plaques using CE-IVUS, can enable the development of an index for plaque vulnerability. In this paper, we present a review of our original work on coronary VV imaging, discuss subsequent reports by others, and also present the latest on the detection of VV based on CE-IVUS.

The treatment of Aortic Aneurysm disease is a growing procedure due to increase of life expectancy in Western Countries and relative incidence. In the past ten years we observed a progressive growth of endovascular over open surgery procedures with a related decline in rupture related deaths. Endo Vascular Aortic Repair [EVAR] is a well known technique of treatment of abdominal aortic aneurysms, that has changed the surgical approach to abdominal aortic aneurysms, as it is performed with low perioperative morbility and mortality rate and shorter hospital stay. However although EVAR offers immediate advantages over open surgical repair, it carries the need of close lifelong surveillance due to specific possible complications including rupture, endoleaks, graft migration and enlargement of aneurysm sac size. Contrast Enhanced Computed Tomography [CTA] is actually considered the standard reference in EVAR followup. However CTA carries high costs, radiation exposure and potential renal impairment. In the last five years several studies have been published on the role of Contrast Enhanced UltraSound [CEUS] in EVAR follow-up asserting high accuracy of this evaluation technique with absence of renal impairment, without radiation risk and at low costs. Especially since introduction of second generation Contrast Agents this evaluation technique is gaining popularity in EVAR follow-up surveillance. The diffusion of CEUS investigations by using new generation of contrast medium with appropriate software represents without any doubt an important step in the EVAR surveillance and could open up new strategies in the evaluation of endovascular aortic procedures gaining a fundamental role in EVAR follow-up.

Rapid advances in microbubble pharmacology together with novel ultrasound technologies for contrast-specific imaging of the macro- and microcirculation have led to a number of new applications for assessment of stroke patients. In particular, ultrasound perfusion imaging has added new perspectives for diagnosis and monitoring of both ischemic and hemorrhagic stroke. Recently, real-time brain perfusion imaging of middle cerebral artery infarctions has been introduced and new quantitative algorithms for evaluation of regional cerebral blood flow are being applied for the first time in humans. Microbubbles enable visualization of carotid artery plaque neovascularization to detect plaque vulnerability. There is growing interest in therapeutic applications of ultrasound, particularly in the field of sonothrombolysis. The treatment of acute ischemic stroke can be improved by ultrasound and microbubbles in combination with thrombolytic drugs. Excitingly, ultrasound and microbubbles may be effective in clot lysis of ischemic stroke even without additional thrombolytic drugs. New therapeutic avenues include opening of the blood-brain barrier (BBB) with ultrasound and microbubbles to enable novel drug delivery to the brain. Microbubbles are also assuming a central role in ultrasound molecular imaging with many targets of interest for evaluating pathophysiologic processes involved in cerebrovascular disease including angiogenesis, inflammation, and thrombus formation.

Contrast-enhanced ultrasonography (CEUS), providing relevant informations not available with non-enhanced ultrasonography, greatly impacted the practice of liver imaging. The characterization of focal liver lesions (FLLs), is obtained in a rapid, accurate and safe way and is considered the main hepatic indication; however CEUS offers other established or emergent relevant applications. Metastases detection and assessment of response to locoregional tumor treatment are accepted applications with specific indications. Needle guidance in case of poorly or non visible target lesions at conventional ultrasound is also accepted. The early assessment of response to systemic treatment, and in particular to antiangiogenic ones, by quantification software is an emergent application. The manageability of CEUS determined also its use in the operating theatre, improving the accuracy of intraoperatory US with a significant impact on final surgical strategy. In cirrhotic patients, the role of CEUS was proven highly accurate and sensitive in the characterization of portal vein thrombosis, by identification of contrast arterial enhancement inside the thrombus, that occurs only in case of neoplastic origin. In recent years microbubbles taken up by Kupffer cells, thus possessing a “postvascular” phase, were registered as ultrasound contrast agent in Japan (Sonazoid). During the post-vascular phase tumoral tissue tend to appear as a contrast defect image due to the lack of Kupffer cells, strongly contributing to tumor staging beside characterization. Newly developed techniques, such as fusion imaging or real-time three dimensional US, in addition to other applications of CEUS, in terms of post-transplantation or cholecystitis-related complications, have been recently proposed and will be discussed.

Contrast echocardiography represents a major technological advancement in the constantly evolving field of cardiovascular imaging. Over the last twenty years numerous studies have been published, illustrating the incremental value of contrast implementation into a variety of clinical scenarios. However, serious concerns have been raised about contrast safety profile, mainly based on postmarketing observational data and several animal studies. The latter have investigated contrast bio effects under experimental conditions that are not consistent with daily clinical practice. On the other hand, numerous clinical trials with large registries have proven otherwise. Not only contrast agents are efficient, but they also demonstrate a remarkable safety profile. So the question is: should we fear contrast utilization or should we consider them as an adjunctive and indispensable clinical tool for daily bedside practice?