Chien Ting Chin

Focused ultrasound and microbubbles for enhanced extravasation

The permeability of blood vessels for albumin can be altered by using ultrasound and polymer or lipid-shelled microbubbles. The region in which the microbubbles were destroyed with focused ultrasound was quantified in gel phantoms as a function of pressure, number of cycles and type of microbubble. At 2MPa the destruction took place in a fairly wide area for a lipid-shelled agent, while for polymer-shelled agents at this setting, distinct destruction spots with a radius of only 1mm were obtained. When microbubbles with a thicker shell were used, the pressure above which the bubbles were destroyed shifts to higher values. In vivo both lipid and polymer microbubbles increased the extravasation of the albumin binding dye Evans Blue, especially in muscle leading to about 6-8% of the injected dose to extravasate per gram muscle tissue 30 min after start of the treatment, while no Evans Blue could be detected in muscle in the absence of microbubbles. Variation in the time between ultrasound treatment and Evans Blue injection, demonstrated that the time window for promoting extravasation is at least an hour at the settings used. In MC38 tumors, extravasation already occurred without ultrasound and only a trend towards enhancement with about a factor of 2 could be established with a maximum percentage injected dose per gram of 3%. Ultrasound mediated microbubble destruction especially enhances the extravasation in the highly vascularized outer part of the MC38 tumor and adjacent muscle and would, therefore, be most useful for release of, for instance, anti-angiogenic drugs.

Targeted Ultrasound-Mediated Delivery of Nanoparticles: On the Development of a New HIFU-Based Therapy and Imaging Device

Ultrasound-mediated delivery (USMD) is an active research topic, as researchers develop applications for therapeutic ultrasound in addition to thermal ablation. In USMD, ultrasound is used in conjunction with microbubbles and drugs, nanoparticles, siRNA, pDNA, stem cells, etc., to facilitate their cellular delivery and uptake using pressure and temperature-mediated mechanisms to bring about a desired therapeutic effect. To investigate the potential of targeted USMD of nanoparticles, pDNA, and stem cells for cardiovascular and other applications, a general-purpose preclinical research tool, therapy imaging probe system (TIPS) was designed. It consists of a wideband annular array, a small-animal acoustic coupler, a motorized positioning system, integrated control software for ultrasound image-guided treatment planning and execution, and triggering electronics that allow ECG and respiration-gated ultrasound exposures. TIPS was then used to enhance delivery of nanoparticles into the murine myocardium and heart vessel walls to demonstrate the feasibility of the technology, pave the way for additional basic research in cardiovascular USMD, and begin to explore the requirements that USMD devices will have to meet to be useful in a clinical setting.

Ultrasound mediated drug delivery

For patients with ulcerative colitis, the most common form of inflammatory bowel disease (IBD), aminosalicylates such as mesalamine provide first line anti-inflammatory therapy. Since efficacy is related to tissue concentration, when administered as a rectal enema lack of retention challenges topical formulation effectiveness. To address this problem, Schoellhammer and his colleagues (Schoellhammer et al 2015) recently described the development and pre-clinical testing of a novel delivery system in which they incorporated low frequency ultrasound (LFU) as a means to enhance transmucosal drug delivery through the gastrointestinal mucosa.

Control and reversal of tumor growth by ultrasound activated microbubbles

It has been known that tumor insonation after intravenous microbubble injection may lead to transient reduction of tumor blood flow. Subsequent experiments showed that repeated induction of tumor blood flow disruption may retard tumor growth. This paper reports an initial investigation of the underlying blood flow disruption phenomena and the first systematic investigation of the control of tumor growth by the combination of moderately intense ultrasound and microbubbles.

Enhanced gene transfection of plasmid DNA in the liver with ultrasound and microbubbles

Ultrasound mediated delivery (USMD) in the presence of microbubbles is a potentially safe and effective method for gene therapy, offering many desired characteristics such as low toxicity, potential for repeated treatment, as well as organ specificity. In this study we tested the capability of USMD to improve gene transfection to mice livers for treating a rare genetic disorder, Glycogen Storage Disease Type 1a. An image guided therapeutic ultrasound system (TIPS, Philips) was used to provide therapeutic ultrasound to mice liver under IACUC approved protocols. Anesthetized healthy mice were placed supine on a heated pad and coupled to the TIPS transducer. Therapeutic plasmid DNA (FLAG-pG6Pase) was injected and microbubbles infused via the tail vein. Treated animals (n=4) received therapeutic ultrasound pulses (1 MHz, 2.5 MPa) over the entire liver. Control animals (n=5) received pDNA but not ultrasound treatment. Post treatment the animals were left to recover and subsequently sacrificed after 5 days. Tissues from the left, middle, caudal, right anterior, and right posterior liver lobes were harvested and stored. Quantitative PCR assays were then performed on the samples to quantify gene transfection. Ultrasound treated animals showed significantly higher levels of G6Pase transfection compared to control animals (p<0.05) in all five lobes of the liver. On average, the treated animals showed 5.4 times more pDNA accumulation in the liver compared to controls. Immunohistochemistry staining for FLAG tag showed increased transgene expression especially around the blood vessels in treated animals. No evidence of toxicity was found up to 5 days post treatment.

Ultrasound activated paclitaxel delivery in mice using a combined therapy and imaging probe system

Localized therapeutic effects of an experimental therapeutic agent on a mouse MC38 tumor model were evaluated using a combined ultrasound therapy and imaging probe system (TIPS). With ultrasound activated paclitaxel delivery, the tumor growth is substantially retarded for at least 3 days. The therapeutic effect of ultrasound is statistically significant (p=0.03) within one day after treatment but gradually decreases with time. Therefore, it is suggested that multiple treatments over a long period are required for cancer treatment. It is also demonstrated in this preclinical study that TIPS is an effective tool for imaging-guided, ultrasound-activated drug delivery.

Triggered drug release from liposome-microbubble pendant complexes: From model systems to tumor therapy in a murine adenocarcinoma model

Liposome-microbubble complexes allow delivery of drugs that cannot be otherwise associated with the bubble shell: water-soluble drugs and proteins. We prepared liposome-microbubble pendants by decorating biotinylated microbubbles with biotinylated liposomes via streptavidin. A model dye calcein was used as a model release marker. Using focused ultrasound (Philips TIPS, 1 MHz, 7 MPa) we were able to release ~30% of the entrapped dye. For an enzyme thrombin, ~11% of the entrapped material was released following pendant insonation. In vivo tumor therapy was performed with doxorubicin-liposome-microbubble pendants in a subcutaneous MC38 murine adenocarcinoma model. Doxorubicin was loaded in liposomes via an ammonium citrate gradient procedure. By using larger liposomes, >>80nm (as in Doxil/Lipodox), we prepared pendants carrying ~1 pg doxorubicin per particle. To avoid tumor blood flow stoppage caused by high-power insonation of microbubbles in tumor vasculature, we applied continuous sine wave ultrasound (Bi...

Ultrasound mediated gene silencing with short-hairpin RNA

Ultrasound-mediated delivery (USMD) of drugs and genes, especially with the enhancement of microbubble or nanoparticle agents, has been demonstrated in various settings for some years. Translation to pre-clinical disease models faces many challenges. This paper reports on the development and characteristics of a device for preclinical translational research in USMD. Feasibility of short-haripin RNA (shRNA) delivery in vivo using this device was demonstrated.