Pancreatic cancer is characterized by extensive stromal desmoplasia, which decreases blood perfusion and impedes chemotherapy delivery. Breaking the stromal barrier could both increase perfusion and permeabilize the tumor, enhancing chemotherapy penetration. Mechanical disruption of the stroma can be achieved using ultrasound-induced bubble activity-cavitation. Cavitation is also known to result in microstreaming and could have the added benefit of actively enhancing diffusion into the tumors. Here, we report the ability to enhance chemotherapeutic drug doxorubicin penetration using ultrasound-induced cavitation in a genetically engineered mouse model (KPC mouse) of pancreatic ductal adenocarcinoma. To induce localized inertial cavitation in pancreatic tumors, pulsed high-intensity focused ultrasound (pHIFU) was used either during or before doxorubicin administration to elucidate the mechanisms of enhanced drug delivery (active vs. passive drug diffusion). For both types, the pHIFU exposures that were associated with high cavitation activity resulted in disruption of the highly fibrotic stromal matrix and enhanced the normalized doxorubicin concentration by up to 4.5-fold compared with controls. Furthermore, normalized doxorubicin concentration was associated with the cavitation metrics (P < 0.01), indicating that high and sustained cavitation results in increased chemotherapy penetration. No significant difference between the outcomes of the two types, that is, doxorubicin infusion during or after pHIFU treatment, was observed, suggesting that passive diffusion into previously permeabilized tissue is the major mechanism for the increase in drug concentration. Together, the data indicate that pHIFU treatment of pancreatic tumors when resulting in high and sustained cavitation can efficiently enhance chemotherapy delivery to pancreatic tumors.

High-intensity focused US (HIFU) is becoming more widely used for noninvasive and minimally invasive ablation of benign and malignant tumors. Recent studies suggest that HIFU can also enhance targeted drug delivery and stimulate an antitumor immune response in many tumors. However, targeting pancreatic and liver tumors by using an extracorporeal source is challenging due to the lack of an adequate acoustic window. The development of an EUS-guided HIFU transducer has many potential benefits including improved targeting, decreased energy requirements, and decreased potential for injury to intervening structures.

The transducer successfully created lesions in gel phantoms and ex vivo bovine livers. In vivo studies demonstrated that targeting and creating lesions in the porcine pancreas and liver are feasible. An EUS-guided HIFU transducer was successfully designed and developed with dimensions that are appropriate for endoscopic use. The feasibility of performing EUS-guided HIFU ablation in vivo was demonstrated in an in vivo porcine model. Further development of this technology will allow endoscopists to perform precise therapeutic ablation of periluminal lesions without breaching the wall of the gastric tract.

Pulsed high-intensity focused ultrasound (pHIFU) has been shown to enhance vascular permeability, disrupt tumor barriers and enhance drug penetration into tumor tissue through acoustic cavitation. Monitoring of cavitation activity during pHIFU treatments and knowing the ultrasound pressure levels sufficient to reliably induce cavitation in a given tissue are therefore very important. Here, three metrics of cavitation activity induced by pHIFU and evaluated by confocal passive cavitation detection were introduced: cavitation probability, cavitation persistence and the level of the broadband acoustic emissions.

These metrics were used to characterize cavitation activity in several ex vivo tissue types (bovine tongue and liver and porcine adipose tissue and kidney) and gel phantoms (polyacrylamide and agarose) at varying peak-rare factional focal pressures (1–12 MPa) during the following pHIFU protocol: frequency 1.1 MHz, pulse duration 1 ms and pulse repetition frequency 1 Hz. To evaluate the relevance of the measurements in ex vivo tissue, cavitation metrics were also investigated and compared in the ex vivo and in vivo murine pancreatic tumors that develop spontaneously in transgenic KrasLSL.G12 D/+; p53 R172 H/+ ; PdxCretg/ (KPC) mice and closely re-capitulate human disease in their morphology.

The cavitation threshold, defined at 50% cavitation probability, was found to vary broadly among the investigated tissues (within 2.5–10 MPa), depending mostly on the water-lipid ratio that characterizes the tissue composition. Cavitation persistence and the intensity of broadband emissions depended both on tissue structure and lipid concentration. Both the cavitation threshold and broadband noise emission level were similar between ex vivo and in vivo pancreatic tumor tissue. The largest difference between in vivo and ex vivo settings was found in the pattern of cavitation occurrence throughout pHIFU exposure: it was sporadic in vivo, but it decreased rapidly and stopped over the first few pulses ex vivo. Cavitation activity depended on the interplay between the destruction and circulation of cavitation nuclei, which are not only used up by HIFU treatment but also replenished or carried away by circulation in vivo. These findings are important for treatment planning and optimization in pHIFU-induced drug delivery, in particular for pancreatic tumors.

Recent studies have shown that shockwave heating and millisecond boiling in high-intensity focused ultrasound fields can result in mechanical fractionation or emulsification of tissue, termed boiling histotripsy. Visual observations of the change in color and contents indicated that the degree of thermal damage in the emulsified lesions can be controlled by varying the parameters of the exposure. The goal of this work was to examine thermal and mechanical effects in boiling histotripsy lesions using histologic and biochemical analysis. The lesions were induced in ex vivo bovine heart and liver using a 2-MHz single-element transducer operating at duty factors of 0.005–0.01, pulse durations of 5–500 ms and in situ shock amplitude of 73 MPa. Mechanical and thermal damage to tissue was evaluated histologically using conventional staining techniques (hematoxylin and eosin, and nicotinamide adenine dinucleotide-diaphorase). Thermal effects were quantified by measuring denaturation of salt soluble proteins in the treated region. According to histologic analysis, the lesions that visually appeared as a liquid contained no cellular structures larger than a cell nucleus and had a sharp border of one to two cells. Both histologic and protein analysis showed that lesions obtained with short pulses (<10 ms) did not contain any thermal damage. Increasing the pulse duration resulted in an increase in thermal damage. However, both protein analysis and nicotinamide adenine dinucleotide-diaphorase staining showed less denaturation than visually observed as whitening of tissue. The number of high-intensity focused ultrasound pulses delivered per exposure did not change the lesion shape or the degree of thermal denaturation, whereas the size of the lesion showed a saturating behavior suggesting optimal exposure duration. This study confirmed that boiling histotripsy offers an effective, predictable way to non-invasively fractionate tissue into sub-cellular fragments with or without inducing thermal damage.

Molecular imaging, the visualization of molecular and cellular markers, is a promising method for detection of dysplasia and early cancer in the esophagus and can potentially be used to identify regions of interest for biopsy or tumor margins for resection. EGFR is a previously reported cell surface receptor with stepwise increases in expression during the progression from Barrett's metaplasia to adenocarcinoma. In this work, a 200 nm fluorescent nanoparticle contrast agent was synthesized for targeted imaging of EGFR through a series of surface modifications to dye-encapsulated polystyrene particles. Amino-functionalized polystyrene particles were PEGylated using a heterobifunctional PEG linker. Subsequently, thiolated M225 antibodies were conjugated to maleimide functional groups on attached PEGs for EGFR targeting. In vitro binding studies using flow cytometry demonstrated specific binding of M225-PEG-NP to EGFR-expressing cells with minimal nonspecific binding in EGFR^– cells. Binding was shown to increase proportionally with the number of conjugated M225 antibodies. Adsorbed formulations with unmodified M225 antibodies, M225 PEG-NP, were synthesized using the same antibody feeds used in M225-PEG-NP synthesis to determine the contribution of adsorbed antibodies to EGFR targeting. Adsorbed antibodies were less efficient at mediated nanoparticle targeting to EGFR than conjugated antibodies. Finally, M225-PEG-NP demonstrated binding to EGFR-expressing regions in human esophageal tissue sections.

High intensity focused ultrasound (HIFU)-induced hyperthermia is a promising tool for cancer therapy. Three-dimensional nonlinear acoustic-bioheat transfer-blood flow-coupling model simulations and in vivo thermocouple measurements were performed to study hyperthermia effects in rabbit auricular vein exposed to pulsed HIFU (pHIFU) at varied duty cycles (DCs). pHIFU-induced temperature elevations are shown to increase with increasing DC. A critical DC of 6.9% is estimated for temperature at distal vessel wall exceeding 44°C, although different tissue depths and inclusions could affect the DC threshold. The results demonstrate clinic potentials of achieving controllable hyperthermia by adjusting pHIFU DCs, while minimizing perivascular thermal injury.

Because tumors are much larger in size compared with the beam width of high-intensity focused ultrasound (HIFU), raster scanning throughout the entire target is conventionally performed for HIFU thermal ablation. Thermal diffusion affects the temperature elevation and the consequent lesion formation. As a result, the lesion will grow continuously over the course of HIFU therapy. The purpose of this study was to investigate the influence of scanning pathways on the overall thermal lesion. Two new scanning pathways, spiral scanning from the center to the outside and spiral scanning from the outside to the center, were proposed with the same HIFU parameters (power and exposure time) for each treatment spot. The lesions produced in the gel phantom and bovine liver were compared with those using raster scanning. Although more uniform lesions can be achieved using the new scanning pathways, the produced lesion areas (27.5 plus/minus 12.3 mm^2 and 65.2 plus/minus 9.6 mm^2, respectively) in the gel phantom are significantly smaller (p < 0.05) than those using raster scanning (92.9 plus/minus 11.8 mm^2). Furthermore, the lesion patterns in the gel phantom and bovine liver were similar to the simulations using temperature and thermal dose-threshold models, respectively. Thermal diffusion, the scanning pathway and the biophysical aspects of the target all play important roles in HIFU lesion production. By selecting the appropriate scanning pathway and varying the parameters as ablation progresses, HIFU therapy can achieve uniform lesions while minimizing the total delivered energy and treatment time.

Recent studies in high intensity focused ultrasound (HIFU) have shown significant interest in generating purely mechanical damage of tissue without thermal coagulation. Here, an approach using millisecond bursts of ultrasound shock waves and repeated localized boiling is presented. In HIFU fields, nonlinear propagation effects lead to formation of shocks only in a small focal region. Significant enhancement of heating due to absorption at the shocks leads to boiling temperatures in tissue in milliseconds as calculated based on weak shock theory. The heated and potentially necrotized region of tissue is small compared to the volume occupied by the mm-sized boiling bubble it creates. If the HIFU pulse is only slightly longer than the time-to-boil, thermal injury is negligible compared to the mechanical injury caused by the exploding boiling bubble and its further interaction with shocks. Experiments performed in transparent gels and various ex vivo and in vivo tissues have confirmed the effectiveness of this emulsification method. In addition, since mm-sized boiling bubbles are highly echogenic, tissue emulsification can be easily monitored in real-time using B-mode ultrasound imaging.

Metrology of high intensity focused ultrasound (HIFU) is critical to the advancement of clinical application of HIFU for safe and effective treatments in patients. Several methods for performing metrology of HIFU systems are available in the research laboratory setting; however, translation of these methods to the clinical setting remains in evolution. From our initial experience with clinical HIFU systems we have realized the importance of accurate acoustic characterization of HIFU systems in order to determine the parameters of the treatment protocol to result in safe and effective treatments. The acoustic parameters of the system, particularly at very high intensities, are very important to understand prior to delivering HIFU therapy to patients. Improved methods of HIFU metrology, especially to determine in situ exposure and dose, will result in a more rational approach to clinical HIFU therapy. Further advances in clinical HIFU therapy will require close cooperation between clinicians and scientists in order to make HIFU therapy safe and effective. Educating clinicians on the importance of metrology will also be important.

Shock-wave heating and millisecond boiling in high intensity focused ultrasound fields have been shown to result in mechanical emulsification of ex-vivo tissue. In this work, the same in situ exposures were applied in vivo in pig liver and in mice bearing 5-7 mm subcutaneous tumors (B16 melanoma) on the hind limb. Lesions were produced using a 2-MHz annular array in the case of pig liver (shock amplitudes up to 98 MPa) and a 3.4-MHz single-element transducer in the case of mouse tumors (shock amplitude of 67 MPa). The parameters of the pulsing protocol (1-500 ms pulse durations and 0.01-0.1 duty factor) were varied depending on the extent of desired thermal effect. All exposures were monitored using B-mode ultrasound. Mechanical and thermal tissue damage in the lesions was evaluated histologically using H&E and NADH-diphorase staining. The size and shape of emulsified lesions obtained in-vivo agreed well with those obtained in ex-vivo tissue samples using the same exposure parameters. The lesions were successfully produced both in bulk liver tissue at depths of 1-2 cm and in superficial tumors at depths less than 1 mm without damaging the skin.

A wide variety of treatment protocols have been employed in high intensity focused ultrasound (HIFU) treatments, and the resulting bioeffects observed include both mechanical as well as thermal effects. In recent studies, there has been significant interest in generating purely mechanical damage using protocols with short, microsecond pulses. Tissue erosion effects have been attained by operating HIFU sources using short pulses of 10–20 cycles, low duty cycles (<1%), and pulse average intensities of greater than 20 kW/cm².

The goal of this work was to use a modified pulsing protocol, consisting of longer, millisecond-long pulses of ultrasound and to demonstrate that heating and rapid millisecond boiling from shock wave formation can be harnessed to induce controlled mechanical destruction of soft tissue. Experiments were performed in excised bovine liver and heart tissue using a 2-MHz transducer. Boiling activity was monitored during exposures using a high voltage probe in parallel with the HIFU source. In situ acoustic fields and heating rates were determined for exposures using a novel derating approach for nonlinear HIFU fields. Several different exposure protocols were used and included varying the duty cycle, pulse length, and power to the source. After exposures, the tissue was sectioned, and the gross lesion morphology was observed. Different types of lesions were induced in experiments that ranged from purely thermal to purely mechanical depending on the pulsing protocol used. Therefore, shock wave heating and millisecond boiling may be an effective method for reliably generating significant tissue erosion effects.

Targeted vascular occlusion is desirable for clinical therapies such as in the treatment of esophageal and gastric varices and varicose veins. The feasibility of ultrasound-mediated endothelial damage for vascular occlusion was studied. A segment of a rabbit auricular vein was treated in vivo with low duty cycle, high peak rarefaction pressure (9 MPa) high-intensity focused ultrasound pulses in the presence of intravenously administered circulating microbubbles, followed by fibrinogen injection, which resulted in the formation of an acute occlusive intravascular thrombus. Further investigation and refinements of treatment protocols are necessary for producing durable vascular occlusion.

High-intensity focused ultrasound (HIFU) is being promoted as a noninvasive method to treat certain primary solid tumors, metastatic disease, and enhance drug delivery. The field of medicine is evolving towards increasing use of noninvasive and minimally invasive therapies such as HIFU. This article provides an overview of current clinical applications of HIFU and future requirements to expand the clinical applications of this technique.

Extracorporeal high-intensity focused ultrasound (HIFU) can be used to ablate tissue noninvasively by delivering focused ultrasound energy from an external source. HIFU for clinical treatment of pancreatic cancer has been reported; however, systematic evaluation of the safety and efficacy of pancreatic ablation with HIFU has not been performed. The objectives of this in vivo study are as follows: (1) assess the safety and feasibility of targeting and ablating pancreatic tissue using the FEP-BY02 HIFU system (Yuande Bio-Medical Engineering, Beijing, China); (2) evaluate a method for estimating in situ acoustic treatment energy in an in vivo setting; and (3) identify the optimal treatment parameters that result in safe and effective ablation of the pancreas.

The pancreata of 12 common swine were treated in vivo. Prior to therapy, blood was drawn for laboratory analysis. Animals were then treated with extracorporeal HIFU at three different acoustic treatment energies (750, 1000 and 1250 J). Endoscopy was performed prior to and immediately following HIFU therapy to assess for gastric injury. Blood was drawn after completion of the treatment and on days 2 and 7 following treatment to assess for biochemical evidence of pancreatitis. Animals were then euthanized 7 d following treatment and a necropsy was performed to assess for unintended injury and to obtain pancreatic tissue for histology to assess efficacy of HIFU ablation. Histologic scoring of pancreatic tissue changes was performed by a pathologist blinded to the treatment energy delivered. The degree of ablation identified on histology correlated with the treatment energy. No collateral tissue damage was seen at treatment energies of 750 and 1000 J. At 1250 J, thermal injury to the abdominal muscles and gastric ulcers were observed. There were no premature deaths, serious illnesses, skin burns or evidence of pancreatitis on biochemical analysis. HIFU treatment of the pancreas is feasible, safe and can be used to ablate tissue noninvasively. A clinical trial in humans examining the use of extracorporeal HIFU for palliation of pain related to pancreatic cancer is planned.

High intensity focused ultrasound (HIFU) is emerging as a modality for treatment of solid tumors. The temperature at the focus can reach over 65°C denaturing cellular proteins resulting in coagulative necrosis. Typically, HIFU parameters are the same for each treated spot in most HIFU control systems. Because of thermal diffusion from nearby spots, the size of lesions will gradually become larger as the HIFU therapy progresses, which may cause insufficient treatment of initial spots, and over-treatment of later ones. It is found that the produced lesion pattern also depends on the scanning pathway. From the viewpoint of the physician creating uniform lesions and minimizing energy exposure are preferred in tumor ablation. An algorithm has been developed to adaptively determine the treatment parameters for every spot in a theoretical model in order to maintain similar lesion size throughout the HIFU therapy. In addition, the exposure energy needed using the traditional raster scanning is compared with those of two other scanning pathways, spiral scanning from the center to the outside and from the outside to the center. The theoretical prediction and proposed algorithm were further evaluated using transparent gel phantoms as a target. Digital images of the lesions were obtained, quantified, and then compared with each other. Altogether, dynamically changing treatment parameters can improve the efficacy and safety of HIFU ablation.

Previous in vivo studies have demonstrated that microvessel hemorrhages and alterations of endothelial permeability can be produced in tissues containing microbubble-based ultrasound contrast agents when those tissues are exposed to MHz-frequency pulsed ultrasound of sufficient pressure amplitudes. The general hypothesis guiding this research was that acoustic (viz., inertial) cavitation, rather than thermal insult, is the dominant mechanism by which such effects arise. We report the results of testing five specific hypotheses in an in vivo rabbit auricular blood vessel model: (1) acoustic cavitation nucleated by microbubble contrast agent can damage the endothelia of veins at relatively low spatial-peak temporal-average intensities, (2) such damage will be proportional to the peak negative pressure amplitude of the insonifying pulses, (3) damage will be confined largely to the intimal surface, with sparing of perivascular tissues, (4) greater damage will occur to the endothelial cells on the side of the vessel distal to the source transducer than on the proximal side and (5) ultrasound/contrast agent-induced endothelial damage can be inherently thrombogenic, or can aid sclerotherapeutic thrombogenesis through the application of otherwise subtherapeutic doses of thrombogenic drugs. Auricular vessels were exposed to 1-MHz focused ultrasound of variable peak pressure amplitude using low duty factor, fixed pulse parameters, with or without infusion of a shelled microbubble contrast agent. Extravasation of Evans blue dye and erythrocytes was assessed at the macroscopic level. Endothelial damage was assessed via scanning electron microscopy (SEM) image analysis. The hypotheses were supported by the data. We discuss potential therapeutic applications of vessel occlusion, e.g., occlusion of at-risk gastric varices.