Intravenous Injection Of Microbubbles Research Articles

Effects of low-intensity ultrasound opening the blood-brain barrier on Alzheimer's disease-a mini review.

Due to the complex pathological mechanisms of Alzheimer's disease (AD), its treatment remains a challenge. One of the major difficulties in treating AD is the difficulty for drugs to cross the blood-brain barrier (BBB). Low-intensity ultrasound (LIUS) is a novel type of ultrasound with neuromodulation function. It has been widely reported that LIUS combined with intravenous injection of microbubbles (MB) can effectively, safely, and reversibly open the BBB to achieve non-invasive targeted drug delivery. However, many studies have reported that LIUS combined with MB-mediated BBB opening (LIUS + MB-BBBO) can improve pathological deposition and cognitive impairment in AD patients and mice without delivering additional drugs. This article reviews the relevant research studies on LIUS + MB-BBBO in the treatment of AD, analyzes its potential mechanisms, and summarizes relevant ultrasound parameters.

Focused Ultrasound-Mediated Delivery of Anti-Programmed Cell Death-Ligand 1 Antibody to the Brain of a Porcine Model.

Immune checkpoint inhibitor (ICI) therapy has revolutionized cancer treatment by leveraging the body's immune system to combat cancer cells. However, its effectiveness in brain cancer is hindered by the blood-brain barrier (BBB), impeding the delivery of ICIs to brain tumor cells. This study aimed to assess the safety and feasibility of using focused ultrasound combined with microbubble-mediated BBB opening (FUS-BBBO) to facilitate trans-BBB delivery of an ICI, anti-programmed cell death-ligand 1 antibody (aPD-L1) to the brain of a large animal model. In a porcine model, FUS sonication of targeted brain regions was performed after intravenous microbubble injection, which was followed by intravenous administration of aPD-L1 labeled with a near-infrared fluorescent dye. The permeability of the BBB was evaluated using contrast-enhanced MRI in vivo, while fluorescence imaging and histological analysis were conducted on ex vivo pig brains. Results showed a significant 4.8-fold increase in MRI contrast-enhancement volume in FUS-targeted regions compared to nontargeted regions. FUS sonication enhanced aPD-L1 delivery by an average of 2.1-fold, according to fluorescence imaging. In vivo MRI and ex vivo staining revealed that the procedure did not cause significant acute tissue damage. These findings demonstrate that FUS-BBBO offers a noninvasive, localized, and safe delivery approach for ICI delivery in a large animal model, showcasing its potential for clinical translation.

Focused ultrasound-mediated delivery of anti-programmed cell death-ligand 1 antibody to the brain of a porcine model

Immune checkpoint inhibitor (ICI) therapy has revolutionized cancer treatment by leveraging the body's immune system to combat cancer cells. However, its effectiveness in brain cancer is hindered by the blood-brain barrier (BBB), impeding the delivery of ICIs to brain tumor cells. This study aimed to assess the safety and feasibility of using focused ultrasound combined with microbubbles-mediated BBB opening (FUS-BBBO) to facilitate trans-BBB delivery of an ICI, anti-programmed cell death-ligand 1 antibody (aPD-L1), to the brain of a large animal model. In a porcine model, FUS sonication of targeted brain regions was performed after intravenous microbubble injection, which was followed by intravenous administration of aPD-L1 labeled with a near-infrared fluorescent dye. The permeability of the BBB was evaluated using contrast-enhanced MRI, while fluorescence imaging and histological analysis were conducted on ex vivo pig brains. Results showed a significant 4.8-fold increase in MR contrast enhancement volume in FUS-targeted regions compared to non-targeted regions. FUS sonication enhanced aPD-L1 delivery by an average of 2.1-fold, according to fluorescence imaging. In vivo MRI and ex vivo staining found the procedure did not cause significant acute tissue damage. These findings demonstrate that FUS-BBBO offers a noninvasive, localized, and safe delivery approach for ICIs in a large animal model, showcasing its potential for clinical translation.

Assessment of Takayasu's arteritis activity by ultrasound localization microscopy.

Ultrasound localization microscopy (ULM) based on ultrafast ultrasound imaging of circulating microbubbles (MB) can image microvascular blood flows invivo up to the micron scale. Takayasu arteritis (TA) has an increased vascularisation of the thickened arterial wall when active. We aimed to perform vasa vasorum ULM of the carotid wall and demonstrate that ULM can provide imaging markers to assess the TA activity. Patients with TA were consecutively included with assessment of activity by the National Institute of Health criteria: 5 had active TA (median age 35.8 [24.5-46.0] years) and 11 had quiescent TA (37.2 [31.7-47.3] years). ULM was performed using a 6.4MHz probe and a dedicated imaging sequence (plane waves with 8 angles, frame rate 500Hz), coupled with the intravenous injection of MB. Individual MB were localised at a subwavelength scale then tracked, allowing the reconstruction of the vasa vasorum flow anatomy and velocity. ULM allowed to show microvessels and to measure their flow velocity within the arterial wall. The number of MB detected per second in the wall was 121 [80-146] in active cases vs. 10 [6-15] in quiescent cases (p=0.0005), with a mean velocity of 40.5 [39.0-42.9] mm.s-1 in active cases. ULM allows visualisation of microvessels within the thickened carotid wall in TA, with significantly greater MB density in active cases. ULM provides a precise visualisation invivo of the vasa vasorum and gives access to the arterial wall vascularisation quantification. French Society of Cardiology. ART (Technological Research Accelerator) biomedical ultrasound program of INSERM, France.

Feasibility of MRI-guided focused ultrasound-mediated intranasal delivery in a large animal model

Intranasal delivery provides non-invasive delivery of drugs from the nose to the brain bypassing the blood-brain barrier (BBB). We previously reported the focused ultrasound (FUS)-mediated intranasal delivery (FUSIN) of various agents in mice using FUS. However, no study has investigated the feasibility of FUSIN in large animals. The objective of this study was to investigate the feasibility of MR-guided FUS-mediated intranasal delivery in a large animal model. Pigs were used as the large animal model due to their clinical relevance to humans. Fluorescence-labeled albumin was mixed with an MRI contrast agent and administrated intranasally to the pig using a catheter. Followed by intravenous injection of microbubbles, FUS sonication was performed at two brain regions, the cortex and brainstem. T1-weighted MR images showed enhancement of MRI contrast agent at the olfactory region of the pig nose. Fluorescence imaging displayed enhanced fluorescence intensity at the FUS-targeted brain regions. These findings suggest that MR-guided FUS can achieve noninvasive and localized delivery of intranasally administered agents to the brain in large animals.

Dynamics of focused ultrasound-enhanced glymphatic transportation in the mouse brain

The glymphatic system, a perivascular network in the brain, regulates the exchange of cerebrospinal fluid and interstitial fluid in the perivascular space. Focused ultrasound combined with microbubbles (FUSMB) has been recently shown feasible to manipulate the glymphatic transportation by enhancing the intranasal delivery of agents in the perivascular space. The objective of this study was to reveal the dynamics of FUSMB-enhanced agent transport in the glymphatic system. A ring-shaped FUS transducer was confocally aligned with an objective of a two-photon microscope (2PM). Fluorescently labeled albumin was administered to the mouse nose, where it transported along olfactory nerve and trigeminal nerve from nose to brain and then spread in perivascular space in the brain. FUS sonication was performed after intravenous injection of microbubbles. In vivo 2PM recorded the dynamics of agent transportation before, during, and after FUS sonication. Time-lapse recording showed that FUSMB enhanced the agent accumulation at the perivascular space. FUS significantly increased the accumulation of the agent in the perivascular space by 1.2-folds immediately after the sonication (p = 0.0325), and 1.6-folds at 5 min after the sonication (p < 0.0001). In conclusion, this study showed, for the first time, direct evidence of FUSMB-enhanced glymphatic transportation in the mouse brain.

Detection and Characterization of Sentinel Lymph Node by Ultrasound Molecular Imaging with B7-H3-Targeted Microbubbles in Orthotopic Breast Cancer Model in Mice.

Accurate identification and assessment of sentinel lymph node (SLN) using noninvasive imaging methods can play a vital role in tumor staging, surgical planning, and prognostic evaluation. In this study, we assessed the efficacy of B7-H3-targeted molecular-ultrasound imaging for the early SLN detection, and characterization in a mouse model of orthotopic breast cancer. We established a mouse breast cancer model with lymph node metastasis by injecting MAD-MB 231 cells which were engineered to express firefly luciferase reporter gene into the fat pad of the right 4th mammary gland in female BALB/c nude mice. The sole lymph node (LN) close to the tumor was regarded as the SLN for imaging investigation, which included metastatic and non-metastatic SLNs. The LN in the right 4th mammary gland from normal mice was used as normal control (normal mice LN). The commercially available preclinical streptavidin-coated, perfluorocarbon-containing lipid-shelled microbubbles (VisualSonics, Toronto, Canada) were used to generate B7-H3-targeted microbubbles (MBB7-H3) and control microbubbles (MBControl). Then, ultrasound molecular imaging (USMI) was performed using a high-resolution transducer (MS250; center frequency, 21MHz; Vevo 2100; VisualSonics, Toronto, Canada) after intravenous injection of microbubbles. The SLN was clearly detected and located under conventional (B-mode) and contrast-enhanced ultrasonography with microbubble injection. The metastatic SLNs showed a markedly higher signal from B7-H3-targeted microbubbles (MBB7-H3) compared to the non-metastatic SLNs and normal LNs. The metastatic SLN was further confirmed by ex vivo bioluminescence imaging and eventually verified by histological analysis. Our findings suggest the potential value of USMI using B7-H3 targeted microbubbles in breast cancer and establish an effective imaging method for the non-invasive detection and characterization of SLN.

Static Magnetic Fields Dampen Focused Ultrasound-mediated Blood-Brain Barrier Opening.

Background Focused ultrasound combined with microbubbles has been used in clinical studies for blood-brain barrier (BBB) opening in conjunction with MRI. However, the impact of the static magnetic field generated by an MRI scanner on the BBB opening outcome has not been evaluated. Purpose To determine the relationship of the static magnetic field of an MRI scanner on focused ultrasound combined with microbubble-induced BBB opening. Materials and Methods Thirty wild-type mice were divided into four groups. Mice from different groups were sonicated with focused ultrasound in different static magnetic fields (approximately 0, 1.5, 3.0, and 4.7 T), with all other experimental parameters kept the same. Focused ultrasound sonication was performed after intravenous injection of microbubbles. Microbubble cavitation activity, the fundamental -physical mechanism underlying focused ultrasound BBB opening, was monitored with passive cavitation detection. After sonication, contrast-enhanced T1-weighted MRI was performed to assess BBB opening outcome. Intravenously injected Evans blue was used as a model agent to evaluate trans-BBB delivery efficiency. Results The microbubble cavitation dose decreased by an average of 2.1 dB at 1.5 T (P = .05), 2.9 dB at 3.0 T (P = .01), and 3.0 dB at 4.7 T (P = .01) compared with that outside the magnetic field (approximately 0 T). The static magnetic field of an MRI scanner decreased BBB opening volume in mice by 3.2-fold at 1.5 T (P < .001), 4.5-fold at 3.0 T (P < .001), and 11.6-fold at 4.7 T (P <.001) compared with mice treated outside the magnetic field. It also decreased Evans blue trans-BBB delivery 1.4-fold at 1.5 T (P = .009), 1.6-fold at 3.0 T (P < .001), and 1.9-fold at 4.7 T (P < .001). Conclusion Static magnetic fields dampened microbubble cavitation activity and decreased trans-blood-brain barrier (BBB) delivery by focused ultrasound combined with microbubble-induced BBB opening. © RSNA, 2021 An earlier incorrect version of this article appeared online. This article was corrected on July 8, 2021.

Focused ultrasound mediated blood\u2013brain barrier opening is safe and feasible in a murine pontine glioma model

Drug delivery in diffuse intrinsic pontine glioma is significantly limited by the blood-brain barrier (BBB). Focused ultrasound (FUS), when combined with the administration of microbubbles can effectively open the BBB permitting the entry of drugs across the cerebrovasculature into the brainstem. Given that the utility of FUS in brainstem malignancies remains unknown, the purpose of our study was to determine the safety and feasibility of this technique in a murine pontine glioma model. A syngeneic orthotopic model was developed by stereotactic injection of PDGF-B+PTEN−/−p53−/− murine glioma cells into the pons of B6 mice. A single-element, spherical-segment 1.5 MHz ultrasound transducer driven by a function generator through a power amplifier was used with concurrent intravenous microbubble injection for tumor sonication. Mice were randomly assigned to control, FUS and double-FUS groups. Pulse and respiratory rates were continuously monitored during treatment. BBB opening was confirmed with gadolinium-enhanced MRI and Evans blue. Kondziela inverted screen testing and sequential weight lifting measured motor function before and after sonication. A subset of animals were treated with etoposide following ultrasound. Mice were either sacrificed for tissue analysis or serially monitored for survival with daily weights. FUS successfully caused BBB opening while preserving normal cardiorespiratory and motor function. Furthermore, the degree of intra-tumoral hemorrhage and inflammation on H&E in control and treated mice was similar. There was also no difference in weight loss and survival between the groups (p > 0.05). Lastly, FUS increased intra-tumoral etoposide concentration by more than fivefold. FUS is a safe and feasible technique for repeated BBB opening and etoposide delivery in a preclinical pontine glioma model.

Subsequent Ultrasound Vascular Targeting Therapy of Hepatocellular Carcinoma Improves the Treatment Efficacy.

Simple SummaryThe incidence of hepatocellular carcinoma (HCC) has consistently increased over the last two decades. Although surgical resection is the preferred treatment for HCC, approximately 80% of the patients are not suited for surgery because the tumors are in advanced stage of the disease and unresectable. Locoregional therapies like anti-vascular ultrasound (AVUS) provide a unique alternative for treatment. In this study we evaluate the use of repeated subsequent AVUS therapy as an approach to optimize the therapeutic efficacy by targeting tumor vasculature at different times. This approach can overcome the heterogeneity in tumor vasculature structure and functionality often observed at time of therapy. For this purpose, we evaluated two regiments of subsequent AVUS treatments with a predetermined time interval between the treatments. The results showed that AVUS when applied subsequently in a shorter duration, boosts the therapy effects showing more than 75% reduction in tumor perfusion post therapy. However, this improvement was lost for a longer duration between subsequent therapies. These findings demonstrate the significance of the viability of tumors for maximizing the therapeutic effects of tumor vasculature targeting therapies.The response of hepatocellular carcinoma (HCC) to anti-vascular ultrasound therapy (AVUS) can be affected by the inherent differences in tumor vascular structure, and the functionality of tumor vessels at the time of treatment. In this study, we evaluate the hypothesis that repeated subsequent AVUS therapies are a possible approach to overcome these factors and improve the therapeutic efficacy of AVUS. HCC was induced in 30 Wistar rats by oral ingestion of diethylnitrosamine (DEN) for 12 weeks. A total of 24 rats received treatment with low intensity, 2.8 MHz ultrasound with an intravenous injection of microbubbles. Treated rats were divided into three groups: single therapy group (ST), 2-days subsequent therapy group (2DST), and 7-days subsequent therapy group (7DST). A sham control group did not receive ultrasound therapy. Tumor perfusion was measured by quantitative contrast-enhanced ultrasound (CEUS) nonlinear and power-Doppler imaging. Tumors were harvested for histologic evaluation of ultrasound-induced vascular changes. ANOVA was used to compare the percent change of perfusion parameters between the four treatment arms. HCC tumors treated with 2DST showed the largest reduction in tumor perfusion, with 75.3% reduction on average for all perfusion parameters. The ST group showed an average decrease in perfusion of 54.3%. The difference between the two groups was significant p < 0.001. The 7DST group showed a reduction in tumor perfusion of 45.3%, which was significant compared to the 2DST group (p < 0.001) but not different from the ST group (p = 0.2). The use of subsequent targeted AVUS therapies applied shortly (two days) after the first treatment enhanced the anti-vascular effect of ultrasound. This gain, however, was lost for a longer interval (1 week) between the therapies, possibly due to tumor necrosis and loss of tumor viability. These findings suggest that complex interplay between neovascularization and tumor viability plays a critical role in treatment and, therefore, must be actively monitored following treatment by CEUS for optimizing sequential treatment.

DDEL-13. FOCUSED ULTRASOUND MEDIATED BLOOD BRAIN BARRIER DISRUPTION IN A MURINE MODEL OF PONTINE GLIOMA: A SAFETY AND FEASIBILITY STUDY

BACKGROUNDDrug delivery remains a major obstacle in DIPG, as the blood brain barrier (BBB) limits the penetration of systemic therapies to the brainstem. Focused ultrasound (FUS) is an exciting new technology that, when combined with microbubbles, can open the BBB permitting the entry of drugs across the cerebrovasculature. Given that the utility of FUS in brainstem tumors remains unknown, the purpose of our study was to determine the safety and feasibility of this technique in a murine pontine glioma model.METHODSA syngeneic orthotopic model was established by stereotactic injection of PDGF-B+PTEN-/-p53-/- murine glioma cells (10,000/1ul) into the pons of B6 albino mice. A single-element, spherical-segment FUS transducer (center frequency=1.5MHz) driven by a function generator through a power amplifier (acoustic pressure=0.7MPa) was used with concurrent intravenous microbubble injection (FUS+MB) to sonicate the tumor on post-injection day 14. BBB opening was confirmed with gadolinium-enhanced MRI and Evans blue. Kondziela inverted screen (KIS) testing was completed to measure motor function. Mice were either immediately sacrificed for histopathological assessment or serially monitored for survival.RESULTSIn mice treated with FUS (n=11), there was no measured deficit in KIS testing. Additionally, the degree of intra-tumoral hemorrhage and inflammation on H&E in control (n=5) and treated mice (n=5) was similar. Lastly, there was no difference in survival between the groups (control, n=6, median=26 days; FUS, n=6, median=25 days, p>0.05).CONCLUSIONFUS+MB is a safe and feasible technique to open the BBB in a preclinical pontine glioma model.

Characterization of focused ultrasound-mediated brainstem delivery of intranasally administered agents

Focused ultrasound-mediated intranasal (FUSIN) delivery is a recently proposed technique that bypasses the blood-brain barrier to achieve noninvasive and localized brain drug delivery. The goal of this study was to characterize FUSIN drug delivery outcome in mice with regard to its dependency on several critical experimental factors, including the time interval between IN administration and FUS sonication (Tlag1), the FUS pressure, and the time for sacrificing the mice post-FUS (Tlag2). Wild-type mice were treated by FUSIN delivery of near-infrared fluorescent dye-labeled bovine serum albumin (800CW-BSA, used as a model agent). 800CW-BSA was intranasally administered to the mice in vivo, followed by intravenous injection of microbubbles and FUS sonication at the brainstem. Fluorescence imaging of ex vivo mouse brain slices was used to quantify the delivery outcomes of 800CW-BSA. Major organs, along with the nasal tissue and trigeminal nerve, were harvested to assess the biodistribution of 800CW-BSA. The delivery outcome of 800CW-BSA was the highest at the brainstem when Tlag1 was 0.5 h, which was on average 24.5-fold, 5.4-fold, and 21.6-fold higher than those of the IN only, Tlag1 = 1.5 h, and Tlag1 = 4.0 h, respectively. The FUSIN delivery outcome at the lowest pressure level, 0.43 MPa, was on average 1.8-fold and 3.7-fold higher than those at 0.56 MPa and 0.70 MPa, respectively. The mean concentration of 800CW-BSA in the brainstem after FUSIN delivery decreased from 0.5 h to 4.0 h post-FUS. The accumulation of 800CW-BSA was low in the heart, lung, spleen, kidneys, and liver, but high in the stomach and intestines. This study revealed the unique characteristics of FUSIN as a noninvasive, efficient, and localized brain drug delivery technique.

Evaluation of the Uncertainty of Passive Cavitation Measurements for Blood–Brain Barrier Disruption Monitoring

Exposure to ultrasound combined with intravenous injection of microbubbles is a technique that can be used to temporarily disrupt the blood–brain barrier. Transcranial monitoring of cavitation can be done with one or more passive cavitation detectors (PCDs). However, the positioning of the PCDs relative to the cavitation site and the attenuation of these signals by the skull are two sources of error in the quantification of cavitation activity. The aim of this study was to evaluate in vitro the amplitude variation of cavitation signals that can be expected for an excised porcine skull model. The variation caused by the relative positioning of the PCD with respect to the cavitation site was quantified. A position-based correction of the signal amplitude was evaluated. Pig skull samples were used to assess variation in signal amplitude caused by bone. The overall coefficient of variation of the signals owing to these measurement biases was estimated at 30.8%.

Enhancement Effect of Microbubble-Enhanced Ultrasound in Microwave Ablation in Rabbit VX2 Liver Tumors.

Objectives One reason for the high recurrence and metastatic rates of tumors such as hepatocellular carcinoma (HCC) treated by microwave ablation (MWA) is the presence of residual foci in the tumor due to heat sink effect. Microbubble-enhanced ultrasound (MEUS) can noninvasively disrupt and block the tumor blood perfusion and has the potential to overcome the heat sink effect and enhance the therapeutic effect of MWA. The study aimed at evaluating the potential additional benefit of microbubble-enhanced ultrasound (MEUS) in hepatocellular carcinoma (HCC) treated by microwave ablation (MWA). Methods In this study, a new strategy of combining MWA with MEUS for treating HCC was proposed. Twenty-four rabbits with VX2 tumors in livers were randomly divided into MEUS + MWA, MEUS alone, MWA alone, and blank control groups, respectively (n = 6). In the MEUS group, the tumors were directly exposed to therapeutic ultrasound for 5 min with a concurrent intravenous injection of microbubbles (0.1 ml/kg diluted into 5 ml saline). In the MWA group, the tumors were treated by MWA for 1 min. In the MEUS + MWA group, tumors were ablated by MWA for 1 min after ultrasound cavitation enhanced by microbubbles as in the MEUS group. In the blank control group, the tumors received probe sham and intravenous saline. Contrast-enhanced ultrasound (CEUS) was performed before treatment and immediately after treatment to display the size, shape, and contour of the tumors. Throughout the treatment process, the local temperature of the treatment area was detected by a temperature needle punctured into the tumor. The blood samples of animals were obtained after treatment for evaluating the liver function. Tumor cell necrosis and apoptotic rates were observed after treatment by histological examination. Results CEUS showed that although perfusion defects appeared in all the treatment groups, especially in the MEUS + MWA group, there was no significant difference between the two groups on the volumes of perfusion defects, which were 1.78 ± 0.31 (cm3) in the MWA group and 1.84 ± 0.20 (cm3) in the combined group (P < 0.01). The time to reach the peak temperature of the treatment area was 21.7 ± 5.0 (s) in the MWA group and 10.3 ± 5.0 (s) in the MEUS + MWA group (P < 0.01). The time to reach the peak temperature of the treatment area was 21.7 ± 5.0 (s) in the MWA group and 10.3 ± 5.0 (s) in the MEUS + MWA group (P < 0.01). The time to reach the peak temperature of the treatment area was 21.7 ± 5.0 (s) in the MWA group and 10.3 ± 5.0 (s) in the MEUS + MWA group (Conclusions These results suggested MEUS treatment alone may significantly reduce tumor blood perfusion and led to a sharp rise in the local temperature of the treatment area to a higher PT using MEUS + MWA with higher rates of necrosis and apoptosis of cancer cells without severe liver function damage, which might be a safe strategy for treating HCC.

ACTR-57. A PHASE 1/2 STUDY TO EVALUATE THE SAFETY AND EFFICACY OF BLOOD-BRAIN BARRIER (BBB) OPENING WITH A NINE-EMITTER IMPLANTABLE ULTRASOUND DEVICE IN RECURRENT GLIOBLASTOMA PATIENTS PRIOR TO CARBOPLATIN

Abstract The blood-brain barrier (BBB) limits penetration of systemically administered therapies to brain tumors and peritumoral tissue and may be responsible for the failure of numerous drugs in recent clinical trials for glioblastoma (GBM). Low intensity pulsed ultrasound (LIPU), with concomitant intravenous microbubble injection (DEFINITY®), can temporarily disrupt the BBB for 6–24 hours and allow for enhanced delivery of drugs to the brain. In previous clinical studies, a 1-cm ultrasound (US) device, SonoCloud-1, was implanted in a burr hole of recurrent GBM patients (n=27) and used to disrupt the BBB prior to carboplatin infusion at AUC5. This ongoing pilot phase 1/2 study (NCT03744026) aims to evaluate the safety and efficacy of transient opening of the BBB by LIPU in rGBM with the SonoCloud-9, a larger nine-emitter implantable device, designed to cover the tumor and surrounding infiltrative regions. The ultrasound device is implanted in a 6 cm x 6 cm bone flap window after tumor debulking surgery. Patients receive a 270-second sonication every month with concomitant microbubble injection to disrupt the BBB followed by carboplatin infusion at AUC4-6. Magnetic resonance imaging (MRI) is performed to verify safety and extent of BBB disruption. This study began enrollment in early 2019 and is an international, open-label, single arm, multi-site, dose-escalation and expansion trial to evaluate the safety of escalating sonication volume (“dose”) with concurrent carboplatin. The number of activated emitters in the implant (3, 6, 9) is increased using a 3 + 3 dose escalation design. Safety data of the escalation phase of the study will be presented.

Cavitation-induced traumatic cerebral contusion and intracerebral hemorrhage in the rat brain by using an off-the-shelf clinical shockwave device

Traumatic cerebral contusion and intracerebral hemorrhages (ICH) commonly result from traumatic brain injury and are associated with high morbidity and mortality rates. Current animal models require craniotomy and provide less control over injury severity. This study proposes a highly reproducible and controllable traumatic contusion and ICH model using non-invasive extracorporeal shockwaves (ESWs). Rat heads were exposed to ESWs generated by an off-the-shelf clinical device plus intravenous injection of microbubbles to enhance the cavitation effect for non-invasive induction of injury. Results indicate that injury severity can be effectively adjusted by using different ESW parameters. Moreover, the location or depth of injury can be purposefully determined by changing the focus of the concave ESW probe. Traumatic contusion and ICH were confirmed by H&E staining. Interestingly, the numbers of TUNEL-positive cells (apoptotic cell death) peaked one day after ESW exposure, while Iba1-positive cells (reactive microglia) and GFAP-positive cells (astrogliosis) respectively peaked seven and fourteen days after exposure. Cytokine assay showed significantly increased expressions of IL-1β, IL-6, and TNF-α. The extent of brain edema was characterized with magnetic resonance imaging. Conclusively, the proposed non-invasive and highly reproducible preclinical model effectively simulates the mechanism of closed head injury and provides focused traumatic contusion and ICH.

Localized Blood–Brain Barrier Opening in Ovine Model Using Image-Guided Transcranial Focused Ultrasound

Transcranial application of focused ultrasound (FUS) combined with vascular introduction of microbubble contrast agents (MBs) has emerged as a non-invasive technique that can temporarily create a localized opening in the blood–brain barrier (BBB). Under image-guidance, we administered FUS to sheep brain after intravenous injection of microbubbles. BBB opening was confirmed by performing dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) to detect the extravasated gadolinium-based magnetic resonance contrast agents. Through pharmacokinetic analysis as well as independent component analysis of the DCE-MRI data, we observed localized enhancement in BBB permeability at the area that subjected to acoustic pressure of 0.48 MPa (mechanical index = 0.96). On the other hand, application of a higher pressure at 0.58 MPa resulted in localized, minor cerebral hemorrhage. No animals exhibited abnormal behavior during the post-FUS survival periods up to 2 mo. Our data suggest that monitoring for excessive BBB disruption is important for safe translation of the method to humans.

Safe long-term repeated disruption of the blood-brain barrier using an implantable ultrasound device: a multiparametric study in a primate model.

OBJECTIVE The main limitation to the efficacy of chemotherapy for brain tumors is the restricted access to the brain because of the limited permeability of the blood-brain barrier (BBB). Previous animal studies have shown that the application of pulsed ultrasound (US), in combination with the intravenous injection of microbubbles, can temporarily disrupt the BBB to deliver drugs that normally cannot reach brain tissue. Although many previous studies have been performed with external focused US transducers, the device described in the current work emits US energy using an unfocused transducer implanted in the skull thickness. This method avoids distortion of the US energy by the skull bone and allows for simple, repetitive, and broad disruption of the BBB without the need for MRI monitoring. The purpose of the present study was to determine if the BBB can be safely and repeatedly disrupted using such an implantable unfocused US device in a primate model. METHODS An 11.5-mm-diameter, 1-MHz, planar US device was implanted via a bur hole into the skull of 3 primates (2 Papio anubis [olive] baboons and 1 Macaca fascicularis [macaque]) for 4 months. Pulsed US sonications were applied together with the simultaneous intravenous injection of sulfur hexafluoride microbubbles (SonoVue) every 2 weeks to temporarily disrupt the BBB. In each primate, a total of 7 sonications were performed with a 23.2-msec burst length (25,000 cycles) and a 1-Hz pulse repetition frequency at acoustic pressure levels of 0.6-0.8 MPa. Potential toxicity induced by repeated BBB opening was analyzed using MRI, PET, electroencephalography (EEG), somatosensory evoked potential (SSEP) monitoring, behavioral scales, and histopathological analysis. RESULTS The T1-weighted contrast-enhanced MR images acquired after each sonication exhibited a zone of hypersignal underneath the transducer that persisted for more than 4 hours, indicating a broad region of BBB opening in the acoustic field of the implant. Positron emission tomography images with fluorine-18-labeled fluorodeoxyglucose (FDG) did not indicate any changes in the cerebral metabolism of glucose. Neither epileptic signs nor pathological central nerve conduction was observed on EEG and SSEP recordings, respectively. Behavior in all animals remained normal. Histological analysis showed no hemorrhagic processes, no petechia, and extravasation of only a few erythrocytes. CONCLUSIONS The studies performed confirm that an implantable, 1-MHz US device can be used to repeatedly open the BBB broadly in a large-animal model without inducing any acute, subacute, or chronic lesions.

Breast tumor response to ultrasound mediated excitation of microbubbles and radiation therapy in vivo.

Acoustically stimulated microbubbles have been demonstrated to perturb endothelial cells of the vasculature resulting in biological effects. In the present study, vascular and tumor response to ultrasound-stimulated microbubble and radiation treatment was investigated in vivo to identify effects on the blood vessel endothelium. Mice bearing breast cancer tumors (MDA-MB-231) were exposed to ultrasound after intravenous injection of microbubbles at different concentrations, and radiation at different doses (0, 2, and 8 Gy). Mice were sacrificed 12 and 24 hours after treatment for histopathological analysis. Tumor growth delay was assessed for up to 28 days after treatment. The results demonstrated additive antitumor and antivascular effects when ultrasound stimulated microbubbles were combined with radiation. Results indicated tumor cell apoptosis, vascular leakage, a decrease in tumor vasculature, a delay in tumor growth and an overall tumor disruption. When coupled with radiation, ultrasound-stimulated microbubbles elicited synergistic anti-tumor and antivascular effects by acting as a radioenhancing agent in breast tumor blood vessels. The present study demonstrates ultrasound driven microbubbles as a novel form of targeted antiangiogenic therapy in a breast cancer xenograft model that can potentiate additive effects to radiation in vivo.

Low Dose Focused Ultrasound Induces Enhanced Tumor Accumulation of Natural Killer Cells

Natural killer (NK) cells play a vital antitumor role as part of the innate immune system. Efficacy of adoptive transfer of NK cells depends on their ability to recognize and target tumors. We investigated whether low dose focused ultrasound with microbubbles (ldbFUS) could facilitate the targeting and accumulation of NK cells in a mouse xenograft of human colorectal adenocarcinoma (carcinoembryonic antigen (CEA)-expressing LS-174T implanted in NOD.Cg-PrkdcscidIl2rgtm1Wjl/SzJ (NSG) mice) in the presence of an anti-CEA immunocytokine (ICK), hT84.66/M5A-IL-2 (M5A-IL-2). Human NK cells were labeled with an FDA-approved ultra-small superparamagnetic iron oxide particle, ferumoxytol. Simultaneous with the intravenous injection of microbubbles, focused ultrasound was applied to the tumor. In vivo longitudinal magnetic resonance imaging (MRI) identified enhanced accumulation of NK cells in the ensonified tumor, which was validated by endpoint histology. Significant accumulation of NK cells was observed up to 24 hrs at the tumor site when ensonified with 0.50 MPa peak acoustic pressure ldbFUS, whereas tumors treated with at 0.25 MPa showed no detectable NK cell accumulation. These clinically translatable results show that ldbFUS of the tumor mass can potentiate tumor homing of NK cells that can be evaluated non-invasively using MRI.