- Meeting abstracts
- Open Access
International Society for Therapeutic Ultrasound Conference 2016
Tel Aviv, Israel. 14-18 March, 2016
Journal of Therapeutic Ultrasound volume 5, Article number: 15 (2017)
O1 The role of bubbles and cavitation in therapy ultrasound
Basic Radiological Sciences Division, Department of Radiology, University of Michigan, Ann Arbor, Michigan, USA
When exposed to sufficiently high ultrasound pressures, microbubbles can be generated spontaneously in tissue and undergo inertial cavitation where collapses result in physical effects. These effects range from petechial haemorrhage to complete cellular disruption, termed Histotripsy, depending on ultrasound parameters. This presentation will explore the mechanisms associated with histotripsy along with the tissue effects and the wide range of potential applications for this mechanical disruption method.
O2 Challenges for clinical trials in therapeutic ultrasound, the need for an evidence base, & trial design
Stanford School of Medicine, Stanford, California, USA
In this lecture, we will compare the design of clinical trials that led to approval of MR guided focused ultrasound for the treatment of uterine fibroids and osseous metastases. The impact of these trials on the evidence base, and thus on adoption by users and coverage by insurers will be compared. We will also review the process of expanding approved FUS applications, either via investigator- or industry-initiated studies or through off-label clinical use.
O3 Prostate HIFU – current status
SonaCare Medical, Indianapolis, Indiana, USA
In this lecture, present status of focused ultrasound for the treatment of localized prostate cancer ablation will be discussed. High intensity focused ultrasound (HIFU) has been used for the ablation of prostate over two decades and has treated thousands of prostate cancer patients. Meanwhile, prostate cancer management is undergoing significant improvements as molecular markers, targeted biopsy and advanced multi-parametric MRI are routinely used to accurately localize the prostate cancer. These advances offer a unique opportunity for focal ablation of localized prostate cancer with HIFU as it plays a significant role in reducing morbidity and treatment cost. This presentation will focus on hardware design, software architecture and HIFU features of the devices. Presentation will demonstrate localization of prostate with ultrasound imaging, treatment planning with 3D volumetric rendering of the prostate with ultrasound and MRI fusion techniques for focal treatment and finally HIFU dose setting with guidance using real time Tissue Change Monitoring (TCM) with the Sonablate device. The presenter will encourage exchange of ideas and discussion for research topics.
O4 Enhancement of drug delivery - clinical challenges and solutions
Constantin Coussios1, Paul C. Lyon1, Michael Gray1, Christophoros Mannaris1, Marie de Saint Victor1, Eleanor Stride1, Robin Cleveland1, Robert Carlisle1, Feng Wu4, Mark Middleton3, Fergus Gleeson2
1Institute of Biomedical Engineering, University of Oxford, Oxford, United Kingdom; 2Department of Radiology, Churchill Hospital, Oxford, United Kingdom; 3Department of Oncology, University of Oxford, Oxford, United Kingdom; 4HIFU Unit, Churchill Hospital, Oxford, UK
There are four key clinical challenges in optimizing drug based strategies: (i) achieving prolonged blood circulation of the therapeutic to enable active or passive accumulation in the target tissue; (ii) mediating triggered release or activation, or active accumulation of the therapeutic, to maximize its concentration at the target site whilst reducing off target side effects; (iii) enabling successful transport of the therapeutic from the blood stream into the target tissue, and achieving a homogenous distribution in that target tissue and (iv) where necessary, further enabling penetration of the therapeutic into the cell. The potential of therapeutic ultrasound with, or without, sonosensitive microparticle or nanoparticle formulations to address these challenges will be explored.
O5 Neuromodulation with ultrasound for beginners
Institut Langevin, Paris, France
In this lecture, the use of transcranial low intensity focused ultrasound for neuromodulation will be discussed. A historical review will be presented, with an emphasis on the experimental setups and the acoustical parameters. Models ranging from slice cultures to intact rodents and primates will be presented, together with recent trials on humans. Potential mechanisms will be described. Based on our own experience, exciting successful neuromodulation as well as disappointing failures will be presented.
O6 Thermometry in ultrasound fields, challenges & solutions in vivo, ex vivo & everywhere else!
Kim Butts Pauly
Stanford School of Medicine, Stanford, California, USA
This talk will cover image-based thermometry methods for guiding focused ultrasound. MR Thermometry based on the proton resonance frequency shift with temperature is linear and reversible in aqueous tissues and is utilized in clinical practice with common temperature resolutions of 1°C. Ultrasound based thermometry based on the speed of sound change with temperature can be used to 45°C in aqueous tissue. Both methods are sensitive to the presence of fat within the aqueous tissue, as well as motion. This talk will cover these basic concepts as well as their use in clinical practice.
O7 Motion compensation
Center for Imaging Sciences, Imaging Division, University Medical Center, Utrecht, the Netherlands
Motion leads to several challenges for HIFU treatment. In this lecture, the effects of respiratory, cardiac, and peristalsis related motion on MRI thermometry will be discussed. In addition, methods for tracking the moving target with the HIFU will be described, as well as gating strategies.
O8 MRgFUS crossing the chasm from proof of concept to mainstream treatment alternative
InSighTec, Haifa, Israel
MRgFUS treatment is a disruptive, non-invasive, outpatient treatment alternative that is capable to treat tumours and functional disorders under real-time monitoring and control via MR thermometry.
Transforming this breakthrough technology from the lab to a mainstream treatment alternative requires gaining the support and the agreement to change by a whole array of stakeholders in different areas some interrelated and some conflicting.
The stakeholders’ current position, changes they will need to go through and possible changes engines are mentioned below:
▪ Physicians (surgeons) need to transform into image guided surgeons where knowledge and understanding of the disease play a dominant role in the procedure outcome. The benefit is the confidence in the safety and efficacy coupled with income that wouldn’t decline. The change engine could be the patients and the payers.
▪ Payers should benefit from covering MRgFUS by saving cost and addressing patients’ demands. The change engine in this case should be the patients, governments and physicians.
▪ Governments should see the benefit of very fast recovery, very low level of adverse events and productivity enhancement. In this case patients and physicians should drive the change.
▪ Patients should benefit from safer treatment, fast recovery next day back to your life, minimal trauma and morbidity. The significant benefit to them should transform them to the dominant driver of this change. They will need to influence physicians, payers and providers to adopt this new treatment.
▪ Providers should adopt the technology and provide this treatment since the data exist proving safety and efficacy, proven cost savings and physicians and patients demand.
The current Medical ecosystem is biased against new technologies since the incumbent system/treatments are reimbursed while the new technologies are not. Could governments perform economic analysis and if found beneficial (example: saving money and improving productivity) decide on limited 2 years reimbursement during which RCT data will be collected based on which private insurance will decide to cover. This model should incentivize the physician, payers and providers to try the new technology.
O9 MR guided focused ultrasound treatment of soft tissue tumours of the extremities
Stanford School of Medicine, Stanford, California, USA
In this lecture, the use of MR guided focused ultrasound for the treatment of soft tissue and osseous tumours will be discussed. Results of treatment of these different types of tumours will be reviewed, including lessons learned from challenging patient treatments. Technical aspects of all parts of a treatment, including patient preparation, positioning, imaging, planning, thermometry, and methods of evaluation, will be described. The talk will also focus on methods developed to address these current challenges and opportunities for future development.
O10 Non-invasive, non-destructive FUS-induced neuro-modulation assessed by recording auditory evoked potentials – initial experience in small/large animals
Shirley Sharabi1,2, Dianne Daniels1,2, David Last1, David Guez1, Yoav Levy3, Alexander Volovick3, Javier Grinfeld3, Itay Rachmilevich3, Talia Amar3, Zion Zibly1, Yael Mardor1,2, Sagi Harnof1,2
1Sheba Medical Center, Ramat-Gan, Israel; 2Tel-Aviv University, Tel-Aviv, Israel, 3InSighTec, Haifa, Israel
MR guided focused ultrasound (MRgFUS) has been extensively studied in recent years as a non-invasive treatment modality. Initial clinical trials have indicated promising treatment response to ablative FUS treatments of patients with brain tumours, neuropathic pain, essential tremor, obsessive compulsive disorder, and Parkinson’s disease. Apart from the ablative applications of FUS, this technology has been extensively evaluated for less destructive applications such as thrombolysis, blood–brain barrier disruption for increased drug delivery, and recently also neuro-modulation.
The objective of the presented study was to demonstrate non-invasive, non-destructive, reversible FUS-induced neuro-modulation in small (rats) and large (pigs) animals by inducing temporary suppression of auditory evoked potentials.
All animal experiments were performed under full anaesthesia. Rats were anesthetized by Xylazine/Ktalar and pigs were anesthetized by Propofol. Rat’s heads were shaved prior to treatment but they did not undergo craniotomy. Pigs underwent craniotomy to avoid FUS reflection/aberration by the skull. EEG was recorded using 3 small cup-shaped electrodes attached to the skin/dura with metal particles-containing gel for optimal sound conductivity (Fig. 2).
The audio stimulation system consisted of a pulse generator connected to the EEG trigger input and to speakers placed near the animal ears, producing a square-wave form at 10 KHz, resulting in 150 “click” sounds per min (Fig. 2). Each measurement consisted of 200 repetitions enabling acquisition of a full measurement in 1’20” min.
The ExAblate Neuro system (InSightec, Tel Aviv, Israel) is a combination of a standard MRI scanner and a FUS delivery system. The FUS device is in the shape of a helmet consisting of 1024 transducers which deliver US energy in the form of “sonications”. The system is designed to provide real-time therapy, planning, thermal dosimetry, and closed loop therapy control. Treatment starts with conventional MRI scans, displayed on the ExAblate computer, used to determine regions of interest of the target volume. During the procedure, the beam path is periodically reviewed to confirm the planned direction through the tissue. The set of sonication volumes is sequentially applied to cover the entire planned volume. The current experiments were performed with a modified ExAblate version developed for neuro-modulation as part of the MAGNET programs supported by the Israeli Ministry of Commerce.
Baseline auditory evoked potentials were recorded by EEG prior to FUS treatment, with the animals in the prone position. The animals were then placed in the supine position, with the skull dipped in degassed water at the centre of the FUS system, for localization MRI scanning followed by FUS treatment. The animals were then returned to the prone position for continuous post-treatment EEG recordings. Rats which did not show recovery of the auditory evoked potentials 30–60 min post treatment were monitored again 48 hours or 1 week post treatment.
The animals were treated by FUS for 52 sec using the Exablate Neuro system at 220 KHz, 12 W, and 100 ms on/ 2900 ms off pulses. Two rats were treated in the thalamus region (targeted at deep auditory tracks) and another two in the frontal cortex region (targeted at peripheral auditory tracks). Two sham rats underwent a similar procedure without activation of the FUS system. One pig was treated in the thalamus region and another in the right motor cortex region.
Auditory evoked potential EEG signals shapes varied from one animal to the other but all were detected 2–10 ms after the trigger. The maximal peak-to-peak height was calculated for each measurement.
The sham rats showed no significant change in the auditory evoked potentials EEG signal.
The rats treated in the thalamus regions showed 50% and 65% suppression of the baseline auditory evoked potentials EEG signal. The first showed no recovery 2 hours post treatment with full recovery measured 1 week post treatment. The second showed no recovery for 1 hour post treatment and full recovery 48 hours post treatment. The rats treated in the cortex regions showed 50% and 67% suppression of the baseline EEG signal. The first showed no recovery 30 min post treatment with full recovery measured 1 week post treatment. The second showed initial recovery 14 min post treatment reaching full recovery within 28 min post treatment.
The pig treated in the thalamus region showed 90% suppression of the baseline signal with no recovery 30 min post treatment. The second pig, treated in the cortex region, showed complete suppression of the baseline signal immediately post treatment with initial recovery noted 18 min post treatment, reaching full recovery 63 min post treatment (Fig. 3).
Our preliminary results suggest that reversible neuro-modulation by non-invasive FUS is feasible. Full recovery was noted in all 4 treated rats and in 1 of the 2 treated pigs. Unfortunately we were not able to monitor the first pig for more than 30 min post treatment.
O11 Biophysical dissection of ultrasonic neuromodulation mechanisms
Michael Plaksin, Yoni Weissler, Shy Shoham, Eitan Kimmel
Faculty of Biomedical Engineering & Russell Berrie Nanotechnology Institute, Technion – Israel Institute of Technology, Haifa, Israel
Low intensity US can noninvasively suppress or excite central nervous system (CNS) activity using different combinations of stimulation parameters. While applications are already emerging, the underlying biophysics remains unclear regarding the relative contribution of possible mechanisms: extracellular bubble cavitation, thermal effects, acoustic radiation pressure and US-induced intramembrane cavitation within the bilayer membrane (the bilayer sonophore or BLS model). Interestingly, both radiation pressure and intramembrane cavitation can induce plasma membrane capacitance changes. Here, we use detailed predictive modelling and find that only intramembrane cavitation can explain all the observed aspects of ultrasonic neuromodulation.
We analyzed the relevant experimental literature using modified Rayleigh–Plesset intramembrane cavitation BLS biomechanics and acoustic radiation pressure gradients (RPG) - induced membrane dynamics. By coupling these biomechanical models to biophysical membrane models we predict dynamical biophysical responses of artificial bilayer membranes, and of three common neocortical single cell Hodgkin-Huxley type models: i) Regular Spiking (RS) cortical pyramidal neuron, ii) Fast Spiking (FS) cortical inhibitory neuron and iii) Low Threshold Spiking (LTS) cortical inhibitory neuron, RS-FS-LTS Hodgkin-Huxley based network model and CNS axon model. In addition, live brain tissue RPG subjected areal strains were evaluated in a viscoelastic brain model.
Only the Neuronal Intramembrane Cavitation Excitation (NICE) models were able to explain US-induced action potential generation through BLS-type pulsating nano-bubbles inside the bilayer plasma membrane: the leaflets' periodic vibrations induce US-frequency membrane capacitance and potential oscillations, leading to slow charge accumulation across the membrane (on a time scale of tens of milliseconds), until action potentials are generated. In contrast, the analysis of RPG-induced membrane capacitance variations associated with membrane area changes explain artificial membrane results, but were found to be highly unlikely sources for neural excitation, when considering the areal strains expected to form in brain tissue during normal sonication. Further, the NICE-LTS inhibitory neurons show a much higher relative sensitivity to sparse ultrasonic stimulation compared to the other neurons, resulting from their T-type voltage gated calcium channels. This model-based prediction was found to explain the results of a significant body of suppression and excitation experimental studies, including in humans.
These results provide a unified theoretical framework for a large body of experiments in multiple preparations across the field of US neuromodulation, lending further support to the hypothesis that intramembrane cavitation is responsible for ultrasonic neuromodulation. They could thus pave the way towards new CNS therapeutic protocols, using the only method that currently allows targeted non-invasive neuromodulation with millimetre spatial resolution essentially anywhere in the brain.
O12 Ultrasonic stimulation of mammalian retina in-vitro
Omer Naor1,2, Nairouz Farah3, Shy Shoham2
1ELSC Center for Brain Sciences, Hebrew University, Jerusalem, Israel; 2Biomedical Engineering, Technion- Israel Institute of Technology, Haifa, Israel; 3Faculty of Life Sciences, Bar Ilan University, Ramat Gan, Israel
Following previous in vivo stimulation of the retina, we aimed to achieve a first direct measurement of the response of mammalian retinal neurons to ultrasonic (US) stimuli, and to study and characterize this response.
We coupled a high-density phased array (986 elements on a 25x35 mm2 area) to a system for multi-electrode-array (MEA) recording with 256 contacts. Mouse retinas were dissected and placed on the MEA, and sonicated at 2.3 MHz, applying varying durations and intensities, as well as stimulated by light. The acquired data were processed to detect action potentials (spikes) elicited by retinal ganglion cells, and analysed to reveal the relations between the stimuli and the responses.
We found prominent spike responses for stimuli in the range of 4.3-7.3 W/cm2 and 0.5-1 s, which disappeared when the focus was steered 1.5 mm away. Furthermore, we found that the relation between the response strength and the stimulation intensity, or duration, followed a logistic sigmoid curve, while the response latency was described by a decreasing exponent. Lastly, we found indications that the observed responses to US stimuli are related to the "2nd OFF" component in the responses to light stimuli.
These findings are the first direct demonstration of the response of the mammalian retina to US stimulation. The properties of the US transducer and the stimulation frequency indicate that non-invasive US stimulation of human retina is feasible, and may potentially evolve as an important tool for diagnosis and treatment of retinal diseases.
O13 Motor response elicitation and pupil dilation using megahertz-range focused ultrasound neuromodulation
Christian Aurup1, Hermes Kamimura2,1, Shutao Wang1, Hong Chen1, Camilo Acosta1, Antonio A. Carneiro2, Elisa E. Konofagou1
1Columbia University, New York, New York, USA; 2Universidade de São Paulo, São Paulo, Brasil
Using transcranial focused ultrasound for the modulation of brain activity has been identified as a possible non-invasive means of treating neurological disorders. Most studies involving sedate rodents use frequencies in the kilohertz range, which allow for optimal transmission of acoustic power through the skull. The trade-off of using lower frequencies involves a lack of target specificity. Higher frequencies must be used in order to modulate activity in a more highly-specified manner. This study demonstrates that focused ultrasound in the megahertz range can be used to evoke motor- and cognitive-related responses in mice under deep anaesthesia by targeting specific brain structures. Contralateral-paired hind limb movements were observed when stimulating cortical regions, demonstrating the ability of MHz-range FUS to stimulate activity in highly-localized brain regions. Additionally, pupil dilation was observed when deep-seated anxiety-related structures were targeted, demonstrating the ability of FUS to modulate cognitive activity in a highly-specified manner.
For this study, wild-type adult male mice were anesthetized with intraperitoneal injections of sodium pentobarbital (65 mg/kg) and fixed in a stereotaxic frame. A single-element FUS transducer with fundamental frequency of 1.94 MHz was fixed to a 3D positioning system for accurate navigation through the brain. A 6x6 mm grid centred +2 mm rostral of the lambda skull suture was sonicated in a random order using a centre frequency of 1.9 MHz, pulse repetition frequency of 1 kHz, 50% duty cycle, 1 second pulse duration, 1 second inter-pulse interval for a total of 10 pulse repetitions. The acoustic pressure applied was varied in order to evaluate thresholds for eliciting physiological responses like motor movement, eye movement, or pupil dilation. Motor movements were validated using video recordings and electromyography via needle electrodes implanted into the biceps femoris of both hind limbs. Videos were recorded using a high-resolution camera focused at the right eye and processed to measure eye movements or changes in pupil size.
The minimum acoustic pressure required to elicit motor movements was 1.45 MPa when targeting the somatosensory cortex, calibrated using an excised mouse skull. Higher pressures increased the success rate from 20% (at the 1.45 MPa threshold) to 70% (1.79 MPa). Targeting eye-motor and anxiety related regions of the brain elicited eye movements and pupil dilations up to 20%. Sonicating the superior colliculus resulted in both eye movement and pupil dilation at a lower threshold pressure (1.20 MPa) than the hippocampus and locus coeruleus which required pressures greater than 1.80 MPa.
This study successfully demonstrated that MHz-range transcranial focused ultrasound can be used to elicit motor- and cognitive-related physiological responses with high specificity in mice in vivo. It was also shown that the success rate of stimulation increased with acoustic pressure for motor movements associated with cortical activity modulation but highly depends on the region of the brain targeted. These findings emphasize the complex and yet to be determined mechanism of action involved in ultrasonic neuromodulation.
O14 Thermal dose effects by MR-guided focused ultrasound on the pig brain tissue - preliminary results
Dong-Guk Paeng1,3, Zhiyuan Xu2, John Snell1, Anders H. Quigg1, Matthew Eames1, Changzhu Jin3, Ashli C. Everstine4, Jason P. Sheehan2, Beatriz S. Lopes5, Neal Kassell1
1Focused Ultrasound Foundation, Charlottesville, Virginia, USA; 2Neurosurgery, University of Virginia, Charlottesville, Virginia, USA; 3Ocean System Engineering, Jeju National University, Jeju, Korea (the Republic of); 4Biology, University of Virginia, Charlottesville, Virginia, USA; 5Pathology, University of Virginia, Charlottesville, Virginia, USA
The objective of this research is to investigate the effects of thermal dose (TD) delivered by magnetic resonance-guided focused ultrasound (MRgFUS) on in vivo pig brain tissue. In current clinical applications of transcranial MRgFUS systems, continuous acoustic wave emission is used to heat brain tissue to peak temperatures over 58°C. However, there are some situations where it has proven difficult to reach the desired peak temperature due to high absorption of acoustic energy by skull bone. There are reports that thermal effects on tissue are well correlated with thermal dose, which suggest that treatment delivery could be prescribed in terms of thermal dose rather than peak temperature or electric/acoustic power. It is also been demonstrated that the thermal dose threshold for permanent tissue damage is about 240 cumulative equivalent minutes (CEM) at 43°C for most of tissue. Currently available transcranial MRgFUS systems only allow the prescription of acoustic power and duration. In order to investigate the effects of thermal dose on in vivo brain tissue, we have developed a closed-loop control system to allow prescription thermal dose. This system monitors tissue heating via MR thermometry and provides pulse width modulation of output acoustic power in order to hold target tissue at a fixed temperature, and hence receives a nearly constant dose rate.
A FUS system (ExAblate 4000 Neuro 650 kHz system, InSightec) was used for sonication and an MRI system (Discovery MR75-3.0T, GE Medical systems) was used for thermometry and pre- and post-imaging. A closed-loop control system was implemented on a personal computer to control pulse width modulation of the FUS system acoustic power in order to maintain a specified temperature based on the MR thermometry. Accumulated thermal dose was calculated in real time and used to stop the sonication so that a prescribed thermal dose was delivered to the targeted tissue. Phantom studies were performed to test the control system to prepare for animal experiments. One acute and six chronic experiments (with three day survival) were conducted to observe the effects of TD on pig brain by behaviour observation and post MR imaging of the brain (1 hour and 70 hours post procedure). Craniotomy was performed to create an acoustic access window, and sonication was applied on 4 spots in the thalamus of each pig. Histology was also performed to compare it with MR imagery. Temperature in the pig brain tissue was estimated by rectal temperature for the MR thermometry baseline. TD was varied from 7 to 200 CEM while the target temperature was changed from 46 to 52 °C with appropriate acoustic power depending on target position and individual pig. This study was approved by the University of Virginia Institutional Animal Care and Use Committee.
From the acute experiment, we could observe the lesions on MR images after 1 hour of sonication and histology subsequently confirmed the lesions. For the chronic experiments, no obvious problem was observed in the behavior of any of the six animals. Eighteen sonication spots in 5 pigs were analyzed through MR images. One pig experiment failed to control temperature due to introduction of air bubbles between the brain and scalp during surgery procedure, and 2 sonication spots were excluded due to technical problems. Large tissue changes were observed in MR images in all 6 spots over 100 CEM.
The diameter of those tissue changes in MR T2-weighted axial images were measured and averaged to 2.9 ± 0. 4 mm. There is inconsistency in generating lesions for TD below 100 CEM. No lesion was shown in some lower TD from 7 CEM and 61 CEM, while some smaller lesions (<2 mm in lesion diameter) were shown in TD from 18 CEM to 85 CEM except one large tissue change of 3.5 mm in diameter at 31 CEM. Some tissue changes were shown in both post MR images after 1 hour and 70 hours of sonication, while some were visible only at the 70 hour time point. Histology of 3 pig experiments is now available and the histology reports support the tissue changes and lesions in MR images. Lesion diameters in MR T2-weighted axial images versus TD in CEM are shown in Fig. 6 for all the results from the chronic pig study.
These preliminary results from pig brain tissue generally confirmed the previous results from rabbit brain tissue in generating tissue changes over a certain TD, even though there are some differences in the FUS systems and the experimental procedures and analysis. For lower thermal dose below 61 CEM, there is significant variability in generating of tissue changes, while large tissue changes whose average diameter is 2.9 mm were observed in MR T2-weighted axial images for higher TD over 100 CEM, which were reported with similar tendency but a little difference in TD from the rabbit brain study. These results may contribute to open the way to prescribe the thermal dose rather than peak temperature or acoustic power for brain treatments, and expand the treatment envelope beyond the current limitations in selecting targets and patients. This project is ongoing and will be further pursued with additional experiments for consolidation of the results and analysis.
O15 In vivo feasibility study of boiling histotripsy with clinical Sonalleve system in a neurological porcine model
Thomas Looi1,4, Vera Khokhlova2, Charles Mougenot5, Kullervo Hynynen3, James Drake1
1Hospital for Sick Children, Toronto, Ontario, Canada; 2University of Washington, Seattle, Washington, USA; 3Sunnybrook Research Institute, Toronto, Ontario, Canada; 4University of Toronto, Toronto, Ontario, Canada; 5Philips Healthcare, Markham, Ontario, Canada
To determine if a clinical focused ultrasound system (Philips Sonalleve) can be used to perform mechanical liquefaction of brain tissue for neurological lesioning through a simulated fontanelle in a porcine model (simulating neo-natal patients). This work will determine the power of the system required to induce lesions using a boiling histotripsy (BH) pulsing protocol. Post-treatment, the lesion volume and border will be measured with MRI imaging and histological examination.
A porcine model was used as the in vivo model with a maximum weight < 6.8 kg (4.9 - 6.8kg). A horse-shoe incision and blunt dissection was used to expose the skull. A craniotomy was performed to create a 4–5 cm2 opening in the skull simulating the fontanelle in a neonatal patient. A degassed mixture of ultrasound gel and water (ratio 10:1) was poured on top of the dura to ensure good acoustic coupling. The scalp was sutured closed with 2–0 Vicryl cutting needle. The animal was placed supine feet first with the craniotomy centred about the Sonalleve V2 system with Flex-M surface coils. Pre-treatment T1-weighted (T1-w), T2-weighted (T2-w) and T2*-weighted (T2*-w) MRI imaging was conducted as a baseline. Each animal was treated at four cluster locations where each cluster consisted of seven sonication points; one point in the centre and six points uniformly distributed over a 4-mm diameter circle. The clusters were located approximately 15 mm deep in the brain, 7 mm off the midline, and separated by 14 mm in a rectangular geometry. In initial treatment on the first animal, the power was increased from 100 to 500 W for each cluster. After initial analysis, the treatment was repeated on second animal with refined power levels of 325, 350, 375, and 425 W. The treatment sequence consisted of 12000 pulses of 1.2 MHz frequency, 1 and 10 ms pulse duration, and 1% duty cycle for both 1 and 10 ms pulse duration. These protocols have been shown to generate BH lesions in ex vivo bovine liver in another Sonalleve system. During treatment, MR thermometry was used to monitor for surface, focal, and far field heating. A dedicated MATLAB-based interface was connected to the Sonalleve cavitation sensor to detect the signal generated during treatment points. After treatment, post T1-w, T2-w and T2*-w MRI scans were completed for comparison. The animals were euthanized, perfusion fixated and their brains were removed for histology. The brain specimens were cut at the centre for the treatment clusters to get a cross-sectional coronal view where each slice was 5 microns. The slides were stained with haematoxylin and eosin (H&E) and examined for lesion presence, blood and border definition.
A total of 4 piglets were sonicated with the following configuration: 1 piglet with 5 clusters (100 – 500 W), 2 piglets with 4 clusters (325-425 W) using 10-ms long pulses and 1 piglet with 4 clusters and 1-ms long pulses. For all power levels, the MR-measured temperature in the near or far field of the treatment was below the noise level. For 10-ms long pulses, 100 and 200 W acoustic powers, no noticeable imaging change was observed during sonication and post-treatment MR evaluation. As power was increased from 300 to 400 W, a temperature increase of up to 5C was measured at the focus. With more discrete power levels, it appeared at 375W that the lesion was more contained whereas higher power levels created wider areas of tissue change. It was observed the timing of MR magnitude of the target cluster changed as the power level increased where the tissue change occurred at 15 s at 300 W, 12 s at 350 W, 10 s at 375 W and 5 s at 425 W. After sonication was completed, the detected temperature rise decreased immediately versus dissipating over time. This would indicate the detected thermometry was due to a phase change of the tissue rather than temperature increase. During treatment, a high amount of lower broad band emissions at frequency < 1.2MHz were detected by a cavitation sensor. Post-treatment MR imaging showed that at power levels between 300 and 400 W, there were areas of hypointensity indicating the lesion. H&E staining confirmed the presence of the mechanical lesion where various anatomical targets were fractionated. Power levels were sufficient to rupture vessels and cause a focused area of haemorrhage at the treatment cluster. It showed that BH treatment dissolved the anterior ventricle wall with presence of elements of blood. H&E staining also showed that maximum lesion diameter was approximate 7 mm in coronal plane therefore the treatment borders matched the treatment plan. For the piglet treated with 1-ms long pulses, the post-treatment imaging change was not noticeable. However, at 375 and 400 W power, H&E slides showed two areas where there was a perforation of the anterior ventricle wall with a lesion size of up to 2 mm. It appeared that the shorter pulse duration generated smaller but more focused lesions.
This pilot study shows that the clinical Sonalleve system is capable of generating mechanical ablation of a brain tissue in an in vivo porcine model using boiling histotripsy pulsing scheme. The power threshold to initiate lesions in brain using 10 ms pulses (375 W) was found to be similar the power levels used in BH studies in ex vivo bovine liver and porcine kidney tissue at a similar depth in tissue (250 – 300 W) in a Sonalleve V2 system at the University of Washington. The treatments can be accelerated by using higher power outputs and shorter pulses. H&E histological evaluation showed that BH treatment caused rupture of vessels focally while also creating wider well defined areas of mechanical ablation with no damage to surrounding tissue. Additional work is underway to characterize the pressure levels generated by the Sonalleve to correlate the power and pressure for treatment.
O16 Musculoskeletal clinical applications of intense therapy ultrasound (ITU): part 1. Clinical study for plantar fasciitis
Michael Slayton1, Richard C. Amodei1, Keegan Compton1, Ashley McNelly2, Daniel Latt2
1Guided Therapy Systems, LLC, Mesa, Arizona, USA; 2School of Medicine, University of Arizona, Tucson, Arizona, USA
Chronic Plantar fasciitis (CPF) is a common cause of plantar heel pain that is a result of a degenerative process of the plantar fascia and its surrounding perifascial structures . It is the most common cause of heel pain, affecting 10% of the U.S. population, and one of the most common foot and ankle problems.
More than twenty different treatments have been used for plantar fasciitis. Conservative treatments (rest, ice, stretching and NSAIDs) have been shown to effectively treat symptoms but 10% of patients fail conservative management and continue to have symptoms within 12 months and beyond. Surgery consisting of partial PF release is often considered with 50% of patients having residual symptoms, in addition to surgical risk exposure.
High frequency ITU, a novel potential approach to treating Plantar Fasciosis, was studied for the creation of small thermal injuries noninvasively inside symptomatic Plantar Fascia (PF). It has been shown to initiate a tissue repair cascade and promote collagen generation in musculoskeletal tissue [2, 3]. A double blinded, randomized, sham controlled clinical study for ITU treatment of chronic Plantar Fasciitis has been conducted by IRB approved clinical protocol to access clinical efficacy of the procedure.
Custom 3.2 MHz high intensity (10 kW/cm2) ultrasound therapy system was designed and fabricated (GTS, Mesa, AZ, USA). Field simulations, testing and Schlieren images verified intensity, high focal pressure (17.3 MPa) and focal distance of 13–15 mm.
Each treatment consisted of 250–320 100 ms pulses creating matrices of small ablative thermal lesions of 4–5 joules at pre-programmed pitch of 1.6 mm. Each patient underwent two treatment sessions in 2 weeks, each treatment time did not exceed 12 minutes. ITU placebo group consisted of the same treatment with energy set to 0.
Treatment effects were assessed with diagnostic imaging ultrasound at 12 MHz (Spark, Ardent Sound, Mesa, AZ, USA) by a certified sonographer. Ultrasound images were analysed to determine symptomatic hypoechoic lesion size with PF.
Patient reported outcomes consisted of PROMIS physical function computer adaptive test (PF-CAT), PROMIS global health, Foot Function Index pain subscale (FFIPS) [4, 5] and a non-validated heel pain specific questionnaire.
Clinical protocol included (35) patients diagnosed with chronic heel pain due to Plantar Fasciitis (more than 3 months) and failed conventional therapy treatments.
Patients were randomized to standard therapy (anti-inflammatory pills, stretching and gel heel cups) plus ITU (“Treatment” group, n=26) of standard therapy plus sham ITU (“Control” group, n=9).
Primary investigator, sonographer and study coordinator administering the study were blinded to group assignments. P-values were calculated via 2-tailed paired T-tests for both treatment and control groups.
Patient-Reported Outcome Measures: Compared to the baseline assessment of Pain, the Treatment Group showed significantly improved pain scores compared to the Control (sham treatment) Group in follow-up visits including 12 weeks after the initial treatment.
Foot Function Index Pain Score: Compared to the baseline assessment, the treatment group pain scores also showed significant improvement compared to the sham group.
Diagnostic Ultrasound Imaging: During the 12 week follow-up period changes to the overall thickness of the PF were not statistically significant, while calculated volume size of hypoechoic lesions within the PF, just distal to the Calcaneus, showed significant change. For the experimental group (n=28) the average hypoechoic lesion volume reduction was followed and compared to the baseline measurements just before the first Treatment; 2 week follow-up and 2nd treatment date (−28%), 4 weeks (−50%), 6 weeks (−66%) and 12 weeks (−80%).
For the control group (n=10), the average hypoechoic lesion volume was followed and compared to the baseline just before the first Treatment; 2 weeks and 2nd sham treatment (+9%), at 4 weeks (+16%), 6 week (+29%) and 12 weeks (+31%). Unlike the experimental group, these lesions grew in size during the follow-up period.
P-values calculated for all outcome results discussed above for both treatment and control groups were below .01, showing the statistical significance of the results.
Results of the double blinded randomized, sham controlled study for the treatment of Plantar Fasciitis with ITU appeared to have statistically significant positive results within 12 weeks post-treatment in 80% of treated subjects.
Both quantitative measurements from diagnostic ultrasound imaging and applied standardized assessment protocols consisting of PROMIS PF-CAT, FFIPS along with Patient Reported Outcome Measures showed statistically significant coincidental improvements in treated subjects vs. control group.
Intense Therapeutic Ultrasound has shown potential for effective treatment of Chronic Plantar Fasciitis. Better designed studies with increased # of subjects will be considered to support ITU as an effective tool for the proposed clinical treatment.
 Neufeld S.K. and Cerrato R.; Plantar fasciitis: evaluation and treatment. J Am Acad Orthop Surg, June 2008, 16(6): p. 338–46.
 Slayton M. and Barton J.; Healing tissue response with ITU (Intense Therapy Ultrasound) in musculoskeletal tissue, feasibility study. Ultrasonics Symposium (IUS), 2014 IEEE International, Chicago, USA, p.1654–1657.
 Slayton M. H., Amodei R. C., McNelly A. and Latt D. L.; Intense Therapy Ultrasound (ITU) for the treatment of Chronic Plantar Fasciitis: Preliminary Results of Clinical Study. 37th International Conference of the IEEE Engineering in Medicine and Biology; Milan, Italy, August 2015.
 Rose, M., Bjorner, J.B., Becker J., Fries J.F. and Ware J.E.; Evaluation of a preliminary physical function item bank supported the expected advantages of the Patient-Reported Outcomes Measurement Information System (PROMIS). J. Clin. Epidemiology, January2008, 61(1): p. 17–33.
 Budiman-Mak E., Conrad K. J. and Roach K. E.; The Foot Function Index; a measure of foot pain and disability. J. Clin. Epidemiology, 1991, 44(6): p. 561–579.
O17 Musculoskeletal clinical applications of intense therapy ultrasound (ITU): part 2. Initial results of clinical study for lateral epicondylitis
Michael Slayton1, Richard C. Amodei1, Keegan Compton1, John Kearney2
1Guided Therapy Systems, Mesa, Arizona, USA; 2The CORE Institute, Phoenix, Arizona, USA
Acute and Chronic pain of the Common Extensor Tendon (CET) region, lateral epicondylitis or tennis elbow is a common pathology of both athletes and non-athletes affecting up to 3% of the population at large , while the prevalence of chronic problems caused by overuse in tennis players can be as high as 40%. Elbow tendinopathy represents an important set of pathologies that account for lost recreation time, decreased quality of life, and work-related disability claims. Conservative treatment of the Epicondylitis or -osis is recommended as the initial strategy by most authors. This strategy includes identification and correction of possible etiological factors, and a symptom related approach. Generally, the initial treatment consists of a multifactorial approach that may include a combination of rest (complete or modified activity), medication (NSAIDs for Epicondylitis), stretching and strength training. More aggressive treatments for CET include: Cortisone injection, Plasma Rich Platelets (PRP), Tenotomy, ESWT.
High frequency ITU, a novel potential treatment for CET, was studied for the creation of small thermal injuries noninvasively inside symptomatic Common Extensor Tendon (CET). It has been shown to initiate a tissue repair cascade and promote collagen generation in musculoskeletal tissue [2–5]. A blinded, randomized, clinical study for ITU treatment of chronic Lateral Epicondylitis has been conducted by IRB approved clinical protocol to assess clinical efficacy of the procedure.
Custom 4.5 MHz high intensity (47.9 KW/cm2) ultrasound therapy system was designed and fabricated (GTS, Mesa, AZ, USA). Field simulations, testing and Schlieren images verified intensity, high focal pressure (37.9 MPa) and focal distance of 6 mm.
Each treatment consisted of 80 14 ms pulses creating matrices of small ablative thermal lesions of 1 joule at manually targeted area set by diagnostic ultrasound imaging. Each subject underwent two treatment sessions 4 weeks apart. Each treatment time did not exceed 10 minutes. Treatment effects were assessed with diagnostic imaging ultrasound at 20 MHz (Spark, Ardent Sound, Mesa, AZ, USA) by a certified sonographer. Ultrasound images were analysed to determine changes in the peri-tendon region, including hypoechoic areas, calcifications and dependent free fluid.
Subject reported outcomes consisted of PRTEE survey , physical examination, Universal Analog Visual Pain Score17 and a Patient Reported Satisfaction Survey .
Clinical protocol includes 25 subjects diagnosed with chronic Tennis Elbow, or Lateral Epicondylosis (more than 3 months) and failed conventional therapy treatments.
Subjects were subjected to standard therapy (stretching and strength exercises, hot and cold compresses and compression support) plus ITU.
Primary investigator, sonographer and study coordinator administering the study were blinded to group assignments. P-values were calculated via 2-tailed paired T-tests at each visit of the clinical study.
The results presented below are initial findings for the first 12 subjects currently being followed through the study.
PRTEE: Patient Reported Tennis Elbow Evaluation Final Score is a weighted Pain Score based on 15 questions grouped into 3 categories: Overall Pain, Functional Disability and Usual Activities. Subjects respond to each question with a Pain Score of 0–10. Each category is then summed and weighted with a maximum score of 100 (Overall Pain 100%, Functional Disability 50% and Usual Activities (50%), n=12, Fig. 12.
Self-Reported Outcome Measures Surveys show a significant improvement and treatment satisfaction with Subjects reporting improvements in elbow pain 100%, improvement in Daily function 83% (vs. 17% no improvement) and treatment satisfaction 83% (vs. 17% not satisfied), n=6, Fig. 13.
Universal Analog Pain Scores also show progressive reduction (−3 on a 10 point scale) throughout the same period, n=12, Fig. 14.
Diagnostic Ultrasound Images: Diagnostic Ultrasound Images show a consistent increase of free fluid 2 weeks after the first treatment, with a progressive reduction in free fluid at 8 weeks in subjects with no to mild peri-tendon calcifications. These subjects correlated well with improving PRTEE survey scores. Subjects with little or no improvement in PRTEE scores consistently presented with moderate to severe peri-tendon calcifications.
P-values calculated for the above reported outcomes were not statistically significant for Visits 2 and 3 (P>0.05) while results for Visits 4 and 5 demonstrated P< 0.05.
Feasibility of Intense Therapeutic Ultrasound treatments of chronic pain in CET region has been established with the initial results (n=12) of the ongoing clinical study.
Significant reduction of pain scores per activities (PRTEE) and Self-Reported Outcome Measures (83% improvement) with average Universal Pain Scores reduction from 5.0 to 2.0 were statistically significant (p<0.05) at 8 and 12 weeks post treatment.
 Hong QN, Durand MJ, Loisel P. Treatment of lateral epicondylitis: where is the evidence? Joint Bone Spine 2004; 71(5):369–373.
 White, W. M., I. R. Makin, et al. (2007). Selective creation of thermal injury zones in the superficial musculoaponeurotic system using intense ultrasound therapy: a new target for noninvasive facial rejuvenation." Arch Facial Plast Surg 9(1): 22–29.
 Gliklich R, White WM, Barthe PG, Slayton MH, Makin IRS. Clinical pilot study of intense ultrasound (IUS) therapy to deep dermal facial skin and subcutaneous tissues. Arch Facial Plast Surg 2007; 9:88–95.
 Slayton M., Barton J, Feasibility of Modulating Healing Tissue Response by ITU (Intense Therapy Ultrasound) in Musculoskeletal Tissue ASLMS 2014 Annual Conference.
 Slayton M., Barton J., Healing tissue response with ITU (Intense Therapy Ultrasound) in musculoskeletal tissue, feasibility study, Ultrasonics Symposium (IUS), 2014 IEEE International, pp. 1654–1657. DOI 10.1109/ULTSYM.2014.010
 Rompe JD1, Overend TJ, MacDermid JC. Validation of the Patient-rated Tennis Elbow Evaluation Questionnaire.J Hand Ther. 2007 Jan-Mar; 2007 (1):3–10: quiz 11.
 Rose, M., et al., Evaluation of a preliminary physical function item bank supported the expected advantages of the Patient-Reported Outcomes Measurement Information System (PROMIS). J Clin Epidemiol, 2008. 61(1): p. 17–33.
O18 Clinical experience of intra-operative high intensity focused ultrasound in patients with colorectal liver metastases. Results of a phase ii study.
David Melodelima1,2, Aurelien Dupre2, Yao Chen2, David Perol2, Jeremy Vincenot1, Jean-Yves Chapelon1, Michel Rivoire2,1
1LabTAU - U1032, INSERM, Lyon, France; 2Centre Leon Berard, Lyon, France
Managing colorectal liver metastases (CLM) is a major clinical challenge, and surgery remains the only potentially curative treatment. Five-year survival rates of up to 51% have recently been reported. However, only 10–20% of patients are eligible for surgery, which is often precluded by the number, size and/or location of metastases, or because the necessary resection will leave insufficient volume of functional liver. Radiofrequency ablation (RFA) is the main technology that has been used in association with surgery as a tool to expand the number of patients who may be candidates for liver directed therapy. However, RFA has several limitations. There is a risk of inadequate treatment due to the heat sink effect of blood flow, RFA does not allow reliable real-time monitoring, and it require intra-parenchymal introduction of a probe. Moreover, only small hepatic volumes can be targeted. These limitations could explain the high rates of local recurrence seen after RFA.
High intensity focused ultrasound (HIFU) has been proven effective in a wide range of clinical applications, especially prostate cancer. The ablation achieved by conventional HIFU is small and ellipsoidal. The dimensions vary according to transducer characteristics but are typically 1–3 mm (transverse) and 8–15 mm (along beam axis). In clinical practice, hundreds of superimposed ablations are required; and the procedure may take up to two hours. Even so, HIFU has several potential advantages in the treatment of liver tumours: there is no need to puncture the parenchyma, the extent of the thermal lesions achieved is not reduced by hepatic perfusion, and it is possible to monitor the effects of therapy in real time. However, extra-corporeal treatment of the liver is difficult because presence of the ribcage may stop propagation of ultrasound waves and respiratory motion may cause targeting problems. HIFU treatment of CLM needs to be improved, and reducing the duration of surgical intervention by increasing the size of ablated fields is particularly important. A HIFU device enabling destruction of larger liver volumes has been developed based on toroidal transducers.
Preliminary in vitro and preclinical work demonstrated the potential, feasibility and safety of such HIFU ablations. During laparotomy in a porcine model we demonstrated that this HIFU device achieves reproducible ablations with an average volume of 7 cm3 (with 20 mm diameter and 25 mm long axis) in 40 seconds. Such preclinical work has to be translated into clinical practice through controlled trials, and the aim of this study was to assess the feasibility and safety of HIFU ablation in patients undergoing hepatectomy for CLM, as well to collect efficacy and accuracy data. This study is registered with Clinical-Trials.gov (NCT01489787).
This study was a prospective, single-centre phase I/II study evaluating the feasibility, safety and accuracy of HIFU during surgery in patients with CLM. The protocol was reviewed and validated by a national ethics committee (CPP Sud-Est IV) according to French and European directives. Since this study was the first use in man of intra-operative hepatic HIFU, ablations were made only in areas of liver scheduled for resection. This allowed real-time evaluation of HIFU ablation while protecting participating patients from any adverse effects related to this new technique. The transducer has a toroidal shape 70 mm in diameter and is divided into 32 ultrasound emitters of 0.13 cm2 operating at 3 MHz. The radius of curvature is 70 mm. A 7.5 MHz ultrasound imaging probe was placed in the centre of the device and was used to guide the treatment. The imaging plane was aligned with the HIFU focal zone.
Six patients were included in the Phase I. Two single thermal ablations were created in each patient. Thirteen patients were included in Phase IIa and two HIFU ablations were to be placed precisely in a target previously identified in ultrasound images (step 1) and then at distance (step 2) from a target. Five patients were included in Phase IIb until now. HIFU ablations were created to ablate metastases (20 mm maximal diameter) with safety margins in all directions. The exposure time varied from 40 s to 370 s according to the diameter of the metastases to be treated.
In agreement with preclinical studies, the demarcation between ablated and non-ablated tissue was clearly apparent in ultrasound images and histology. The dimensions measured on ultrasound imaging were correlated (r=0.88, p<0.0001) with dimensions measured during histological analysis. All HIFU ablations were obtained in 40 seconds. The average dimensions obtained from each HIFU ablation were a diameter of 21.0 ± 3.9 mm and a depth of 27.5 ± 6.0 mm. The phase IIa study showed both that the area of ablation could be precisely targeted on a previously implanted metallic mark and that ablations could be created deliberately to avoid such a mark. Ablations were achieved with a precision of 1–2 mm. In Phase IIb, one metastasis of 10 mm in diameter was ablated in 40 seconds with safety margins. Using electronic focusing metastases of 2 cm in diameter were ablated with safety margins (>3 mm in all directions) in 370 seconds. The dimensions of HIFU ablations created in 370 s were a diameter of 48 mm and a long axis of 51 mm.
This new HIFU device safely achieved large volume liver ablations in 40 s, with a precision of one to two millimetres under real-time monitoring. HIFU ablations of small metastases (<20 mm) and peri-lesional healthy liver were successfully created with planned safety margins of at least 3 mm in all directions.
O19 Chemotherapy in oral cancer
Wei Guo, Guoxin Ren, Guofeng Shen
Department of Oral Maxillofacial and Head Neck Oncology, Shanghai 9th People's Hospital, Shanghai, China
To evaluate the efficacy and the main side-effects during the clinical trial of this new ultrasound hyperthermia system combined with chemotherapy in oral cancer, meanwhile, to observe the preliminary clinical response of this combined therapeutic modality.
Thirty four cases of oral squamous cell carcinoma entered this clinical trial, from which 23 had advanced oral carcinoma and were treated with new ultrasound hyperthermia system combined plus docetaxel–cisplatin–fluorouracil regimen (test group). Eleven patients only received chemotherapy with docetaxel–cisplatin–fluorouracil regimen (control group). The thermo-index were detected during the course of hyperthermia, the chief-complain of the patients were also recorded. The systemic physiological, biochemical and immunological index were tested before and after the treatment respectively. The therapeutic response was estimated 1 month after 2 cycles of the treatment.
Twenty three cases of oral squamous cell carcinoma enrolled the clinical trial of local ultrasound hyperthermia combined with chemotherapy. 230 times of ultrasound hyperthermia in total were performed. The ultrasound hyperthermia system operated smoothly, no malfunction was found. The main thermo-index were: the maximum heating temperature was108 F, the average heating temperature was 106 F, the minimum heating temperature was 104 F, the fraction of heating time more than 108 F was 0.46, the average treatment time was 37.74±8.88min. PR+CR was 71% (test group). The main local side-effects were low-grade pain (6/23). The incidence of adverse effects was similar between both study groups, no bone marrow suppression (over III).
The system combined with docetaxel–cisplatin–fluorouracil regimen is effective and safe in the treatment of advanced oral cancer. The main side-effects of local ultrasound hyperthermia combined with chemotherapy are low-grade pain or tolerable pain. There is no immune function and obtains satisfying short-term response.
O20 Non-thermal, non-cavitational, 20kHz Ultrasound applicators in wound healing
Michael Neidrauer1, Leonid Zubkov1, Michael S. Weingarten2, David J. Margolis3, Peter A. Lewin1
1The School of Biomedical Engineering, Science and Health Systems, Drexel University, Philadelphia, Pennsylvania, USA; 2Department of Surgery, College of Medicine, Drexel University, Philadelphia, Pennsylvania, USA; 3Biostatistics and Epidemiology, University of Pennsylvania, Philadelphia, Pennsylvania, USA
This talk examines the challenges associated with the design of clinically viable ultrasound applicators operating at the relatively low frequency (20 kHz) and intensity (<100 mW/cm2, spatial peak, temporal peak) levels, and tailored to treatment of chronic wounds, such as venous or diabetic ulcers. These challenges were associated with the architecture and weight, and principle and efficiency of operation, including electrical power consumption. The ultimate goal of this work was to test the efficacy of the applicators in human subjects.
A fully wearable Band-Aid™-like, dial-in delivery, battery-operated ultrasound applicator was designed. The applicator included light weight (<25g) piezoelectric flexural transducer and was powered by 10-12V fully rechargeable lithium-ion batteries (total weight <200g); it was able to operate for up to 4 hours between re-charging. To emphasize the uniqueness of the design, it might be useful to note that typically, the thickness of the capacitive piezoelectric element is inversely proportional to the frequency, therefore a 20 kHz element would need to be 10 cm thick. Such element would be bulky and require hundreds of volts (demanding a large power amplifier), in excitation signal thus eliminating any chance of being a portable design. To overcome this, a mechanical displacement amplifier, which translates 2 MHz ultrasound waves into 20 kHz output at the desired pressure amplitude (55 kPa; i.e. 100 mW/cm2) with only 12 volts excitation was chosen as a preferred solution.
The applicators were extensively tested to ensure that the ultrasound field energy was below the level needed to generate inertial cavitation and any temperature elevation that would exceed 1°C. Also, the uniformity of the acoustic field distribution was verified. The pilot study included 32 individuals between ages of 18 and 80 having venous (n=23) or diabetic (n= 9) wounds that remained open for a minimum of 8 weeks. In compliance with the IRB study protocol the subjects were randomly assigned to either treatment or control group, with an equal chance of being assigned to receive active ultrasound treatment or sham (current standard care). Treatment sessions lasted 15 minutes and were administered once a week for a period of 12 treatments, or until the wound’s closure. Clinical efficacy was evaluated by measuring the reduction in wound area over time. For both etiologies, i.e. both venous and diabetic wounds the rate of closure was statistically faster (p<.05) in the treated group compared to the control group.
The study findings show that the ultrasound treated venous ulcer group had statistically improved (p<0.04) rate of wound size change (reduction of 14.3%/week) compared to the rate of wound size change for the control group (increase of 3.6%/week on average). Diabetic wound closure was achieved typically after 4 sessions for treated wounds, as opposed to 7 sessions for the control group. Time to heal was also statistically faster (p< .05) for treated wounds (~5 weeks) when compared to non-treated wounds (~12 weeks).
Overall, the results from this study support the notion that low frequency ultrasound treatment can successfully improve healing outcomes in chronic wounds with different morphology and etiology. The evaluated device used safe levels (<100mW/cm2 ISPTP) of ultrasound energy and featured unique portability, which opens possibility for personalized home treatment of chronic wounds in the future.
NIH Grant R01 EB9670, NSF 1064802, Wallace H. Coulter Foundation.
O21 Tests of thermal ablation with a 230 kHz transcranial MRI-guided focused ultrasound system in a large animal model
Nathan McDannold1, Jonathan Sutton1, Natalia Vykhodtseva1, Margaret Livingstone2
1Radiology, Brigham and Women's Hospital, Boston, Massachusetts, USA; 2Neurobiology, Harvard Medical School, Boston, Massachusetts, USA
Thermal ablation with transcranial MRI-guided focused ultrasound (TcMRgFUS) is being tested clinically for as an alternative to surgery for functional neurosurgery and brain tumour resection. The current TcMRgFUS system, which operates at 650–670 kHz, is limited by skull heating to a small central region in the brain. Use of a lower acoustic frequency will reduce skull heating, but at the same time the focal heating will decrease and the risks of uncontrolled cavitation (the formation of microbubbles) increase. The purpose of this study was to evaluate the feasibility of thermal ablation in nonhuman primates using a system that operates at a lower acoustic frequency and to determine whether it can increase the “treatment envelope” for TcMRgFUS.
The experiments were approved by our institutional animal committee. Thermal ablation with the 230 kHz ExAblate Neuro system (InSighTec) was tested over five sessions in three rhesus macaques. In each session a target in the thalamus was sonicated transcranially at 40–50 s at acoustic power levels ranging from 90–560 W. The TcMRgFUS system software modulated the acoustic power in real time with a closed-loop controller that maintained a low-level of acoustic emissions, which are correlated with cavitation activity. MR temperature imaging (MRTI) was acquired at 3T (LX, GE) in a single plane using a 14 cm surface coil (TR/TE: 29/13 ms; flip angle: 30°). Measurements of the peak temperature rise at the focus and on the outer brain surface were compared for the different animals as a function of the applied acoustic energy. For the brain surface we measured the average temperature at the hottest 5% of the voxels and over a two voxel wide strip and we also normalized the measurement by the outer skull area. Temperature measurements were used to calculate the accumulated thermal dose, which was then compared to post-sonication T2-weighted, T2*-weighted, and contrast-enhanced T1-weighted MRI. The focal and skull-induced heating on the brain surface were compared to an earlier study performed in macaques with a 650 kHz version of this system.
Focal heating sufficient to create an MRI-evident thermal lesion was achieved in 4/6 targets; the peak thermal dose exceeded 240 CEM43°C at these targets (Fig. 15). Heating at the focus was slightly higher than that measured on the brain surface. The focal heating increased linearly as a function of the applied energy at a rate of 3.2 ± 0.4°C per kJ (R2: 0.81). The surface area of the outer skull ranged from 47–55 cm2. For the hottest 5% of the voxels on the brain surface included in the MRTI imaging plane, the temperature rise increased linearly as a function of temperature at a rate of 126.6 ± 7.3°C per kJ/cm2. For a two voxel wide strip over the entire brain surface, this increase was 62.7 ± 7.5°C per kJ/cm2. The extent of MRI-evident changes (apparent oedema in T2-weighted MRI, BBB disruption post-contrast, no petechiae in T2*-weighted MRI) were consistent with 240 CEM43°C contours. One lesion imaged one week after FUS increased in size.
Analyses of the MRTI and post-sonication MRI suggest that the lesions were consistent with thermal mechanisms. The temperature rise increased linearly with the applied energy, and no evidence of cavitation-related petechiae were evident after sonication. The MRI-evident lesions were consistent with isodose contours drawn at 240 CEM43°C, a conservative threshold often used to guide thermal ablation. However, since it is known that thermal damage can take several hours to manifest in MRI and the lesion we imaged at one week increased in size, it is likely that the size of the lesion was underestimated by this dose value.
Similar tests in macaques with a version of this system operating at 670 kHz (Hynynen et al., Eur J Radiol 2006; 59: 149–56) measured skull-induced heating of 130°C per kJ/cm2 of outer skull surface, more than twice of that measured here (63°C per kJ/cm2). While no or minimal focal heating was observed at 670 kHz, with this 230 kHz system we were able to reach ablation-level thermal dose values. Thus, these preliminary results thus suggest that this low frequency system can expand the area of the brain that can be targeted for thermal ablation without overheating the skull. The closed-loop feedback system successfully maintained a low level of acoustic emission (and presumably microbubble activity) and immediately stopped the sonication when excessive levels were detected. However, additional work is needed to understand whether low-level cavitation activity played a role in the focal heating, to characterize the lesions in histology, and to examine whether safe cavitation levels can be maintained in tumours where the cavitation threshold may vary.
O22 Growth slowdown in a brain metastasis model by antibody delivery using focused ultrasound-mediated blood–brain barrier disruption
Thiele Kobus1,2, Yong-Zhi Zhang2, Natalia Vykhodtseva2, Nathan McDannold2
1Radiology and Nuclear Medicine, Radboud University Medical Center, Nijmegen, Netherlands; 2Radiology, Brigham and Women's Hospital, Boston, Massachusetts, USA
HER2-targeting antibodies prolong survival in patients with HER2-positive breast cancer metastases outside the brain. However, the response of brain metastases to these drugs is poor and it is hypothesized that the blood–brain barrier (BBB) limits drug delivery to the brain. We aim to improve delivery by temporary disruption of the BBB using focused ultrasound (FUS). Here we evaluate the treatment benefit of combining two antibody therapies that target the HER2-receptor with FUS-mediated BBB disruption in a breast cancer brain metastasis model.
MDA-MB-361 HER2-positive human cancer cells were injected in the right brain hemisphere of nude rats. The animals were divided in three treatment groups of 10 animals each: the control-group received no treatment; the antibody-only group received trastuzumab and pertuzumab (antibodies that target the HER2-receptor); and FUS+antibody-group received trastuzumab and pertuzumab in combination with FUS-mediated BBB disruption. The six weekly treatments started five weeks after tumour implantation, when the tumour diameter was around 2 mm. The FUS treatments took place in a 7T MR-scanner using a single-element, spherically-focused 690 kHz-transducer. Trastuzumab and pertuzumab were injected before the first sonication. At the start of each sonication (duration 60s, 10-ms bursts, burst repetition frequency 1 Hz), the ultrasound contrast agent Optison (100 μl/kg) was injected. The complete tumour was treated in 4 to 14 sonications that were separated 1 to 1.5 mm. Peak negative pressure amplitudes in water between 0.46 and 0.62 MPa were used.
Before and after the sonications, MR imaging was performed consisting of T2-weighted (T2w), T1w and T2*w imaging to locate the tumour, confirm BBB disruption and study the presence of hemorrhages. In two animals tumour leakiness was studied by comparing T1w imaging before and after gadolinium injection before the tumours were sonicated. The difference in signal intensity change in pre- and post-contrast T1w images was determined between the tumour and contralateral brain region (= SI%). In all FUS-treated animals BBB disruption was confirmed with contrast-enhanced T1w imaging and quantified using the same method as for the tumour leakiness. Pre- and post-sonication T2*w images were inspected for hypo-intense regions, which can indicate extravasated erythrocytes.
Every other week, high-resolution T2w imaging was performed to determine tumour volume. The growth rate (r) was determined by fitting the tumour volumes to the following formula: volume(t)=a*exp(r*t), in which t is the time in days. The growth rate of each tumour was determined for the treatment period (week 5 to 11) and the follow-up period (week 11 till sacrifice). An animal was classified as ‘responder’ if the growth rate was lower than the mean growth rate of the control animals minus two standard deviations.
The animals were euthanized if the tumour size exceeded 13 mm in diameter or if the condition of the animal was poor. From nine animals, histology was obtained (hematoxylin and eosin (H&E)) and the brains of five animals were stained for HER2.
BBB disruption was successful in all sessions with an average SI% of 21.2% (range 4.5 – 77.6%). The mean SI% of two tumours before BBB disruption during the six treatment weeks were 0.4% and 0.6%, indicating that the tumours were not leaky before disruption (Fig. 16). In 33% (20/60) of the FUS-sessions, regions were present that were clearly more hypo-intense on post- than on pre-sonication T2*w images, suggesting hemorrhages. In the remaining 67% of the sessions, no or a small difference in hypo-intensity was observed.
In the FUS+antibody-group, 4/10 animals were classified as responders during the treatment period (week 5 to week 11) with an average growth rate of 0.010±0.007, compared to 0.043±0.013 for the non-responders. There was no difference in the average SI% of the responding rats (21.8%±16.7) and the non-responding rats (20.7%±9.7). None of the control or antibody-only animals were classified as responder. When the FUS+antibody-animals are grouped, no significant differences in mean growth rates were observed between the control, antibody-only and FUS+antibody animals for the treatment period, nor for the follow-up period.
High-resolution T2w imaging showed that the tumour was homogenous in almost all animals till week 13–15, when cystic and necrotic areas started to develop. The tumours showed also a heterogeneous appearance on H&E stained sections and the complete tumour was expressing the HER2-receptor in the examined animals.
In this study, we demonstrate that FUS-mediated BBB disruption in combination with antibody therapy can slow down the growth of brain metastasis from breast cancer. As the tumours were not leaky before BBB disruption and no difference in growth rates was observed in the antibody-only group, the disruption of the BBB is necessary for drug delivery to these brain metastasis. Interestingly, only part of the rats responded to the treatment, the other animals had the same growth rate as the control-group. This is in line with a previous study (Park et al. 2012, J. Control. Release), where antibody therapy was combined with FUS in a different brain metastasis model and in part of the animals a strong response was observed, while the other animals did not respond. We did not observe a difference in tumour volume at the start of the treatment, in HER2 expression on histopathology, or in contrast-enhancement on MR images between the responders and non-responders to explain this. Better understanding of why certain animals respond is needed and will help in translating this technique to the clinic.
O23 Long term follow up of 6 essential tremor patients treated with MR-guided focused ultrasound thalamotomy
Michael Schwartz1,2, Yuexi Huang2, Nir Lipsman1, Jennifer Jain2, Martin Chapman2, Tejas Sankar3, Andres Lozano1, Kullervo Hynynen2
1Surgery (Neurosurgery), University of Toronto, Toronto, Ontario, Canada; 2Sunnybrook Health Sciences Centre, Toronto, Ontario, Canada; 3Surgery (Neurosurgery), University of Alberta, Edmonton, Alberta, Canada
To determine the factors influencing outcome after MR-guided focused ultrasound (MRgFUS) thalamotomy for essential tremor.
Between May 2012 and February 2013, 6 patients were treated with MRgFUS thalamotomy. The first 4 patients have been reported (Lipsman et al., Lancet Neurol. 2013 May; 12(5): 462–8). Prospective recording of data from preoperative screening to 3 month follow up according to an experimental protocol using the clinical rating scale for tremor (CRST), and then a 2 year follow up examination and assessment were done. Maximum temperature at the focus of sonication was determined and post-treatment day 1 MRI scans were reviewed for lesion size and location. A point derived by our current method of determining the probable location of the ventral intermediate nucleus (VIM) of the thalamus was taken as the starting point for each treated case and a deviation vector from that point to the centre of each lesion produced was plotted.
For the 6 patients, the mean CRST A scale (rest, posture and action) for the treated hand and arm prior to treatment was 7.2. At 1 week post-treatment, the mean CRST A was 0.67. At 3 months, the mean CRST A was 0.83 and at 2 years approximately 3.33. (At 2 years, the CRST A for 2 patients was estimated from a narrative account). Mean lesion size, excluding one very large lesion with a volume 2.7 times the mean volume of the other 5 patients, was 90.2 mm3. Only 2 patients had no decline in thalamotomy effect. Of these, one had a relatively large lesion of 107.5 mm3, which was located at the predicted location of the VIM nucleus. The other patient’s lesion was located 2.4 mm lateral to the predicted VIM location. He had prominent ataxia following his treatment. This subsided by 3 months. The patient with the large lesion, treated for left hand tremor, had persistent tingling of the left side of his mouth, and his left thumb and 3 fingers but excellent early relief of tremor. Although there was some persisting reduction of tremor at 2 years, he could no longer write, nor drink without spilling. The centre of his lesion was 1.3 mm lateral and 0.3 mm posterior to the expected VIM location. In its superior-inferior dimension his lesion measured 9.4 mm. The maximum temperature achieved at treatment was 63°C, higher than the mean of 58°C. This decline may, in part, be due to the progress of his condition, with increasing tremor on his untreated side and the onset of tremor in both legs. Two of the patients with a decline in function had relatively small lesions.
Not all patients have lasting benefit from MRgFUS thalamotomy. The effect of relatively large lesions may be more durable, but lesioning temperatures of greater than 60°C should likely be avoided.
O24 Skull bone marrow injury caused by MR-guided focused ultrasound for cerebral functional procedures
Michael Schwartz1,2, Robert Yeung1, Yuexi Huang1, Nir Lipsman2, Jennifer Jain1, Martin Chapman1, Andres Lozano2, Kullervo Hynynen1
1Sunnybrook Health Sciences Centre, Toronto, Ontario, Canada; 2Surgery (Neurosurgery), University of Toronto, Toronto, Ontario, Canada
To determine the factors that lead to bone marrow injury in the skulls of patients undergoing MRgFUS cerebral ablations for the treatment of movement disorders such as ventral intermediate (VIM) nucleus thalamotomy for tremor and pallidotomy for L-dopa induced dyskinesia (LID), and to follow the course of healing.
All patients undergoing functional cerebral ablations produced for the mitigation of movement disorders are followed with serial MRI scans according to study protocol to assess the evolution of the cerebral lesions. In one patient subjected to very high power sonication in the attempt to produce a pallidotomy for LID, skull lesions were noticed on a follow up MRI scan, produced 4 months after his MRgFUS procedure. A review of other treated patients is currently underway.
Multiple ovoid lesions throughout the calvarium, new since the immediate post-treatment MRI scan done January 30, 2015, were seen on the MRI scan done May 12, 2015 (Fig. 17). Their appearance resembles that of bone infarcts. The MRI scan was repeated on October 2, 2015. Many of the ovoid lesions were still visible. This patient underwent sonication increasing to a maximum power of 1100 W for 31 seconds. During the procedure, the scalp and skull were constantly cooled with flowing degassed water at 13 °C. Despite this sonication, the target locus in the globus pallidus reached only 48 °C.
High power and duration sonication for functional cerebral lesions may cause injury to skull bone marrow. A review of all patients treated with MRgFUS for movement disorders is currently underway to determine whether there have been other cases, and to determine the threshold for bone marrow injury.
O25 In vitro study using MR-guided focused pulsed ultrasound for destroying clots using thrombolytic drugs
Christakis Damianou1, Nikolaos Papadopoulos2
1Cyprus University of Technology, Limassol, Cyprus; 2City University, London, United Kingdom
In this paper an extensive study of using MR-guided focused pulsed ultrasound system is presented for the treatment of stroke using thrombolytic drugs in a model in vitro.
A single element spherically focused transducer of 4 cm diameter; focusing at 10 cm and operating at 1 MHz was used. The transducer was mounted in an MR compatible robotic system of 3 axes. The artery was modelled using a silicone tube. Tissue was modelled using agar-evaporation-silica gel. Coagulated blood was used to model thrombus. A thermocouple was placed in the thrombus in order to measure the thrombus temperature.
The effect of power, presence of bubbles, temperature, presence of agar-evaporation milk-siligal gel, time of sonication, pulse repetition frequency, presence of standing waves, flow velocity were investigated. The goal was to maintain a temperature increase of less than 1°C during the application of pulsed ultrasound (called safe temperature). With the application of ultrasound alone or thrombolytic drug alone there was no notable destruction of the thrombus.
With the combination of ultrasound and thrombolytic drugs sufficient destruction occurred after 30 min, but the rate of destruction of thrombus (mL/min) is considered low. Thus, the clinical use of focused ultrasound for sonothrombolysis despite the full parametric study that we performed is considered pessimistic.
O26 Low frequency in-vivo cavitation mapping
Alexander Volovick1, Javier Grinfeld1, Yoav Levy1, Omer Brokman1, Eyal Zadicario1, Ori Brenner2, David Castel3
1INSIGHTEC, Tirat Carmel, Israel, 2Weizmann Institute, Rehovot, Israel, 3Sheba Medical Center, Ramat Gan, Israel
Current trans-cranial Magnetic Resonance guided Focused Ultrasound Surgery (tcMRgFUS) treatments are limited to the targets that are located at the centre of the skull. Superficial and peripheral targets are prone to lower acoustic intensity reaching the focal point due to the geometry of the skull and the ultrasound transversal through it. A possible solution for this problem is reduction of the transmitting frequency. Reducing the frequency is a trade-off between higher skull penetration and lower tissue absorbing. In addition, lower frequency has a lower threshold to induce cavitation. Previously it was reported () that the transient cavitation threshold for the muscle tissue acts linearly with frequency as where Pth is the cavitation pressure threshold and f is the transmitting frequency. However this threshold was not reported to cause any unintentional pathological damage. In the presented work, different levels of acoustic intensities resulting in various occurrences of cavitation were applied to an in-vivo pig brain model, reaching various temperature levels and causing various pathological effects.
36 subjects underwent craniotomy and were sonicated using various ultrasonic parameters (ExAblate Neuro low frequency system, 220 Khz, by INSIGHTEC, Ltd). The temperature rise was measured using MR thermometry (1.5T GE HDx MRI by GE Healthcare, with an integrated Head Coil, by InSighTec) and cavitation signal was measured and recorded using integrated hydrophones. 20 subjects underwent 2 weeks follow up with post procedural MR imaging one, five, seven and fourteen days after the procedure. The brains were harvested, fixed in formalin and sliced to 3 mm slices. Macro-pathological slices for randomly selected subjects were also performed.
Figure 18 represents the graph of temperature rise as a function of applied energy. Figure 19 represents the graph of cavitation activity as a function of maximal acoustic power. Figure 20 shows a follow up imaging for 2 lesions performed on a single subject, Fig. 21 is a macro pathology slide for the subject presented in Fig. 20. In general the temperature rise grew linearly with the applied acoustic energy; the cavitation activity was linearly dependent to the maximum applied acoustic power. For sonications that were in the central cavitation activity area as presented on Fig. 19 the lesions were well defined and increased in size till the 5th day follow up and then reduced in size becoming scarf tissue as presented on Figs. 20 and 21. Macro pathology revealed that tissue rapture is seen on the micro level; however it is well defined and limited within the lesion area.
Cavitation threshold levels that were observed in brain agree with the levels reported in literature. Additional levels of cavitation were observed and associated with effects on tissue as seen in MRI and histology. The data collected suggests cavitation levels that can be applied while keeping lesioning effect to the confined area and avoiding haemorrhages in tissue. Integrating a real time control over the level of cavitation and keeping the level below the safety threshold results in safe and effective tissue ablation in brain.
O27 Real-time, transcranial passive cavitation mapping for monitoring of the focused ultrasound-induced blood–brain barrier opening in primates
Shih-Ying Wu1, Julien Grondin1, Wenlan Zheng1, Marc Heidmann1, Maria Eleni Karakatsani1, Carlos J. Sierra Sánchez1, Vincent Ferrera3, Elisa E. Konofagou1, 2
1Biomedical Engineering, Columbia University, New York, New York, USA; 2Radiology, Columbia University, New York, New York, USA; 3Neuroscience, Columbia University, New York, New York, USA
Real-time cavitation monitoring during the focused ultrasound (FUS) induced blood–brain barrier (BBB) opening is crucial in assessing and controlling the BBB opening outcomes and safety. Currently, passive cavitation detection using a single-element detector for quantification of the stable and inertial cavitation doses has shown good correlation with the opening volume and the molecular delivery efficiency in nonhuman primates (NHPs). However, an off-line magnetic resonance imaging (MRI) is required for confirming the targeting or opening area in the brain after the FUS procedure. It is therefore essential to develop transcranial cavitation mapping in providing the spatial distribution of cavitation intensity in real time in order to precisely assess and control the BBB opening outcomes with accurate targeting during the procedure. The aim of this study is to develop real-time cavitation mapping using time-exposure acoustics and passive sparse matrix beamforming with the performance evaluated in both the in vitro primate skull (NHP and human) experiments and the in vivo NHP experiments during BBB opening.
Similar to array-based passive cavitation detectors, a linear probe (L7-4, Philips) with a programmable data acquisition system (Vantage, Verasonics) was used to acquire cavitation signal passively during the sonication (frequency: 0.5 MHz, pulse length: 10 ms, PRF: 5 Hz, pressure: 150–600 kPa) with in-house made microbubbles (lipid-shelled and monodisperse with a diameter of 4–5 μm) and a single-element FUS transducer with a coaxially aligned flat-band hydrophone as a single-element passive cavitation detector. Time-exposure acoustics for an integration of a series of passive cavitation images over the exposure time reconstructed by dynamic receive beamforming using sparse matrix calculation in graphic processing unit (GPU; Tesla K40, NVIDIA) were developed for the reconstruction of passive cavitation maps in real time. For the in vitro experimental setup, a phantom with a channel of 4 mm in diameter and an infusion pump was used to mimic the vessel with microbubble circulation (a concentration of 2×105 bubbles/mL with a flow rate of 1 mL/min), and the FUS transducer and the linear array were both focused at the channel orthogonally. B-mode imaging with the linear array was performed before acquiring the passive cavitation signal in order to confirm the alignment of both the linear array and the FUS transducer to the channel. Three sets of passive cavitation maps were acquired using this in vitro setup: 1) without the skull, 2) with the NHP skull and 3) with the human skull placed between the phantom and the linear probe. The effects of acoustic pressure, exposure time and aperture size to the intensity and focal size of the cavitation maps were all evaluated, as well as the computational cost in GPU and the sensitivity through the skull. Furthermore, the in vivo cavitation maps were acquired during the sonication for BBB opening in NHPs (duration: 2 min) and compared with the BBB opening outcomes in the MRI.
The results of the in vitro experiments showed that the cavitation location corresponded to the microbubble disruption in B-mode images, and both the intensity and the focal area of the cavitation maps increased with pressure. Increasing the exposure time eliminated the interference outside of the focus and enhanced the focalization by minimizing the focal area, while both the intensity and the focal area reached a plateau at the exposure time of 62.5 μs. The focal size especially in the axial direction increased with decreasing aperture size, suggesting an improved focalization by using a larger aperture size of the array-based PCD. The computational time for the exposure time of 62.5 μs was 9.5 s, which can be decreased to 0.27 s in achieving real-time monitoring by decreasing the exposure time to 1.44 μs. By placing the skull, it was found that the cavitation signals were still detectable through the NHP and human skulls at 300 kPa and 600 kPa with the mapping system, respectively. For the in vivo experiments, the BBB opening in NHP were successfully monitored with passive cavitation mapping targeting at the caudate and the hippocampus, the deep subcortical structure in the brain.
A real-time cavitation mapping technique using time-exposure acoustics and passive sparse matrix beamforming has been developed with the performance and the sensitivity through the primate skull evaluated, and was used for monitoring of the BBB opening in NHPs. This novel transcranial monitoring technique providing both the spatial and intensity information of cavitation in real time during the FUS procedure is promising in assessing and controlling the targeting, treatment efficacy, and safety precisely.
O28 Blood brain barrier opening using focused ultrasound for the reduction of amyloid beta plaques in synergy with antibodies in a rabbit model fed with high cholesterol diet
Christakis Damianou, Marinos Yiannakou
Cyprus University of Technology, Limassol, Cyprus
An animal model that creates Amyloid beta (Ab) plaques in the brain was implemented by delivering high cholesterol diet in rabbits for 3 months. The goal was to reduce the plaques using focused ultrasound (FUS) in combination with external antibodies.
A single spherically focused transducer was used which operates at 1 MHz, has focal length of 10 cm and diameter of 3 cm. The rabbit was placed in a custom made MRI compatible positioning device. Theoflavin staining was used in order to measure the plaque load at the end of each experiment.
Using pulse FUS the blood brain barrier (BBB) was opened repeatedly up to 5 times at three day intervals. The opening of the BBB disruption was imaged using contrast-enhanced T1-weighted fast spin echo. By increasing the number of sessions, the number of plaques decreases (both for internal and external antibodies). With the use of FUS only (internal antibodies) the drop of average number of plaques/mm2 was reduced by 20% (in 5 sessions). The effect of external antibodies was more drastic. With 5 BBB sessions the average number of plaques/mm2 was reduced by 60%.
This study demonstrated that by opening the BBB, it will be possible to deliver internal and external antibodies to the brain, which eliminates Alzheimer disease (AD) plaques. More important by opening the BBB frequently (up to 5 times in this study) the reduction in the number of plaques is enhanced. Therefore FUS has the potentials to be used non-invasively for the treatment of AD.
O29 Correlation between down-regulation of p-glycoprotein and blood–brain barrier disruption in rat brain by mri-guided focused ultrasound and microbubbles
HongSeok Cho1, Hwayoun Lee1, Mun Han2, Jong-Ryul Choi1, Taekwan Lee1, Sanghyun Ahn1, Yongmin Chang2, Juyoung Park1
1Daegu-Gyeongbuk Medical Innovation Foundation, Daegu, Republic of Korea; 2Kyungpook National University, Daegu, Korea (the Republic of )
Blood–brain barrier (BBB) is composed of both physical barrier with tight junctions and functional barrier with active efflux transporters. Mechanism of the functional barrier is mediated by P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP) in brain endothelial cells. Past studies have shown that focused ultrasound (FUS) combined with microbubbles can disrupt the physical barrier of the BBB by exerting mechanical stress on the tight junctions; however, no study was performed to investigate the impact of FUS and microbubbles on the functional barrier of the BBB. Therefore, this study investigated the impact of BBB disruption induced by FUS and microbubbles on the expression of P-gp. We also investigated correlation between magnitude of the BBB opening and the down-regulation of P-gp.
A single target region was sonicated (0.5~0.7 MPa) transracially in one hemisphere in 3 rats using a 1 MHz FUS transducer; the other hemisphere served as a control. For the BBB disruption, 10 ms bursts were applied at 1Hz pulse repetition frequency (PRF) for 120 s and combined with IV injection of a microbubble ultrasound contrast agent (Definity 0.1 ml/kg). An MR contrast agent (Magnevist 0.4 mM/kg) and Evans Blue (0.15 ml/kg) were injected immediately after the sonication to indicate area of the BBB disruption in MR image and fluorescence spectroscopy, respectively. In order to measure the P-gp expression using a confocal fluorescence microscopy, the brains were fixed after perfusion and then stained immunohistochemically with a monoclonal antibody (C219) which reacts with a P-gp epitope.
A T1 contrast enhanced MR image and Evans Blue fluorescent intensity at the sonicated regions indicated localized BBB disruption (Fig. 22). Both the MR contrast intensity and the Evans Blue fluorescent intensity were significantly increased in the targeted regions compared to the control regions (p<0.001). The fluorescence intensity of the P-gp expression at the confirmed locations of the BBB disruption was reduced by an average of 63.2±18.4% compared to the control area in all three rats. From the three sonicated regions, a total of 31 locations were selected and the P-gp fluorescence intensities were measured to observe the correlation between the degree of the BBB opening and the P-gp expression. Both the Evans Blue intensity and the MR contrast intensity were significantly correlated with the P-gp expression intensity (r=−0.72, p<0.001; r=−0.62, p<0.001, respectively). Histologic analysis on the sonicated region of the brain tissue revealed no apparent damage in the endothelial cells, and no significant amount of extravasated red blood cells was observed.
This study demonstrates that the BBB disruption induced by FUS and microbubbles reduces the expression of P-gp, and the level of the down-regulation of P-gp is significantly correlated with the magnitude of the BBB opening. These results suggest FUS + microbubble as a promising mean for the brain drug delivery through the BBB by overcoming both the physical and the functional barrier of the BBB.
O30 MR-guided focused ultrasound blood–brain barrier disruption through an intact human skull in a rat model using a clinical body system
Nicholas Ellens1, Ari Partanen1,2, Keyvan Farahani1,3, Raag Airan1
1Radiology, Johns Hopkins University, Baltimore, Maryland, USA; 2Philips, Andover, Massachusetts, USA; 3National Cancer Institute, Bethesda, Maryland, USA
MR-guided focused ultrasound (MRgFUS) can be used for a large range of non-invasive therapies using mechanical and thermal mechanisms. For instance, it has been demonstrated that a combination of low intensity focused ultrasound and synthetic microbubble scan be used to safely, locally, and transiently disrupt the blood–brain barrier (BBB).
Typically, transcranial MRgFUS thermal ablation requires large aperture phased arrays with many elements operating at lower frequencies and higher powers than body systems due to high transmission losses through the skull. However, in low pressure applications at low duty cycles, such as BBB disruption or neuromodulation, it may be feasible to use 'body' systems that have smaller aperture transducers with fewer elements . The objectives of this study were to use a clinical body MRgFUS system to:
Quantify the ultrasound transmission and focal distortion through a human cranium at different orientations and in different focus locations inside the skull to assess the possible treatment envelope with such a device.
Demonstrate the ability to disrupt the blood–brain barrier (BBB) in rat neurovasculature through a human cranium.
A clean, degassed human cranium (gift of Dr. Quiñones-Hinojosa, JHU Neurosurgery) was mounted in degassed water above a 256-element phased-array transducer (14 cm focal length) of a clinical body MRgFUS system (Sonalleve V2, Philips, Vantaa, Finland). Hydrophone measurements were made with a lipstick hydrophone (Onda Corp., Sunnyvale, USA) mounted to a 3D stage, both in water and through the human skull in 11 locations (three orientations, 3–4 depths at each, ranging between the centre of the cranium and 2 cm from the skull surface). 40-cycle sonications at 1 MHz were applied at different acoustic powers ranging from 5 to 20 W (water) and 20 to 500 W (trans-skull). In each location, the full-width at half-maximum (FWHM) of the ultrasound focal point (in three dimensions) and its peak negative pressure were measured. This setup and the locations sampled are shown in Fig. 23.
Hydrophone measurements demonstrated a great deal of variation in ultrasound transmission with changing transducer/skull incidence angle and skull thickness. The pressure attenuation ranged from −5.8 dB to −9.3 dB (mean +/−standard deviation, −8 dB +/− 1 dB), and the FWHM varied between 1.83 mm and 3.79 mm (2.2 mm +/− 0.3 mm in the anterior/posterior direction, 2.7 +/− 0.7 mm in the left/right direction). The insertion losses as a function of depth at different orientations are shown in Fig. 24. The dorsal midline skull (with the ultrasound beam passing through the sagittal and coronal sutures) was thicker than either of the angled approaches (passing through the parietal and temporal bones) and experienced a greater insertion loss for most sonication target depths. For both angled approaches, the insertion loss decreased as the focus was moved away from the skull surface. For the centered approach, the insertion loss increased slightly as the focus moved closer to the skull surface. It appears that the intervening skull thickness dominates the attenuation, though the angle of incidence seen by transducer elements also affects transmission.
Gadolinium enhancement in the brain on post-sonication T1-weighted MRI indicated successful BBB disruption. Disruption accuracy is shown in Figs. 25 and 26. For sonications of sufficient pressure to produce BBB disruption, the region of gadolinium enhancement did not appear to be shifted at all from the desired target. For higher estimated in situ pressures (>0.55 MPa), the disruption region was larger and less uniform, as evident in the 0.58 MPa point in Fig. 25. Even without element-by-element refocusing, the focusing quality and targeting precision appeared to be adequate. Post-experiment, there were no signs of gross damage to the animals to suggest significant off-target sonication.
Transcranial FUS focal point distortion was minimal despite the lack of element-by-element transducer refocusing. The clinical MRgFUS body transducer and driving electronics had sufficient power and aperture to generate the in situ pressures for BBB disruption through a human skull, using a variety of clinically practical approaches and patient orientations.
Though transcranial thermal ablation typically requires high acoustic powers, a lower operating frequency, and large aperture arrays with high number of elements, this study demonstrates that a clinical body MRgFUS system with a smaller transducer may be a safe and feasible alternative for non-invasive BBB disruption and other low-pressure therapeutic ultrasound transcranial applications, potentially offering a wide treatment envelope.
 Hynynen, K., et al. (2001). Radiology, 220(3), pp. 640–646.
 Airan, R. D., et al. (2015). 23rd ISMRM.
O31 Interim results from a phase 1 clinical trial to disrupt the blood–brain barrier by pulsed ultrasound
Alexandre Carpentier3,4, Michael Canney1, Alexandre Vignot1, Cyril Lafon2, Jean-Yves Chapelon2, Jean-yves Delattre4,5, Ahmed Idbaih5
1CarThera, Lyon, France; 2INSERM, U1032, LabTau, Lyon, France; 3Assistance Publique Hopitaux de Paris, Hopital de la Pitie Salpetriere, Department of Neurosurgery, Paris, Paris, France; 4Universite Paris, UPMC, Paris, France; 5Assistance Publique Hopitaux de Paris, Hopital de la Pitie Salpetriere, Department of Neuro-Oncology, Paris, France
Pulsed ultrasound, coupled with peripheral injection of microbubbles, has been shown in pre-clinical studies to be an effective method for enhancing the delivery of chemotherapy to the brain. In this work, an intra-skull implantable ultrasound device, SonoCloud®, was developed for temporarily disrupting the BBB. The device was implanted in patients with recurrent glioblastoma (GBM) undergoing systemic carboplatin chemotherapy in a Phase 1 clinical trial and the safety of repeated BBB disruption was assessed.
A Phase 1 clinical trial began in July 2014 at a single centre at the Hospital Pitie Salpetriere in Paris, France. Patients with recurrent GBM with an enhancing volume of less than 35 mm in diameter were included in the trial. Participants were implanted with a 11.5-mm diameter biocompatible 1 MHz ultrasound transducer, which was fixed into the skull bone thickness. The device was either implanted during a regular surgical resection of the enhancing region or during a unique ambulatory procedure under local anaesthesia. Once a month, the device was connected to an external radiofrequency generator using a transdermal needle, and patients received a two minute pulsed ultrasound exposure in combination with systemic administration of an ultrasound contrast agent (7 min mean total duration procedure). BBB disruption was assessed immediately after sonications using dynamic T1-weighted imaging with a gadolinium based MR contrast agent. Systemic intravenous injection of carboplatin chemotherapy was delivered immediately following MR imaging. Patients followed a progression of ultrasound dose in which the pressure was increased from 0.5 to 0.8 MPa throughout the course of the study.
As of July 2015, eleven patients had been included in the study with a total of 25 BBB disruption procedures performed. No adverse effects were observed in patients treated. BBB opening was clearly observed in 12/25 treatments on contrast-enhanced T1w imaging. The procedure was safely tolerated in all patients. No evidence of acute haemorrhage, petechia, ischemia or oedema was observed in post-sonication SWAN T2*, Diffusion or FLAIR MRI sequences.
The BBB was safely opened by pulsed ultrasound using an implantable ultrasound device in patients with recurrent glioblastoma. Additional follow up and recruitment will be used to further evaluate the safety and potential efficacy of such an approach.
Work supported by CarThera and the Hospital Pitie Salpetriere.
O32 Investigation of temperature dependent changes in MR signal intensity, t1 and t2* in cortical bone
Henrik Odéen2, Bradley Bolster1, Eun Kee Jeong2, Dennis L. Parker2
1Siemens Healthcare, Salt Lake City, Utah, USA; 2Radiology, University of Utah, Salt Lake City, Utah, USA
For MR guided focused ultrasound treatments in or close to bone, such as for transcranial applications focusing through the intact skull bone and treatments of bone metastases, significant heating can occur in the bone due to its high ultrasound absorption. MR imaging of bone is in general challenging due to the short T2 relaxation time of bone. For MR temperature imaging the short T2 also severely decreases the accuracy that can be achieved with the standard proton resonance frequency shift method. Instead researchers have investigated the temperature dependence of the MR signal intensity (SI) and T1 relaxation time for temperature monitoring (1–4). Miller (1) and Fielden (3) et al. showed that the SI from cortical bone decreases with increasing temperature using ultrashort echo time (UTE) pulse sequences, and Ramsay et al. (2) found that, contrary to what Miller and Fielden observed, the SI increases with increasing temperature using a short TE gradient recalled echo (GRE) pulse sequence. Han et al. (4) further showed that T1 increases with temperature, also using UTE.
In this work we investigate the temperature dependence of the SI (dSI/dT) and the T1 and T2* relaxation times (dT1/dT and dT2*/dT, respectively) using a 3D UTE pulse sequence to investigate which parameter has the highest sensitivity to temperature change.
All imaging was performed on a 3T MRI scanner (MAGNETOM PrismaFit, Siemens Healthcare, Erlangen, Germany) using a 3D UTE pulse sequence. The sequence utilizes radial, ramped sampling of k-space in 3D starting at the k-space centre after a 80 μs non-selective hard RF pulse, allowing TEs down to 50 μs. T2* was measured by performing an exponential fit to data acquired at TE = 50, 90, 130, 170, and 250 μs (other scan parameters are listed in Table 1). The 50 μs TE was also used for dSI/dT calculations. T1 was measured using the variable flip angle (VFA) method (5) with TR = 11 ms and FA = 8 and 36° (other scan parameters are listed in Table 1).
An approximately 4-cm long bovine femur (marrow and connective tissue removed) was placed in a phantom holder that allowed heated water to circulate around the bone, Fig. 27. One fiber optic probe measured the water temperature, and three probes were inserted in 1-mm diameter, 2-cm deep, holes drilled in the bone to measure the temperature of the bone. The water was warmed to ~22, 35, 50, and 65°C and data was collected when all 4 probes measured within 1°C. The whole setup was places in a 20-channel RF head coil for signal detection.
Figure 28 shows 2D maps of SI and T1 and T2* relaxation times for the 4 different temperatures. Mean and standard error values from a 9x9 ROI close to each probe is shown in Fig. 29, together with calculated changes in %/°C. A decrease in SI of 0.3-0.5%/°C, and increases in T1 and T2* of 0.5-0.9%/°C and 0.6-0.9%/°C, respectively, was observed. Using the temperature dependent spoiled GRE signal equation (6) and the observed values for dT1/dT and dT2*/dT, dSI/dT can be closely predicted.
The decrease in SI is in accordance with previously published results by Miller and Fielden. The measured change in T1 using UTE agrees well with the 0.6%/°C reported by Han, although we measured higher absolute T1 values (~160-175 ms at 25°C, compared to ~115-125 ms as reported by Han).
The effect of T1 and T2* on SI are counter-acting each other (both increasing), which reduces the sensitivity of dSI/dT. This may suggest that dT1/dT and dT2*/dT are more suitable candidates for bone MR thermometry, and Fig. 29 also shows higher sensitivity for relaxation times that for SI. However, it should be noted that SI can be detected from a single image, whereas T1 and T2* measurements utilize two or more images, therefor resulting in longer scan times.
Future studies will acquire data with longer TEs (out to ~10 ms) to investigate the difference in temperature sensitivity for shorter and longer T2 components in the bone. These results will be compared to the increase in SI with temperature observed by Ramsay using a TE ≈ 1 ms. The temperature dependence of the long and short T2* components can be found by a multi-exponential fit. Flip angle mapping will also be implemented to improve the accuracy of the T1 measurements (7).
O33 Spatially-segmented MRI brain and water bath reconstruction for undersampled transcranial mr-guided focused ultrasound thermometry
Pooja Gaur1, Xue Feng2, Samuel Fielden2, Craig Meyer2, Beat Werner3, William Grissom1
1Vanderbilt University, Nashville, Tennessee, USA; 2University of Virginia, Charlottesville, Virginia, USA; 3University Children's Hospital, Zurich, Switzerland
MR-guided focused ultrasound (MRgFUS) brain systems deliver targeted thermal energy into the brain using a hemispheric array of transducers that surround the head with an intervening water bath (Fig. 30a). During treatment the localized heating (hot spot) is measured from a change in image phase between baseline (pre-treatment) and dynamic (during treatment) images. Accelerating temperature mapping by undersampling k-space is desirable to increase spatiotemporal resolution and coverage, but is difficult to do with parallel imaging since coils must be placed outside the transducer, far away from the head. Multiple groups have instead developed accelerated temperature mapping methods that exploit temporal correlations between baseline and dynamic images [1, 2]. However, circulation of the water bath to cool the skull causes dynamic signal changes that are not captured by baseline images (Fig. 30b), which breaks those correlations and results in artefacts throughout the temperature maps. We propose a spatially-segmented approach for reconstructing temperature maps in brain MRgFUS, in which we separately estimate a water bath image without a baseline, and a temperature map in the brain with a baseline. The method can estimate artefact-free temperature maps from undersampled data during brain MRgFUS treatments using a single receive coil.
Our iterative approach alternates between updating the parameters of a k-space hybrid signal model which is fit in the brain region of the image , and a baseline-free estimate of the water bath image. The fitting of k-space hybrid brain model results in a phase drift-corrected brain image without the temperature phase shift and a sparse temperature phase shift map. An algorithm to fit the model is described in . The water bath is reconstructed using a POCS algorithm that alternately enforces data consistency, consistency with a water bath support mask (brain and water bath masks are obtained from a baseline image), and sparsity in the Coiflet domain using soft thresholding . Figure 30c illustrates the overall undersampled dynamic image model.
To test the method, a gel-filled human skull phantom was sonicated by an InSighTec ExAblate Neuro 4000 transcranial MRgFUS system (InSighTec Ltd, Haifa, Israel) while imaging with a GE 3T MR750 scanner (GE Healthcare, Waukeshaw, WI). 27 single-slice 2DFT gradient echo images were collected with the body coil and 28 x 28 x 0.3 cm3 field of view, 256 x 128 acquisition matrix, 30° flip angle, 13 ms TE, and 28 ms TR. Images and maps were reconstructed to a 128 x 128 matrix and retrospectively randomly undersampled by 2x, with full sampling over 22 central k-space lines. Temperature maps were reconstructed by fitting the k-space hybrid model to the entire image, or to the brain only with keyhole or POCS methods used to reconstruct the water bath image.
Figure 30d shows the temperature reconstruction results. When the k-space hybrid model is fit to the entire image without distinguishing between the brain and water bath, phase artefacts obscure the hot spot in the reconstructed temperature map and (in this case) lead to an overestimation of the temperature rise in the sonicated region across image dynamics (RMSE across dynamics: 0.0121°C). Restricting the temperature reconstruction to within the brain, in combination with keyhole reconstruction of the water bath image (using the baseline image’s k-space to fill in missing k-space lines), produces temperature maps with lower errors in the hot spot but still large errors outside (RMSE across dynamics: 0.0039°C). The proposed k-space brain/POCS bath approach yields a more accurate estimate of the water bath image (not shown), resulting in much lower in-brain temperature artefacts (RMSE across dynamics: 0.0029°C).
Unpredictable water bath motion confounds model-based approaches to accelerated MR temperature mapping, resulting in large temperature artefacts due to aliased water bath signal. We demonstrated that a spatially-segmented reconstruction that applies a model-based reconstruction in the brain and a POCS reconstruction in the water bath can reconstruct temperature maps without undersampling artefacts at a moderate acceleration factor using a single receive coil. Future work will focus on integrating the approach with other accelerated temperature mapping methods  and extending it to non-Cartesian trajectories . The method is compatible with multiple receive coils.
This work was supported by the Focused Ultrasound Foundation and NIBIB T32EB014841.
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O34 Efficient volumetric thermometry for MR-guided FUS brain treatment monitoring, using multiple-echo spirals and mixed update rates
Michael Marx, Pejman Ghanouni, Kim Butts Pauly
Radiology, Stanford University, Stanford, California, USA
MR-guided focused ultrasound (MRgFUS) brain treatments are currently guided by one thermometry sequence: single-slice 2DFT MR thermometry. In this work, we divided treatment monitoring into two tasks, with different thermometry design goals for each, and developed sequences optimized for these goals: “Focal Spot Localization” and “Monitoring”. These sequences achieve greater imaging performance by utilizing multi-echo spiral thermometry, region-specific update rates, and MASTER slice interleaving.
Currently, focal spot targeting confirmation requires several low-power sonications to obtain high-resolution measurements in three dimensions. We developed a Focal Spot Localization sequence that obtains high-resolution measurements in-plane, at improved temperature precision compared to 2DFT, while also providing multiple-slices for through-plane characterization. This would reduce the number of sonications required, improving treatment time. Additionally, even lower-power sonications could be detected, improving patient safety.
During ablative treatment sonications, single-slice monitoring cannot detect through-plane shifts of the ultrasound focus, or unexpected out-of-plane heating. We developed a multi-rate thermometry Monitoring sequence that interleaves different sequences at different update rates to simultaneously achieve fast and precise focal monitoring, 3-dimensional focal spot measurement, and full brain monitoring. Fast and precise multi-slice monitoring of the focus ensures accurate thermal dose estimates for treatment feedback, while (slower) full-brain monitoring ensures patient safety.
All sequences were implemented using RTHawk (HeartVista, Menlo Park, CA) on a GE 3T 750 Signa scanner (GE Healthcare, Milwaukee, WI) equipped with the InSighTec Exablate Neuro (InSighTec, Haifa, Israel). All imaging was performed with the body coil, as is normally used with the Exablate system. Conventional 2DFT was implemented as a gold standard for comparison. The new sequences all use a 36 cm FOV to ensure that the transducer and water bath do not alias. Volunteer imaging was done with informed consent under IRB approval. Multi-frequency reconstruction was performed on all spiral data. Temperature uncertainty was measured as the per-voxel temporal standard deviation of temperature measurements, and averaged within manually segmented ROIs. Performance was also tested inside the transducer, using a gel phantom. Sequence parameters are compiled in Table 2.
The Focal Spot Localization sequence is a 5-slice 3-echo thermometry sequence, with doubled in-plane resolution as compared to conventional 2DFT (1.1x1.1 mm vs 1.09x2.18 mm), and acquisition time of 7 s. The Monitoring sequence interleaves 3 distinct sequences at different rates to monitor 29 total slices. The “Focus” is monitored using 3-slice 3-echo spiral imaging, for high-speed high-precision measurement of focal heating. Two adjacent slices, the “Boundary”, are monitored at half the temporal rate (also using 3-echo spiral) to fully characterize the focal spot. The remaining 24 slices, “Background”, were acquired using 8 blocks of 3-slice MASTER, with spiral readouts. Use of MASTER improves temperature uncertainty, compared to traditional slice interleaving, by increasing echo time. Limiting each MASTER block to 3 slices reduces inherent diffusion and motion-encoding artefacts. The three component sequences were interleaved such that Focus utilized 45% (15% per-slice) of the timeline, Boundary used 15% (7.5% per-slice), and Background the remaining 40% (1.7% per-slice).
Figure 31a compares temperature uncertainty in vivo between 2DFT and Focal Spot Localization while Fig. 31b compares 2DFT with Monitoring for the same volunteer. Large images compare the centre slice, while stacks of images at the right show additional slices monitored by the new sequences. Figure 31c compares 2DFT and Monitoring uncertainty within the transducer. The new sequences obtained better uncertainty than 2DFT, with average values compiled in Table 2. Relative “Efficiency” is also listed in Table 2, which accounts for differences in speed and voxel volume. Each multi-echo spiral sequence is more than 150% as efficient as 2DFT. Background is 69% as efficient (but collects 3 slices per TR, for an effective 120% efficiency).
In this work, we have shown that multi-slice multi-echo spiral thermometry is an effective imaging approach for volumetric treatment monitoring. Improved imaging performance was successfully used to achieve imaging objectives for different aspects of ablative treatments. Focal spots may be localized faster and with reduced heating using the higher-resolution higher-precision Focal Spot Localization sequence. The mixed update rate Monitoring sequence successfully delivers high-speed high-precision monitoring of the targeted focus, while improving safety by simultaneously monitoring the full brain at a lower update rate. These sequences have also been validated within the transducer, to help ensure they will work in the clinical setting.
O35 Towards MR-guided focused ultrasound treatments near metallic hardware
Hans Weber, Valentina Taviani, Kim Butts Pauly, Pejman Ghanouni, Brian Hargreaves
Radiology, Stanford University, Stanford, California, United States
To demonstrate how MRI can be used in FUS treatments near metallic hardware for treatment planning, sonication monitoring and treatment assessment.
Using standard MRgFUS product sequences, we treated a 73-year-old patient with a metastasis in the right femur that was painful despite prior radiation and surgical stabilization with a metallic rod, demonstrating the inability of both echo-planar imaging (EPI) and gradient-recalled echo (GRE) imaging to be used for conventional proton resonance frequency (PRF) shift thermometry.
Based on the experience from this patient treatment, we have proposed an imaging strategy for MRgFUS near metallic hardware using multi-spectral imaging (MSI) techniques (Fig. 32a). Both MAVRIC-SL [Koch et al.; MRM 2011; pp. 65:71] and 2DMSI [Hargreaves et al.; ISMRM 2014, #615] are spin-echo-train-based imaging approaches that enable artefact-reduced imaging near metal. MAVRIC-SL compensates for distortions induced by field inhomogeneities by additional encoding along the slice dimension. Its ability to adjust the image contrast makes it a promising candidate both for treatment planning and assessment. 2DMSI limits the excitation to finite spectral and spatial regions (“frequency bins”) that can be imaged with minimal artefact quickly enough to be used for sonication monitoring.
We preliminarily tested the feasibility of MAVRIC-SL for treatment planning and assessment in a 65-year-old patient without metal hardware and with a metastasis in the right pelvis undergoing MRgFUS treatment for pain palliation. We used a GE 3T MRI system equipped with an InSightec ExAblate2000 FUS system. Proton density (PD) weighted MAVRIC-SL images were acquired prior to the treatment in addition to standard 2D fast-spin-echo (FSE) images. After treatment, T1-weighted MAVRIC-SL images were acquired before and after gadolinium injection in addition to standard 3D fast RF-spoiled GRE images with 2-point-Dixon fat suppression (LAVA-Flex, see Fig. 32c for details).
We tested the feasibility of 2DMSI to monitor a 25 s sonication in an acrylamide egg-white phantom containing the CoCr stem of a knee replacement and a pork loin sample containing a CoCr augment plate from the same hardware, each placed in a container filled with water. We acquired a time series of 10 single-slice 2DMSI images with a temporal resolution of 8 s/frame and the sonication starting after the third image. Each 2DMSI image was composed of 12 frequency bins ranging from −4.5 kHz to +5.4 kHz, and was acquired with TE = 30 ms, (bin) TR = 500 ms, 280 x 280 mm FOV, 5 mm slice thickness and 128 x 128 matrix size (effective number of phase encodings after half Fourier). To monitor the temperature-induced signal change, we subtracted the mean signal of the 3 baseline images from all 10 images voxel-by-voxel.
Figure 32b shows the GRE and EPI image acquired in the patient with metallic hardware. With both techniques, the stabilized femur and its surrounding area are not visible due to distortions and signal dephasing. For comparison, the MAVRIC-SL image depicts the anatomy including the bone marrow surrounding the metallic rod.
Figure 32c presents the acquired images for treatment planning and assessment in the patient without metal. PD-weighted MAVRIC-SL achieves a contrast comparable that of the conventional FSE image and allows for localization of the metastasis in the lower part of the right pelvis. The T1-weighted MAVRIC-SL pre/post contrast difference image reveals the treatment area similar to the LAVA-Flex water difference image.
Figure 33 shows the thermometry results in the phantom and the tissue sample. In both cases, metal-induced field inhomogeneities of up to ± 4 kHz cause strong distortions and signal loss in the GRE image, whereas 2DMSI clearly depicts the area around the metal. In the phantom, the 2DMSI difference images yield a clear signal change at the focal spot next to the metallic stem and a noticeable change in signal at the focal spot next to the metallic plate in the tissue sample, despite a 70% reduction in SNR due to the lower water content. Averaging over the frames during sonication improves the localization of the focal spot.
We have presented initial results for our proposed imaging strategy for MRgFUS in the presence of metallic hardware.
MAVRIC-SL is an established technique for artefact-reduced imaging near metal. Here, we have shown that the image contrast can be adjusted to yield the relevant information for both planning and assessment of MRgFUS treatments. The reduced image resolution compared to the standard FSE protocol (to keep the scan duration at an acceptable length) did not noticeably reduce diagnostic image quality for the treating radiologist.
We have also demonstrated that 2DMSI enables the measurement of temperature-induced signal changes in close proximity to metallic hardware and thus in regions where conventional PRF shift thermometry fails. The bin-selective approach allows for a temporal resolution of 8 s/frame, which is sufficiently high to resolve the temperature evolution in sonications as short as 20 s, which is the typical duration in MRgFUS treatments. In case of less severe metal-induced artefacts, the number of frequency bins could be reduced to increase the temporal resolution. Averaging over time frames could facilitate the detection of the focal spot in lower SNR cases. For the given temporal resolution and the low temperature sensitivity of the T2 relaxation time of aqueous tissues, the measured signal change is expected to be highly dominated by the temperature sensitivity of the PD, whereas a higher temporal resolution is expected to increase the T1 weighting. While the latter provides higher temperature sensitivity and thus lowers the SNR requirements, PD weighting offers the benefit of a tissue-independent temperature mechanism that could facilitate quantitative thermometry.
In conclusion, the proposed imaging strategy has the potential to enable MRgFUS treatments near metallic hardware. The patient population at greater risk for cancers, and hence osseous metastases, overlaps with the elderly demographic more likely to have metallic hardware such as joint replacements. Further, orthopaedic bone stabilization is often used as a treatment for osseous metastases at risk for fracture. Overcoming these technical limitations is therefore important to allow the use of MRgFUS in a larger patient population.
O36 Thermal monitoring of HIFU using thermal memory effect of phase-change nano droplet
Jun Tanaka, Kentaro Kikuchi, Ayumu Ishijima, Takashi Azuma, Kosuke Minamihata, Satoshi Yamaguchi, Teruyuki Nagamune, Ichiro Sakuma, Shu Takagi
The University of Tokyo, Tokyo, Japan
Phase change nano droplets (PCND), whose diameters are 200–400 nm, are droplets of perfluorocarbon (PFC) covered with phospholipid layers. Since they can be vaporized by ultrasound and transformed to microbubbles, they will be utilized as ultrasound contrast agents and ultrasound therapy sensitizers. There are several types of PCND which are often used in research, whose internal compositions are different, such as perfluoropentane (PFP) and perfluorohexane (PFH).Its boiling point can be adjusted by changing the mixture ratio of PFP and PFH.
The lifetimes of generated microbubbles changed from PCND are different from type to type. In this study, the lifetime dependence to ambient temperature at the moment of its vaporization was investigated. If remaining efficiency depends on the ambient temperature, this drug has a potential to be used as indicator of thermal memory effects and temperature monitoring agents.
Experimental setup for ultrasound exposure and high-speed imaging are shown in Fig. 34.In this study, we used the two types of PCND, whose internal compositions were PFP and the mixture (MIX; PFP: PFH = 1:1), respectively. Their main differences are boiling points (PFP: 29°C, MIX: 40°C), and the boiling point of MIX is estimated by thermodynamic calculation.
For the observation of the vaporization of PCND, we used two-layer structure of polyacrylamide gel phantom (layer with PCND and layer without PCND), and set the layer with PCND at the focal point of the transducer. PCND were vaporized by ultrasound (5 MHz centre frequency, 5 cycle bursts, Peak Negative Pressure = 3.5 MPa), which were irradiated with an arbitrary ultrasound beam controller (Verasonics) and a linear array transducer (EUP-L73S, Hitachi Aloka Medical). Time-lapse behaviours of PCND through phase changes were recorded with the high-speed camera (HPV-1A, SHIMADZU), coupled with inverted microscope (NIKON Eclipse Ti-U). The ambient temperature (gel phantom temperature) conditions were controlled with a hot water bath and thermo plate (TOKAI HIT) on the stage of the microscope.
First, we observed the vaporization of the two types of PCND at 37 °C. The high-speed images of vaporization at 37 °C are shown in Fig. 35.
As to MIX (B.P. = 40 °C), generated microbubbles disappeared soon (within 10 μs) after the ultrasound exposure. On the other hand, as to PFP (B.P.=29 °C ), generated microbubbles remained for a while (more than thirty seconds). Because the main difference of these two types of PCND is the boiling point, we assumed that temperature is the key factor, and tried to control the difference between the boiling point and the ambient temperature. Then, we did some vaporization experiment at various temperatures. Some of the high-speed images of vaporization at 26 °C, 48 °C are shown in Fig. 36.
As to PFP (B.P. = 29 °C), generated microbubbles disappeared soon after the sonication at 26 °C, although they remained for some time at 37 °C. As to MIX (B.P.=40 °C), generated microbubbles had long lifetimes at 48 °C, although they had very short lifetimes at 37 °C. It can be considered that the lifetimes of generated microbubbles are greatly affected by not only the internal composition, but also the ambient temperature. Besides, behaviours after vaporization at 26~48 °C are shown in Fig. 37. Whether generated microbubbles will remain or disappear was switched around the boiling point.
We found that the lifetime of microbubble highly depended on the ambient temperature at the moment of vaporization. This effect has a potential to be used as indicator of thermal memory effects and temperature monitoring agents.
Authors thanks to Dr. Kawabata and Mrs. Asami in Hitachi Central Research Laboratory.
O37 Localized blood brain barrier opening of the macaque brain using a high frequency 512-elements FUS transducer and ultrasound contrast agent
Mathieu D. Santin1,2, Laurent Marsac3,4, Guillaume Maimbourg5, Morgane Monfort2, Benoit Larrat4, Chantal François2, Stéphane Lehéricy1,2, Mickael Tanter6, Jean-Franҫois Aubry4
1Centre de NeuroImagerie de Recherche - CENIR, Paris, France; 2Inserm U 1127, CNRS UMR 7225, Sorbonne Universités, UPMC Univ Paris 06 UMR S 1127, Institut du Cerveau et de la Moelle épinière, ICM, Paris, France; 3Supersonic Imagine, Aix-en-Provence, France; 4Institut Langevin, CNRS, Paris, France; 5Institut Langevin, Université Denis Diderot, Paris, France; 6Institut Langevin, INSERM, Paris, France
Blood Brain Barrier (BBB) protects the brain from most of the pathogen circulating within the bloodstream. Thus, the BBB renders a difficult brain penetration for most of the drugs used for brain therapy. Focused ultrasound has the ability to bypass the BBB in small regions considered as targets for various drugs such as chemotherapies. Being able to open the BBB in a restricted focal zone is a necessary step before using this method for therapies such as drug delivery. Here we developed an in-house primate-dedicated stereotactic frame mounted on a multielement focused ultrasound array to perform BBB opening, using combination of FUS and ultrasound contrast agent (UCA). Stereotactic images were loaded in a planning software that allows controlling the localization of the BBB opening. Sonications were real-time monitored using a passive cavitation detector (PCD) to detect signatures of stable or transient inertial cavitation originating from the UCA.
Experiments were conducted on an anesthetized macaque (Macaca fascicularis) each month during one year. Animal was anesthetized with a mixture of Ketamine (3mg/kg) and dexmedetomidine (15μg/kg) and the anaesthesia was maintained by an infusion of Alfaxan (Alfaxalone, 0.2mg/kg/min). The temperature of the animals was maintained at ~37°C using a heated water blanket. Animal physiology was monitored during the whole experiment. A homemade stereotactic frame holding the monkey head was affixed to a 512-element transducer resonating at 1 MHz (SuperSonic Imagine, France). Images were imported in a planning software in which all the positions of the head of the monkey in regard to the transducer were stored, along 6 axis of freedom (2 rotations and one translation for the transducer and one rotation and two translations for the head). Once the coordinate of the target was chosen in the stereotactic frame coordinates, the planning software allowed determining the position of the frame and the transducer. MRI was performed using a 3T Siemens Verio system (Siemens, Germany). Body coil was used for excitation and an 8-channel phased-array coil (Life Services LLC, USA) dedicated to primates was used for reception. T1 longitudinal relaxation time was obtained at baseline using an MP2RAGE sequence prior to BBB opening1. Ultrasound excitation consisted of a 0.6 MPa Peak Negative pressure (as estimated at focus in the brain) sinusoidal tone burst of 10 ms, with a pulse repetition frequency of 1 Hz during 120s. Excitation followed a bolus injection of 1.5 mL of the UCA (Sonovue, Bracco, Switzerland) and lasted for 2 minutes.
The backscattered signal from microbubbles during insonification was recorded using a wideband (−6dB bandwidth: 4.5 - 14.4 MHz) transducer (Imasonic, France) acting as a PCD. The PCD was fixed at the right temporal bone window of the monkey perpendicularly to the FUS beam. This minimizes signal contamination from the main excitation field.
A bolus of 1.5 mL of an MRI contrast agent (Dotarem, Guerbet, France) was injected 5 minutes after the end of ultrasound excitation. A second MP2RAGE dataset was obtained 10 minutes after contrast agent injection to monitor the localization of the BBB opening resulting in a T1 decrease in the region of interest (ROI) induced by the contrast agent.
A conventional clinical scan consisting on a T1-, T2- and T2*-weighted MRI and DTI was performed at the end of the experiments to assess clinical status of the animal.
This study was conducted in a living macaque during one year. Figure 38 shows the apparatus in position with the monkey and inside MRI. After BBB opening, T1 decrease was obtained in the ROI defined by the planning software (example on Fig. 39), indicating that BBB was opened in the targeted ROI. T1 values were 1000 +/− 70 ms before and 662 +/ -31 ms after the HIFU procedure resulting in a ~33% decrease in T1. The size of the area of BBB opening was 3.2 mm in diameter, and 5.6 mm along the axis of the beam.
Figure 40 shows the typical spectra obtained before and during infusion of gas microbubbles. Level of harmonics increased significantly for third (+13.6 dB) and fourth harmonics (+21.6 dB), but no significant increase in broadband noise was detected, suggesting that no transient inertial cavitation occurred during the insonification. After the experiment, the animal recovered and no side effects were observed during the 3 weeks following each BBB opening procedure. At the end of all the BBB opening procedures, T1-, T2- and T2*-weighted scans and DTI did not show any evidence of tissue damage (oedema, haemorrhages or bleeding) induced by the ultrasound procedures (Fig. 41).
The procedure allowed successful repeated transient opening of the BBB in a small ROI in a living primate with no side effects. No signatures of transient inertial cavitation were detected during experiments. This suggests that this method is safe for the animal. This study will be replicated in other animals with the long-term objective of developing a system suitable for human applications.
This work was supported by the Bettencourt Schueller Foundation and the "Agence Nationale de la Recherche" under the program “Future Investments” with the reference ANR-10-EQPX-15.
O38 Enhanced neurorestoration through triple treatment with focused-ultrasound facilitated delivery of the neurotrophic factor neurturin
Maria Eleni Karakatsani1, Gesthimani Samiotaki1, Shutao Wang1, Camilo Acosta1, Eliza R. Feinberg2, Elisa E. Konofagou1,3
1Biomedical Engineering, Columbia University, New York, New York, USA; 2Biological Sciences, Columbia University, New York, New York, USA; 3Radiology, Columbia University, New York, New York, USA
Currently, the existing Central Nervous System (CNS) drug delivery techniques are confined to either targeted but invasive or to non-targeted and non-invasive methods. Focused Ultrasound (FUS) coupled with the systemic administration of microbubbles has been proven to open the Blood Brain Barrier (BBB) locally, transiently and non-invasively, thus facilitating the diffusion of neurotrophic factors. Neurturin, a member of the glial derived neurotrophic factors (GDNF) family has been demonstrated to have restorative effects on the depleted by Parkinson’s disease dopaminergic neurons (DA). Moreover, our group has shown the bioavailability and downstream signalling of Neurturin in wild type mice and the restorative effect in Parkinsonian mice. Despite the promising results, the potential of multiple treatments with Neurturin in reversing the disease phenotype is still to be determined. The aim of the current study was to investigate the neurorestorative effect of triple delivery sessions of the neurotrophic factor Neurturin in a Parkinsonian mouse model.
For this study, twelve wild type mice (12 months old) were infused with sub-acute dosages of MPTP causing apoptotic degeneration in the nigrostriatal pathway. After the stabilization of the lesions and the decontamination period, the entire cohort was sonicated on the left hemisphere (ipsilateral side) targeting twice the Caudate Putamen region (CPu), to cover the entire area, and once the Sabstantia Nigra region (SN). Half of the mice received an IV injection of 0.5mg Neurturin accounting for the treated group, FUS+/NTN+, while the rest constituted the control group, FUS+/NTN-. Magnetic resonance imaging (MRI) was performed after each sonication to verify the accuracy of the BBB opening in terms of targeting. The procedure was repeated once biweekly to a total of three treatments. Following the third treatment, the survival period lasted for 28 days letting the neurotrophic factor to develop its restorative effects. On the 29th day, the mice were sacrificed and coronally sectioned for tissue processing. The brain slices of both the SN and the CPu were stained for tyrosine hydroxylase positive cells (TH+) with a custom protocol. The stained slices were imaged to count the TH+ nerve cell nuclei on the SN while the axons and dendrites were quantified by a custom MATLAB algorithm. For each mouse the contralateral side was compared to the ipsilateral side to eliminate inter-animal variation in the number of nuclei and projections. A quantification algorithm was used to compute the percentage of the relative difference (RD) between the two hemispheres, i.e., RD = (Ipsilateral – Contralateral)*100%. The process was repeated for all slices that cover the entire SN region and averaged across the mice. The error of the technique was measured as the standard deviation from the mean.
There was no significant difference in the number of neurons between the ipsi- and contralateral sides. This result was in accordance with our knowledge of Neurturin restoring impaired neurons and not regenerating them. The RD was found to be significantly higher for the FUS+/NTN+ compared to the FUS+/NTN- group. This significance strengthens with the negative percentage of the FUS+/NTN- group implying a possible sensitivity of the Parkinsonian brain in multiple sonications.
These findings indicate a potential of multiple treatments on the reversal of the Parkinsonian phenotype as is the first time significance is reported. To strengthen this argument a second cohort of 20 mice is currently undergoing the same treatment aiming to apply various other imaging and quantification techniques to investigate the restoration of the functionality of the previously depleted neurons. Nonetheless, the current findings are essential considering the therapeutic effect of multiple treatments with FUS enhanced drug delivery in patients.
O39 Long term effects of single vs repeated low intensity pulsed focused ultrasound treatment with microbubbles
Zsofia I. Kovacs1, Tsang-Wei Tu1, Georgios Z. Papadakis1,2, William C. Reid2, Dima A. Hammoud2, Joseph A. Frank1,3
1Frank Laboratory, Radiology and Imaging Sciences, National Institute of Health, Bethesda, Maryland, United States; 2Center for Infectious Disease Imaging (CIDI), Radiology and Imaging Sciences, National Institute of Health, Bethesda, Maryland, United States; 3National Institute of Biomedical Imaging and Bioengineering, National Institute of Health, Bethesda, Maryland, United States
One potential issue for using MR-guided pulsed Focused Ultrasound (pFUS) to open the blood brain barrier (BBB) is the lack of data on the long term effects. Safety determination in the brain have been limited to the MR characterization after repeated BBB opening that can be achieved without haemorrhage, oedema and behavioural changes in non-human primates [1,2]. We use multimodal imaging technics to characterize long term effects of pFUS + MB in the rat brain to evaluate the effects of repeated BBB opening by pFUS and microbubbles (MB) on morphology to the rat striatum and hippocampus as monitored by magnetic resonance imaging (MRI), positron emission tomography (PET) and histology over 12 weeks.
Female rats were divided into two groups and received either pFUS + MB (OptisonTM, GE Healthcare, Little Chalfont, UK) once or six times targeting the striatum and the contralateral hippocampus. 200 μl of MB were administered intravenously over 1 minute starting 30 sec before pFUS. Rats received 3 daily doses of 300 mg/kg 5-Bromo-2′-deoxy-uridine (BrdU, Sigma Aldrich, St. Louis, MO) intraperitoneally before sonication to label proliferating cells in vivo.
0.3 MPa acoustic pressure was applied in 10 ms burst length and 1% duty cycle (9 focal points, 120 sec/9 focal points - striatum, 120 sec/4 focal points - hippocampus) using a single-element spherical FUS transducer (centre frequency 589.636 kHz; focal number 0.8; aperture 7.5 cm; FUS Instruments, Toronto, Ontario, Canada). T2, T2* and Gd-enhanced T1-weighted images were obtained by 3.0 T MRI (Philips, Amsterdam, Netherlands), T2, T2*, diffusion tensor imaging (DTI) and chemical exchange saturation transfer (CEST) imaging was performed by 9.4 T MRI (Bruker, Billerica, MA). Parameters for DTI: 3D spin echo EPI; TR/TE 700 ms/37 ms; b-value 800 s/mm2 with 17 encoding directions; voxel size 200 μm, isotropic. Diffusion weighted images were corrected for B0 susceptibility induced EPI distortion, eddy current distortions, and motion distortion with b-matrix reorientation using Tortoise. Parameter for glucoCEST: 2D fast spin echo with (MT) and without (M0) magnetization transfer (MT) pulses (TR/TE 3.5 s/11.5 ms; in plane resolution: 200 μm, thickness: 0.8 mm; MT pulse: 3 μT, 1 s). The MT offset frequencies (Δω) were set from −2 kHz to +2 kHz with 100 Hz stepping to detect the proton metabolites of glucose (1.2 ppm, 2.1 ppm, 2.9 ppm). Fractional anisotropy (DTI-FA) and the asymmetry of magnetization transfer ratio (MTRasym) were derived for mapping structural injury and glucose metabolism.
Quantitative of glucose uptake was performed with FDG-PET (Siemens, Munich, Germany). Each rat received under anaesthesia (O2 3–4 L/min & Isoflurane at 3–3.5 4%) 0.7 – 1.1 mCi of 18F-FDG via tail vein injection and was allowed to regain consciousness for an uptake period of 30 minutes in total. They were anesthetized again and a PET/CT study was acquired using Siemens Inveon Multimodality scanner (Siemens Medical Solutions USA, Inc.). CT scan was performed for localization and attenuation correction purposes. PET images were reconstructed using OSEM3D/MAP algorithm, with Ramp projection filter, scattered corrected, 2 OSEM3D iterations, 18 MAP iterations, 128 × 128 image size, and approximately 0.5mm resolution at the centre of the field of view (FOV).
Animals were euthanized 6 or 12 weeks after the first pFUS treatment. Histological evaluation of brain and tracking of BrdU tagged cells was performed at different time points. Values were compared to baseline.
Preliminary results showed contrast enhancement on T1-weighted MRI in rats receiving a single sonication, indicating BBB disruption in the striatum and the hippocampus. Gd-extravasation or T2 and T2* abnormalities were not seen in the brain 1 day post pFUS + MB at 9.4 T MRI. Hypointense regions appeared on T2* MRI 2 weeks after pFUS + MB (Figs. 43 and 44) consistently with microhemorrhage within the parenchyma that decreased in volumes by week 3. White matter fiber structure- and gray matter-abnormalities on DTI MRI were detected in regions with T2* abnormalities (Fig. 44) suggestive of increased astrogliosis (Fig. 44a) and transient axonal damage (Fig. 44b). GlucoCEST showed loss of contrast as early as 1 day post pFUS and these changes persisted up to week 3 (Fig. 44a).
Qualitative analysis of MRI and GlucoCEST as well as 18F-FDG uptake with PET showed no difference between the sonicated region and the contralateral hemisphere 6 weeks post sonication.
We have observed a complex graded molecular and cellular sterile inflammatory response in the brain up to 24 hrs after pFUS + MB. However, little is known about the long term effects in rats using advanced imaging techniques. The DTI data showed that pFUS caused a low degree of structural injury at the location of sonication. However, the decrease in glucose concentration revealed by glucoCEST indicated that the pFUS could cause hypo-metabolism in the brain even after 3 weeks post sonication. These preliminary