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Background Previous work has demonstrated that drugs can be delivered across the blood-brain barrier by exposing circulating microbubbles to a sequence of long ultrasound pulses. Although this sequence has successfully delivered drugs to the brain, concerns remain regarding potentially harmful effects from disrupting the brain vasculature. Purpose To determine whether a low-energy, rapid, short-pulse ultrasound sequence can efficiently and safely deliver drugs to the murine brain. Materials and Methods Twenty-eight female wild-type mice underwent focused ultrasound treatment after injections of microbubbles and a labeled model drug, while three control mice were not treated (May-November 2017). The left hippocampus of 14 mice was exposed to low-energy short pulses (1 MHz; five cycles; peak negative pressure, 0.35 MPa) of ultrasound emitted at a rapid rate (1.25 kHz) in bursts (0.5 Hz), and another 14 mice were exposed to standard long pulses (10 msec, 0.5 Hz) containing 150 times more acoustic energy. Mice were humanely killed at 0 (n = 5), 10 (n = 3), or 20 minutes (n = 3) after ultrasound treatment. Hematoxylin-eosin (H-E) staining was performed on three mice. The delivered drug dose and distribution were quantified with the normalized optical density and coefficient of variation. Safety was assessed by H-E staining, the amount of albumin released, and the duration of permeability change in the blood-brain barrier. Statistical analysis was performed by using the Student t test. Results The rapid short-pulse sequence delivered drugs uniformly throughout the parenchyma. The acoustic energy emitted from the microbubbles also predicted the delivered dose (r = 0.97). Disruption in the blood-brain barrier lasted less than 10 minutes and 3.4-fold less albumin was released into the brain than with long pulses. No vascular or tissue damage from rapid short-pulse exposure was observable using H-E staining. Conclusion The rapid short-pulse ultrasound sequence is a minimally disruptive and efficient drug delivery method that could improve the treatment, diagnosis, and study of neurologic diseases. © RSNA, 2019 Online supplemental material is available for this article. See also the editorial by Klibanov and McDannold in this issue.
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Barreira Hematoencefálica/metabolismo , Sistemas de Liberação de Medicamentos/métodos , Sonicação/métodos , Animais , Sistemas de Liberação de Medicamentos/instrumentação , Feminino , Corantes Fluorescentes/farmacocinética , Hipocampo/química , Camundongos , Camundongos Endogâmicos C57BL , Microbolhas , Sonicação/instrumentação , Distribuição TecidualRESUMO
Ultrasound-driven microbubbles have been used in therapeutic applications to deliver drugs across capillaries and into cells or to dissolve blood clots. Yet the performance and safety of these applications have been difficult to control. Microbubbles exposed to ultrasound not only volumetrically oscillate, but also move due to acoustic radiation, or Bjerknes, forces. The purpose of this work was to understand the extent to which microbubbles moved and clustered due to secondary Bjerknes forces. A microbubble population was exposed to a 1-MHz ultrasound pulse with a peak-rarefactional pressure of 50-100 kPa and a pulse length of 20 ms. Microbubbles exposed to low-pressure therapeutic ultrasound were observed to cluster at clustering rates of 0.01-0.02 microbubbles per duration (in ms) per initial average inter-bubble distance (in µm), resulting in 1 to 3 clustered microbubbles per initial average inter-bubble distance (in µm). Higher pressures caused faster clustering rates and a larger number of clustered microbubbles. Experimental data revealed clustering time scales, cluster localizations, and cluster sizes that were in reasonable agreement with simulations using a time-averaged model at low pressures. This study demonstrates that clustering of microbubbles occurs within a few milliseconds and is likely to influence the distribution of stimuli produced in therapeutic applications.
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Despite the promise of microbubble-mediated focused ultrasound therapies, in vivo findings have revealed over-treated and under-treated regions distributed throughout the focal volume. This poor distribution cannot be improved by conventional pulse shapes and sequences, due to their limited ability to control acoustic cavitation dynamics within the ultrasonic focus. This paper describes the design of a rapid short-pulse (RaSP) sequence which is comprised of short pulses separated by µs off-time intervals. Improved acoustic cavitation distribution was based on the hypothesis that microbubbles can freely move during the pulse off-times. Flowing SonoVue® microbubbles (flow velocity: 10 mm/s) were sonicated with a 0.5 MHz focused ultrasound transducer using RaSP sequences (peak-rarefactional pressures: 146-900 kPa, pulse repetition frequency: 1.25 kHz, and pulse lengths: 5-50 cycles). The distribution of cavitation activity was evaluated using passive acoustic mapping. RaSP sequences generated uniform distributions within the focus in contrast to long pulses (50 000 cycles) that produced non-uniform distributions. Fast microbubble destruction occurred for long pulses, whereas microbubble activity was sustained for longer durations for shorter pulses. High-speed microscopy revealed increased mobility in the direction of flow during RaSP sonication. In conclusion, RaSP sequences produced spatiotemporally uniform cavitation distributions and could result in efficient therapies by spreading cavitation throughout the treatment area.
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Background: Focused ultrasound (FUS) in combination with microbubbles has recently shown great promise in facilitating blood-brain barrier (BBB) opening for drug delivery and immunotherapy in Alzheimer's disease (AD). However, it is currently limited to systems integrated within the MRI suites or requiring post-surgical implants, thus restricting its widespread clinical adoption. In this pilot study, we investigate the clinical safety and feasibility of a portable, non-invasive neuronavigation-guided FUS (NgFUS) system with integrated real-time 2-D microbubble cavitation mapping. Methods: A phase 1 clinical study with mild to moderate AD patients (N=6) underwent a single session of microbubble-mediated NgFUS to induce transient BBB opening (BBBO). Microbubble activity under FUS was monitored with real-time 2-D cavitation maps and dosing to ensure the efficacy and safety of the NgFUS treatment. Post-operative MRI was used for BBB opening and closure confirmation as well as safety assessment. Changes in AD biomarker levels in both blood serum and extracellular vesicles (EVs) were evaluated, while changes in amyloid-beta (Aß) load in the brain were assessed through 18F-Florbetapir PET. Results: BBBO was achieved in 5 out of 6 subjects with an average volume of 983±626 mm3 following FUS at the right frontal lobe both in white and gray matter regions. The outpatient treatment was completed within 34.8±10.7 min. Cavitation dose significantly correlated with the BBBO volume (R 2>0.9, N=4), demonstrating the portable NgFUS system's capability of predicting opening volumes. The cavitation maps co-localized closely with the BBBO location, representing the first report of real-time transcranial 2-D cavitation mapping in the human brain. Larger opening volumes correlated with increased levels of AD biomarkers, including Aß42 (R 2=0.74), Tau (R 2=0.95), and P-Tau181 (R 2=0.86), assayed in serum-derived EVs sampled 3 days after FUS (N=5). From PET scans, subjects showed a lower Aß load increase in the treated frontal lobe region compared to the contralateral region. Reduction in asymmetry standardized uptake value ratios (SUVR) correlated with the cavitation dose (R 2>0.9, N=3). Clinical changes in the mini-mental state examination over 6 months were within the expected range of cognitive decline with no additional changes observed as a result of FUS. Conclusion: We showed the safety and feasibility of this cost-effective and time-efficient portable NgFUS treatment for BBBO in AD patients with the first demonstration of real-time 2-D cavitation mapping. The cavitation dose correlated with BBBO volume, a slowed increase in pathology, and serum detection of AD proteins. Our study highlights the potential for accessible FUS treatment in AD, with or without drug delivery.
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Background : Focused ultrasound (FUS) in combination with microbubbles has recently shown great promise in facilitating blood-brain barrier (BBB) opening for drug delivery and immunotherapy in Alzheimer's disease (AD). However, it is currently limited to systems integrated within the MRI suites or requiring post-surgical implants, thus restricting its widespread clinical adoption. In this pilot study, we investigate the clinical safety and feasibility of a portable, non-invasive neuronavigation-guided FUS (NgFUS) system with integrated real-time 2-D microbubble cavitation mapping. Methods : A phase 1 clinical study with mild to moderate AD patients (N = 6) underwent a single session of microbubble-mediated NgFUS to induce transient BBB opening (BBBO). Microbubble activity under FUS was monitored with real-time 2-D cavitation maps and dosing to ensure the efficacy and safety of the NgFUS treatment. Post-operative MRI was used for BBB opening and closure confirmation as well as safety assessment. Changes in AD biomarker levels in both blood serum and extracellular vesicles (EVs) were evaluated, while changes in amyloid-beta (Aß) load in the brain were assessed through 18F-florbetapir PET. Results : BBBO was achieved in 5 out of 6 subjects with an average volume of 983 ± 626 mm3 following FUS at the right frontal lobe both in white and gray matter regions. The outpatient treatment was completed within 34.8 ± 10.7 min. Cavitation dose significantly correlated with the BBBO volume (R 2 > 0.9, N = 4), demonstrating the portable NgFUS system's capability of predicting opening volumes. The cavitation maps co-localized closely with the BBBO location, representing the first report of real-time transcranial 2-D cavitation mapping in the human brain. Larger opening volumes correlated with increased levels of AD biomarkers, including Aß42 (R 2 = 0.74), Tau (R 2 = 0.95), and P-Tau181 (R 2 = 0.86), assayed in serum-derived EVs sampled 3 days after FUS (N = 5). From PET scans, subjects showed a lower Aß load increase in the treated frontal lobe region compared to the contralateral region. Reduction in asymmetry standardized uptake value ratios (SUVR) correlated with the cavitation dose (R 2 > 0.9, N = 3). Clinical changes in the mini-mental state examination over 6 months were within the expected range of cognitive decline with no additional changes observed as a result of FUS. Conclusion : We showed the safety and feasibility of this cost-effective and time-efficient portable NgFUS treatment for BBBO in AD patients with the first demonstration of real-time 2-D cavitation mapping. The cavitation dose correlated with BBBO volume, a slowed increase in pathology, and serum detection of AD proteins. Our study highlights the potential for accessible FUS treatment in AD, with or without drug delivery.
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Doença de Alzheimer , Barreira Hematoencefálica , Imageamento por Ressonância Magnética , Microbolhas , Humanos , Barreira Hematoencefálica/metabolismo , Barreira Hematoencefálica/diagnóstico por imagem , Doença de Alzheimer/terapia , Doença de Alzheimer/diagnóstico por imagem , Masculino , Idoso , Feminino , Projetos Piloto , Imageamento por Ressonância Magnética/métodos , Peptídeos beta-Amiloides/metabolismo , Pessoa de Meia-Idade , Biomarcadores/metabolismo , Sistemas de Liberação de Medicamentos/métodos , Idoso de 80 Anos ou maisRESUMO
OBJECTIVE: Passive acoustic mapping (PAM) provides the spatial information of acoustic energy emitted from microbubbles during focused ultrasound (FUS), which can be used for safety and efficacy monitoring of blood-brain barrier (BBB) opening. In our previous work with a neuronavigation-guided FUS system, only part of the cavitation signal could be monitored in real time due to the computational burden although full-burst analysis is required to detect transient and stochastic cavitation activity. In addition, the spatial resolution of PAM can be limited for a small-aperture receiving array transducer. For full-burst real-time PAM with enhanced resolution, we developed a parallel processing scheme for coherence-factor-based PAM (CF-PAM) and implemented it onto the neuronavigation-guided FUS system using a co-axial phased-array imaging transducer. METHODS: Simulation and in-vitro human skull studies were conducted for the performance evaluation of the proposed method in terms of spatial resolution and processing speed. We also carried out real-time cavitation mapping during BBB opening in non-human primates (NHPs). RESULTS: CF-PAM with the proposed processing scheme provided better resolution than that of traditional time-exposure-acoustics PAM with a higher processing speed than that of eigenspace-based robust Capon beamformer, which facilitated the full-burst PAM with the integration time of 10 ms at a rate of 2 Hz. In vivo feasibility of PAM with the co-axial imaging transducer was also demonstrated in two NHPs, showing the advantages of using real-time B-mode and full-burst PAM for accurate targeting and safe treatment monitoring. SIGNIFICANCE: This full-burst PAM with enhanced resolution will facilitate the clinical translation of online cavitation monitoring for safe and efficient BBB opening.
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Barreira Hematoencefálica , Terapia por Ultrassom , Animais , Barreira Hematoencefálica/diagnóstico por imagem , Neuronavegação , Acústica , Ultrassonografia/métodos , Terapia por Ultrassom/métodos , MicrobolhasRESUMO
Optogenetics employs engineered viruses to genetically modify cells to express specific light-sensitive ion channels. The standard method for gene delivery in the brain involves invasive craniotomies that expose the brain and direct injections of viruses that invariably damage neural tissue along the syringe tract. A recently proposed alternative in which non-invasive optogenetics is performed with focused ultrasound (FUS)-mediated blood-brain barrier (BBB) openings has been found to non-invasively facilitate gene delivery for optogenetics in mice. Although gene delivery can be performed non-invasively, validating successful viral transduction and expression of encoded ion channels in target tissue typically involves similar invasive techniques, such as craniotomies in longitudinal studies and/or postmortem histology. Functional ultrasound imaging (fUSi) is an emerging neuroimaging technique that can be used to transcranially detect changes in cerebral blood volume following introduction of a stimulus. In this study, we implemented a fully non-invasive combined FUS-fUSi technique for performing optogenetics in mice. FUS successfully delivered viruses encoding the red-shifted channelrhodopsin variant ChrimsonR in all treated subjects. fUSi successfully identified stimulus-evoked cerebral blood volume changes preferentially in brain regions expressing the light-sensitive ion channels. Improvements in cell-specific targeting of viral vectors and transcranial ultrasound imaging will make the combined technique a useful tool for neuroscience research in small animals.
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Barreira Hematoencefálica , Encéfalo , Técnicas de Transferência de Genes , Canais Iônicos , Optogenética , Ultrassonografia , Animais , Camundongos , Barreira Hematoencefálica/diagnóstico por imagem , Barreira Hematoencefálica/metabolismo , Encéfalo/diagnóstico por imagem , Encéfalo/fisiologia , Canais Iônicos/metabolismo , Optogenética/métodosRESUMO
BACKGROUND: Focused ultrasound (FUS) is a non-invasive neuromodulation technology that is being investigated for potential treatment of neurological and psychiatric disorders. FUS combined with microbubbles can temporarily open the intact blood-brain barrier (BBB) of animals and humans, and facilitate drug delivery. FUS exposure, either with or without microbubbles, has been demonstrated to alter the behavior of non-human primates (NHP), and previous studies have demonstrated the transient and long-term effects of FUS neuromodulation on functional connectivity using resting state functional MRI. The behavioral effects of FUS vary depending on whether or not it is applied in conjunction with microbubbles to open the BBB, but it is unknown whether opening the BBB affects functional connectivity differently than FUS alone. OBJECTIVE: To compare the effects of applying FUS alone (FUS neuromodulation) and FUS with microbubbles (FUS-BBB opening) on changes of resting state functional connectivity in NHP. METHODS: We applied 2 min FUS exposure without (neuromodulation) and with microbubbles (BBB opening) in the dorsal striatum of lightly anesthetized non-human primates, and acquired resting state functional MRI 40 min respectively after FUS exposure. The functional connectivity (FC) in the cortex and major brain networks between the two approaches were measured and compared. RESULTS: When applying FUS exposure to the caudate nucleus of NHP, we found that both FUS neuromodulation can activate FC between caudate and insular cortex, while inhibiting the FC between caudate and motor cortex. FUS-BBB opening can activate FC between the caudate and medial prefrontal cortex, and within the frontotemporal network (FTN). We also found both FUS and FUS-BBB opening can significantly activate FC within the default mode network (DMN). CONCLUSION: The results suggest applying FUS to a deep brain structure can alter functional connectivity in the DMN and FTN, and that FUS neuromodulation and FUS-mediated BBB opening can have different effects on patterns of functional connectivity.
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Barreira Hematoencefálica , Encéfalo , Animais , Humanos , Encéfalo/diagnóstico por imagem , Encéfalo/fisiologia , Primatas , Córtex Cerebral/diagnóstico por imagem , Microbolhas , Imageamento por Ressonância MagnéticaRESUMO
Focused ultrasound (FUS) can be used to open the blood-brain barrier (BBB), and MRI with contrast agents can detect that opening. However, repeated use of gadolinium-based contrast agents (GBCAs) presents safety concerns to patients. This study is the first to propose the idea of modeling a volume transfer constant (Ktrans) through deep learning to reduce the dosage of contrast agents. The goal of the study is not only to reconstruct artificial intelligence (AI) derived Ktrans images but to also enhance the intensity with low dosage contrast agent T1 weighted MRI scans. We successfully validated this idea through a previous state-of-the-art temporal network algorithm, which focused on extracting time domain features at the voxel level. Then we used a Spatiotemporal Network (ST-Net), composed of a spatiotemporal convolutional neural network (CNN)-based deep learning architecture with the addition of a three-dimensional CNN encoder, to improve the model performance. We tested the ST-Net model on ten datasets of FUS-induced BBB-openings aquired from different sides of the mouse brain. ST-Net successfully detected and enhanced BBB-opening signals without sacrificing spatial domain information. ST-Net was shown to be a promising method of reducing the need of contrast agents for modeling BBB-opening K-trans maps from time-series Dynamic Contrast-Enhanced Magnetic Resonance Imaging (DCE-MRI) scans.
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Rationale: Bilateral sonication with focused ultrasound (FUS) in conjunction with microbubbles has been shown to separately reduce amyloid plaques and hyperphosphorylated tau protein in the hippocampal formation and the entorhinal cortex in different mouse models of Alzheimer's disease (AD) without any therapeutic agents. However, the two pathologies are expressed concurrently in human disease. Therefore, the objective of this study is to investigate the effects of repeated bilateral sonications in the presence of both pathologies. Methods: Herein, we investigate its functional and morphological outcomes on brains bearing both pathologies simultaneously. Eleven transgenic mice of the 3xTg-AD line (14 months old) expressing human amyloid beta and human tau and eleven age-matched wild-type littermates received four weekly bilateral sonications covering the hippocampus followed by working memory testing. Afterwards, immunohistochemistry and immunoassays (western blot and ELISA) were employed to assess any changes in amyloid beta and human tau. Furthermore, we present preliminary data from our clinical trial using a neuronavigation-guided FUS system for sonications in AD patients (NCT04118764). Results: Interestingly, both wild-type and transgenic animals that received FUS experienced improved working memory and spent significantly more time in the escape platform-quadrant, with wild-type animals spending 43.2% (sham: 37.7%) and transgenic animals spending 35.3% (sham: 31.0%) of the trial in the target quadrant. Furthermore, this behavioral amelioration in the transgenic animals correlated with a 58.3% decrease in the neuronal length affected by tau and a 27.2% reduction in total tau levels. Amyloid plaque population, volume and overall load were also reduced overall. Consistently, preliminary data from a clinical trial involving AD patients showed a 1.8% decrease of amyloid PET signal 3-weeks after treatment in the treated hemisphere compared to baseline. Conclusion: For the first time, it is shown that bilateral FUS-induced BBB opening significantly and simultaneously ameliorates both coexistent pathologies, which translated to improvements in spatial memory of transgenic animals with complex AD, the human mimicking phenotype. The level of cognitive improvement was significantly correlated with the volume of BBB opening. Non-transgenic animals were also shown to exhibit similar memory amelioration for the first time, indicating that BBB opening results into benefits in the neuronal function regardless of the existence of AD pathology. A potential mechanism of action for the reduction of the both pathologies investigated was the cholesterol metabolism, specifically the LRP1b receptor, which exhibited increased expression levels in transgenic mice following FUS-induced BBB opening. Initial clinical evidence supported that the beta amyloid reduction shown in rodents could be translatable to humans with significant amyloid reduction shown in the treated hemisphere.
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Doença de Alzheimer , Humanos , Camundongos , Animais , Recém-Nascido , Lactente , Doença de Alzheimer/metabolismo , Peptídeos beta-Amiloides/metabolismo , Memória Espacial , Encéfalo/metabolismo , Camundongos Transgênicos , Modelos Animais de DoençasRESUMO
Phase-change nanodroplets have attracted increasing interest in recent years as ultrasound theranostic nanoparticles. They are smaller compared to microbubbles and they may distribute better in tissues (e.g. in tumours). They are composed of a stabilising shell and a perfluorocarbon core. Nanodroplets can vaporise into echogenic microbubbles forming cavitation nuclei when exposed to ultrasound. Their perfluorocarbon core phase-change is responsible for the acoustic droplet vaporisation. However, methods to quantify the perfluorocarbon core in nanodroplets are lacking. This is an important feature that can help explain nanodroplet phase change characteristics. In this study, we fabricated nanodroplets using lipids shell and perfluorocarbons. To assess the amount of perfluorocarbon in the core we used two methods, 19F NMR and FTIR. To assess the cavitation after vaporisation we used an ultrasound transducer (1.1 MHz) and a high-speed camera. The 19F NMR based method showed that the fluorine signal correlated accurately with the perfluorocarbon concentration. Using this correlation, we were able to quantify the perfluorocarbon core of nanodroplets. This method was used to assess the content of the perfluorocarbon of the nanodroplets in solutions over time. It was found that perfluoropentane nanodroplets lost their content faster and at higher ratio compared to perfluorohexane nanodroplets. The high-speed imaging indicates that the nanodroplets generate cavitation comparable to that from commercial contrast agent microbubbles. Nanodroplet characterisation should include perfluorocarbon concentration assessment as critical information for their development.
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Fluorocarbonos , Nanopartículas , Ultrassonografia , Nanopartículas/química , Volatilização , Meios de Contraste/química , Fluorocarbonos/química , MicrobolhasRESUMO
Optogenetics has revolutionized the capability of controlling genetically modified neurons in vitro and in vivo and has become an indispensable neuroscience tool. Using light as a probe for selective neuronal activation or inhibition and as a means to read out neural activity has dramatically enhanced our understanding of complex neural circuits. However, a common limitation of optogenetic studies to date is their invasiveness and spatiotemporal range. Direct viral injections into the brain tissue along with implantation of optical fibers and recording electrodes can disrupt the neuronal circuitry and cause significant damage. Conventional approaches are spatially limited around the site of the direct injection and insufficient in examining large networks throughout the brain. Lastly, optogenetics is currently not easily scalable to large animals or humans. Here, we demonstrate that optogenetic excitation can be achieved entirely non-invasively through the intact skull in mice. Using a needle-free combination of focused ultrasound-mediated viral delivery and extracorporeal illumination with red light, we achieved selective neuronal activation at depths up to 4 mm in the murine brain, confirmed through cFos expression and electrophysiology measurements within the treated areas. Ultrasound treatment significantly reduced freezing time during recall in fear conditioning experiments, but remote light exposure had a moderate effect on the freezing behavior of mice treated with viral vectors. The proposed method has the potential to open new avenues of studying, but also stimulating, neuronal networks, in an effort to elucidate normal or dysfunctional brain activity and treat neurological diseases. Finally, the same non-invasive methodology could be combined with gene therapy and applied to other organs, such as the eye and the heart.
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Neurônios , Optogenética , Animais , Encéfalo/fisiologia , Terapia Genética , Humanos , Camundongos , Neurônios/fisiologia , Optogenética/métodos , Estimulação LuminosaRESUMO
OBJECTIVE: Focused ultrasound (FUS) has emerged as a non-invasive technique to locally and reversibly disrupt the blood-brain barrier (BBB). Here, we investigate the use of diffusion tensor imaging (DTI) as a means of detecting FUS-induced BBB opening at the absence of an MRI contrast agent. A non-human primate (NHP) was repeatedly treated with FUS and preformed circulating microbubbles to transiently disrupt the BBB (n = 4). T1- and diffusion-weighted MRI scans were acquired after the ultrasound treatment, with and without gadolinium-based contrast agent, respectively. Both scans were registered with a high-resolution T1-weighted scan of the NHP to investigate signal correlations. DTI detected an increase in fractional anisotropy from 0.21 ± 0.02 to 0.38 ± 0.03 (82.6 ± 5.2% change) within the targeted area one hour after BBB opening. Enhanced DTI contrast overlapped by 77.22 ± 9.2% with hyper-intense areas of gadolinium-enhanced T1-weighted scans, indicating diffusion anisotropy enhancement only within the BBB opening volume. Diffusion was highly anisotropic and unidirectional within the treated brain region, as indicated by the direction of the principal diffusion eigenvectors. Polar and azimuthal angle ranges decreased by 35.6% and 82.4%, respectively, following BBB opening. Evaluation of the detection methodology on a second NHP (n = 1) confirmed the across-animal feasibility of the technique. In conclusion, DTI may be used as a contrast-free MR imaging modality in lieu of contrast-enhanced T1 mapping for detecting BBB opening during focused-ultrasound treatment or evaluating BBB integrity in brain-related pathologies.
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Barreira Hematoencefálica , Terapia por Ultrassom , Animais , Barreira Hematoencefálica/diagnóstico por imagem , Meios de Contraste , Imagem de Tensor de Difusão , Sistemas de Liberação de Medicamentos , Imageamento por Ressonância Magnética , MicrobolhasRESUMO
Passive acoustic mapping enables the spatiotemporal monitoring of cavitation with circulating microbubbles during focused ultrasound (FUS)-mediated blood-brain barrier opening. However, the computational load for processing large data sets of cavitation maps or more complex algorithms limit the visualization in real-time for treatment monitoring and adjustment. In this study, we implemented a graphical processing unit (GPU)-accelerated sparse matrix-based beamforming and time exposure acoustics in a neuronavigation-guided ultrasound system for real-time spatiotemporal monitoring of cavitation. The system performance was tested in silico through benchmarking, in vitro using nonhuman primate (NHP) and human skull specimens, and demonstrated in vivo in NHPs. We demonstrated the stability of the cavitation map for integration times longer than 62.5 [Formula: see text]. A compromise between real-time displaying and cavitation map quality obtained from beamformed RF data sets with a size of 2000 ×128 ×30 (axial [Formula: see text]) was achieved for an integration time of [Formula: see text], which required a computational time of 0.27 s (frame rate of 3.7 Hz) and could be displayed in real-time between pulses at PRF = 2 Hz. Our benchmarking tests show that the GPU sparse-matrix algorithm processed the RF data set at a computational rate of [Formula: see text]/pixel/sample, which enables adjusting the frame rate and the integration time as needed. The neuronavigation system with real-time implementation of cavitation mapping facilitated the localization of the cavitation activity and helped to identify distortions due to FUS phase aberration. The in vivo test of the method demonstrated the feasibility of GPU-accelerated sparse matrix computing in a close to a clinical condition, where focus distortions exemplify problems during treatment. These experimental conditions show the need for spatiotemporal monitoring of cavitation with real-time capability that enables the operator to correct or halt the sonication in case substantial aberrations are observed.
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Acústica , Microbolhas , Algoritmos , Animais , Barreira Hematoencefálica/diagnóstico por imagem , UltrassonografiaRESUMO
An emerging approach with potential in improving the treatment of neurodegenerative diseases and brain tumors is the use of focused ultrasound (FUS) to bypass the blood-brain barrier (BBB) in a non-invasive and localized manner. A large body of pre-clinical work has paved the way for the gradual clinical implementation of FUS-induced BBB opening. Even though the safety profile of FUS treatments in rodents has been extensively studied, the histological and behavioral effects of clinically relevant BBB opening in large animals are relatively understudied. Here, we examine the histological and behavioral safety profile following localized BBB opening in non-human primates (NHPs), using a neuronavigation-guided clinical system prototype. We show that FUS treatment triggers a short-lived immune response within the targeted region without exacerbating the touch accuracy or reaction time in visual-motor cognitive tasks. Our experiments were designed using a multiple-case-study approach, in order to maximize the acquired data and support translation of the FUS system into human studies. Four NHPs underwent a single session of FUS-mediated BBB opening in the prefrontal cortex. Two NHPs were treated bilaterally at different pressures, sacrificed on day 2 and 18 post-FUS, respectively, and their brains were histologically processed. In separate experiments, two NHPs that were earlier trained in a behavioral task were exposed to FUS unilaterally, and their performance was tracked for at least 3 weeks after BBB opening. An increased microglia density around blood vessels was detected on day 2, but was resolved by day 18. We also detected signs of enhanced immature neuron presence within areas that underwent BBB opening, compared to regions with an intact BBB, confirming previous rodent studies. Logistic regression analysis showed that the NHP cognitive performance did not deteriorate following BBB opening. These preliminary results demonstrate that neuronavigation-guided FUS with a single-element transducer is a non-invasive method capable of reversibly opening the BBB, without substantial histological or behavioral impact in an animal model closely resembling humans. Future work should confirm the observations of this multiple-case-study work across animals, species and tasks.
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Barreira Hematoencefálica/metabolismo , Barreira Hematoencefálica/efeitos da radiação , Neuronavegação/métodos , Ondas Ultrassônicas , Animais , Comportamento Animal , Transporte Biológico/efeitos da radiação , Biomarcadores , Barreira Hematoencefálica/diagnóstico por imagem , Cognição , Imageamento por Ressonância Magnética , Microbolhas , Modelos Animais , Primatas , Característica Quantitativa HerdávelRESUMO
PURPOSE: Glioblastoma (GBM) is a devastating disease. With the current treatment of surgery followed by chemoradiation, outcomes remain poor, with median survival of only 15 months and a 5-year survival rate of 6.8%. A challenge in treating GBM is the heterogeneous integrity of the blood-brain barrier (BBB), which limits the bioavailability of systemic therapies to the brain. There is a growing interest in enhancing drug delivery by opening the BBB with the use of focused ultrasound (FUS). We hypothesize that an FUS-mediated BBB opening can enhance the delivery of etoposide for a therapeutic benefit in GBM. METHODS AND MATERIALS: A murine glioma cell line (Pdgf+, Pten-/-, P53-/-) was orthotopically injected into B6(Cg)-Tyrc-2J/J mice to establish the syngeneic GBM model for this study. Animals were treated with FUS and microbubbles to open the BBB to enhance the delivery of systemic etoposide. Magnetic resonance (MR) imaging was used to evaluate the BBB opening and tumor progression. Liquid chromatography tandem mass spectrometry was used to measure etoposide concentrations in the intracranial tumors. RESULTS: The murine glioma cell line is sensitive to etoposide in vitro. MR imaging and passive cavitation detection demonstrate the safe and successful BBB opening with FUS. The combined treatment of an FUS-mediated BBB opening and etoposide decreased tumor growth by 45% and prolonged median overall survival by 6 days: an approximately 30% increase. The FUS-mediated BBB opening increased the brain tumor-to-serum ratio of etoposide by 3.5-fold and increased the etoposide concentration in brain tumor tissue by 8-fold compared with treatment without ultrasound. CONCLUSIONS: The current study demonstrates that BBB opening with FUS increases intratumoral delivery of etoposide in the brain, resulting in local control and overall survival benefits.
Assuntos
Antineoplásicos Fitogênicos/administração & dosagem , Barreira Hematoencefálica/fisiologia , Neoplasias Encefálicas/tratamento farmacológico , Etoposídeo/administração & dosagem , Glioblastoma/tratamento farmacológico , Ultrassonografia de Intervenção/métodos , Animais , Antineoplásicos Fitogênicos/análise , Barreira Hematoencefálica/diagnóstico por imagem , Neoplasias Encefálicas/química , Neoplasias Encefálicas/diagnóstico por imagem , Neoplasias Encefálicas/mortalidade , Linhagem Celular Tumoral , Cromatografia Líquida , Meios de Contraste/administração & dosagem , Progressão da Doença , Etoposídeo/análise , Glioblastoma/química , Glioblastoma/diagnóstico por imagem , Glioblastoma/mortalidade , Imageamento por Ressonância Magnética , Masculino , Camundongos , Microbolhas , Sonicação , Espectrometria de Massas em TandemRESUMO
Drug delivery in diffuse intrinsic pontine glioma is significantly limited by the blood-brain barrier (BBB). Focused ultrasound (FUS), when combined with the administration of microbubbles can effectively open the BBB permitting the entry of drugs across the cerebrovasculature into the brainstem. Given that the utility of FUS in brainstem malignancies remains unknown, the purpose of our study was to determine the safety and feasibility of this technique in a murine pontine glioma model. A syngeneic orthotopic model was developed by stereotactic injection of PDGF-B+PTEN-/-p53-/- murine glioma cells into the pons of B6 mice. A single-element, spherical-segment 1.5 MHz ultrasound transducer driven by a function generator through a power amplifier was used with concurrent intravenous microbubble injection for tumor sonication. Mice were randomly assigned to control, FUS and double-FUS groups. Pulse and respiratory rates were continuously monitored during treatment. BBB opening was confirmed with gadolinium-enhanced MRI and Evans blue. Kondziela inverted screen testing and sequential weight lifting measured motor function before and after sonication. A subset of animals were treated with etoposide following ultrasound. Mice were either sacrificed for tissue analysis or serially monitored for survival with daily weights. FUS successfully caused BBB opening while preserving normal cardiorespiratory and motor function. Furthermore, the degree of intra-tumoral hemorrhage and inflammation on H&E in control and treated mice was similar. There was also no difference in weight loss and survival between the groups (p > 0.05). Lastly, FUS increased intra-tumoral etoposide concentration by more than fivefold. FUS is a safe and feasible technique for repeated BBB opening and etoposide delivery in a preclinical pontine glioma model.
Assuntos
Barreira Hematoencefálica/efeitos dos fármacos , Neoplasias do Tronco Encefálico/tratamento farmacológico , Sistemas de Liberação de Medicamentos , Glioma/tratamento farmacológico , Animais , Transporte Biológico/efeitos dos fármacos , Tronco Encefálico/diagnóstico por imagem , Tronco Encefálico/efeitos dos fármacos , Neoplasias do Tronco Encefálico/diagnóstico por imagem , Neoplasias do Tronco Encefálico/genética , Neoplasias do Tronco Encefálico/patologia , Modelos Animais de Doenças , Etoposídeo/farmacologia , Azul Evans/farmacologia , Gadolínio/farmacologia , Glioma/diagnóstico por imagem , Glioma/genética , Glioma/patologia , Humanos , Imageamento por Ressonância Magnética , Camundongos , PTEN Fosfo-Hidrolase/genética , PTEN Fosfo-Hidrolase/farmacologia , Ponte/diagnóstico por imagem , Ponte/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-sis/genética , Proteínas Proto-Oncogênicas c-sis/farmacologia , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/farmacologia , UltrassonografiaRESUMO
Focused ultrasound (FUS)-mediated blood-brain barrier (BBB) opening is currently being investigated in clinical trials. Here, we describe a portable clinical system with a therapeutic transducer suitable for humans, which eliminates the need for in-line magnetic resonance imaging (MRI) guidance. A neuronavigation-guided 0.25-MHz single-element FUS transducer was developed for non-invasive clinical BBB opening. Numerical simulations and experiments were performed to determine the characteristics of the FUS beam within a human skull. We also validated the feasibility of BBB opening obtained with this system in two non-human primates using U.S. Food and Drug Administration (FDA)-approved treatment parameters. Ultrasound propagation through a human skull fragment caused 44.4 ± 1% pressure attenuation at a normal incidence angle, while the focal size decreased by 3.3 ± 1.4% and 3.9 ± 1.8% along the lateral and axial dimension, respectively. Measured lateral and axial shifts were 0.5 ± 0.4 mm and 2.1 ± 1.1 mm, while simulated shifts were 0.1 ± 0.2 mm and 6.1 ± 2.4 mm, respectively. A 1.5-MHz passive cavitation detector transcranially detected cavitation signals of Definity microbubbles flowing through a vessel-mimicking phantom. T1-weighted MRI confirmed a 153 ± 5.5 mm3 BBB opening in two non-human primates at a mechanical index of 0.4, using Definity microbubbles at the FDA-approved dose for imaging applications, without edema or hemorrhage. In conclusion, we developed a portable system for non-invasive BBB opening in humans, which can be achieved at clinically relevant ultrasound exposures without the need for in-line MRI guidance. The proposed FUS system may accelerate the adoption of non-invasive FUS-mediated therapies due to its fast application, low cost and portability.
Assuntos
Barreira Hematoencefálica/diagnóstico por imagem , Neuronavegação/métodos , Transdutores , Animais , Desenho de Equipamento , Humanos , Neuronavegação/instrumentação , Primatas , UltrassonografiaRESUMO
Non-invasive blood-brain barrier (BBB) opening using focused ultrasound (FUS) is being tested as a means to locally deliver drugs into the brain. Such FUS therapies require injection of preformed microbubbles, currently used as contrast agents in ultrasound imaging. Although their behavior during exposure to imaging sequences has been well described, our understanding of microbubble stability within a therapeutic field is still not complete. Here, we study the temporal stability of lipid-shelled microbubbles during therapeutic FUS exposure in two timescales: the short time scale (i.e., µs of low-frequency ultrasound exposure) and the long time scale (i.e., days post-activation). We first simulated the microbubble response to low-frequency sonication, and found a strong correlation between viscosity and fragmentation pressure. Activated microbubbles had a concentration decay constant of 0.02 d-1 but maintained a quasi-stable size distribution for up to 3 weeks (< 10% variation). Microbubbles flowing through a 4-mm vessel within a tissue-mimicking phantom (5% gelatin) were exposed to therapeutic pulses (fc: 0.5 MHz, peak-negative pressure: 300 kPa, pulse length: 1 ms, pulse repetition frequency: 1 Hz, n=10). We recorded and analyzed their acoustic emissions, focusing on emitted energy and its temporal evolution, alongside the frequency content. Measurements were repeated with concentration-matched samples (107 microbubbles/ml) on day 0, 7, 14, and 21 after activation. Temporal stability decreased while inertial cavitation response increased with storage time both in vitro and in vivo, possibly due to changes in the shell lipid content. Using the same parameters and timepoints, we performed BBB opening in a mouse model (n=3). BBB opening volume measured through T1-weighted contrast-enhanced MRI was equal to 19.1 ± 7.1 mm3, 21.8 ± 14 mm3, 29.3 ± 2.5 mm3, and 38 ± 20.1 mm3 on day 0, 7, 14, and 21, respectively, showing no significant difference over time (p-value: 0.49). Contrast enhancement was 24.9 ± 1.7 %, 23.7 ± 11.7 %, 28.9 ± 5.3 %, and 35 ± 13.4 %, respectively (p-value: 0.63). In conclusion, the in-house made microbubbles studied here maintain their capacity to produce similar therapeutic effects over a period of 3 weeks after activation, as long as the natural concentration decay is accounted for. Future work should focus on stability of commercially available microbubbles and tailoring microbubble shell properties towards therapeutic applications.