Article

# Targeted delivery of antibodies through the blood–brain barrier by MRI-guided focused ultrasound

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## Abstract

The blood-brain barrier (BBB) is a persistent obstacle for the local delivery of macromolecular therapeutic agents to the central nervous system (CNS). Many drugs that show potential for treating CNS diseases cannot cross the BBB and there is a need for a non-invasive targeted drug delivery method that allows local therapy of the CNS using larger molecules. We developed a non-invasive technique that allows the image-guided delivery of antibody across the BBB into the murine CNS. Here, we demonstrate that subsequent to MRI-targeted focused ultrasound induced disruption of BBB, intravenously administered dopamine D(4) receptor-targeting antibody crossed the BBB and recognized its antigens. Using MRI, we were able to monitor the extent of BBB disruption. This novel technology should be useful in delivering macromolecular therapeutic or diagnostic agents to the CNS for the treatment of various CNS disorders.

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... [21][22][23][24] FUS represents a very promising strategy to overcome this limitation. 25 Several studies have reported positive results when using FUS to enhance antibody penetration across the BBB for the treatment of Alzheimer's disease [25][26][27][28] and a small number of studies have investigated the effects of combining FUS with antibody delivery on tumor growth, immune response, and animal survival in mouse or rat models of glioma. [29][30][31][32][33] While these studies reported promising preclinical results, there are a number of critical limitations that must be addressed before the clinical translation is considered. ...
... [21][22][23][24] FUS represents a very promising strategy to overcome this limitation. 25 Several studies have reported positive results when using FUS to enhance antibody penetration across the BBB for the treatment of Alzheimer's disease [25][26][27][28] and a small number of studies have investigated the effects of combining FUS with antibody delivery on tumor growth, immune response, and animal survival in mouse or rat models of glioma. [29][30][31][32][33] While these studies reported promising preclinical results, there are a number of critical limitations that must be addressed before the clinical translation is considered. ...
... It is often suggested that FUS-induced extravasation of molecules of interest in the healthy brain is sufficient evidence to infer that FUS will increase extravasation in nonenhancing HGG tissue. 16,25,29,[33][34][35][36] However, due to the significant differences in vascular architecture and regional blood flow between a healthy brain and nonenhancing HGG tissue, it is unlikely that mechanisms of FUS-induced BBB disruption will be consistent between a healthy brain and nonenhancing HGG tissue. ...
Article
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Background High-grade glioma (HGG) remains a recalcitrant clinical problem despite many decades of research. A major challenge in improving prognosis is the inability of current therapeutic strategies to address a clinically significant burden of infiltrating tumor cells that extend beyond the margins of the primary tumor mass. Such cells cannot be surgically excised nor efficiently targeted by radiation therapy. Therapeutic targeting of this tumor cell population is significantly hampered by the presence of an intact blood-brain barrier (BBB). In this study we performed a preclinical investigation of the efficiency of MR-guided Focused Ultrasound (FUS) to temporarily disrupt the BBB to allow selective delivery of a tumor-targeting antibody to infiltrating tumor. Methods Structural MRI, dynamic-contrast enhancement MRI and histology were used to fully characterize the MR-enhancing properties of a patient derived xenograft (PDX) orthotopic mouse model of HGG, and to develop a reproducible, robust model of non-enhancing HGG. PET-CT imaging techniques were then used to evaluate the efficacy of FUS to increase 89Zr-radiolabeled antibody concentration in non-enhancing HGG regions, and adjacent non-targeted tumor tissue. Results The PDX mouse model of HGG has significant tumor burden lying behind an intact BBB. Increased antibody uptake in non-enhancing tumor regions is directly proportional to the FUS-targeted volume. FUS locally increased antibody uptake in FUS-targeted regions of the tumor with an intact BBB, while leaving untargeted regions unaffected. Conclusions FUS exposure successfully allowed temporary BBB disruption, localized to specifically targeted, non-enhancing, infiltrating tumor regions and delivery of a systemically administered antibody was significantly increased.
... The ability to observe BBB disruption in vivo is conferred by CE-T1 MRI, without requiring postmortem histological analysis. Multiple investigations have shown correlative relationships between the extravasation of the MRI contrast agent and that of histological tracers [61,62] and some therapeutic agents, including Herceptin [31], doxorubicin [63], and nanoparticles [23,38]. A few studies have employed the use of T2/T2* weighted MRI to track the uptake of superparamagnetic iron oxide (SPIO)-labelled drug molecules into sonicated brain parenchyma. ...
... Reported histopathological outcomes have been identified at a variety of endpoints, from immediately following [115] to months after initial sonication [123], highlighting both the potential acuteness and chronicity at which ultrasound-mediated BBB disruption may exert unwanted biological effects. Histopathological assessments generally reinforce pathological findings on MRI, but in some studies [62,92,93], they appear to highlight pathological change in the absence of any on MRI, despite equivalent timing of data acquisition, suggesting higher sensitivity for adverse pathological change. ...
... Additionally, significant patient neurological comorbidity in these trials makes it difficult to directly attribute adverse events to ultrasound-mediated BBB disruption. X [76] X [60] X X [77] X X [68] X [78] X X X [61] X X X X (PCR + WB) [31] X X X X [62] X X X X X X [54] X X X [52] X X X X [9] X X [10] X X X X [11] X X [53] X ...
Article
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Ultrasound-mediated blood–brain barrier (BBB) disruption has garnered focus as a method of delivering normally impenetrable drugs into the brain. Numerous studies have investigated this approach, and a diverse set of ultrasound parameters appear to influence the efficacy and safety of this approach. An understanding of these findings is essential for safe and reproducible BBB disruption, as well as in identifying the limitations and gaps for further advancement of this drug delivery approach. We aimed to collate and summarise protocols and parameters for achieving ultrasound-mediated BBB disruption in animal and clinical studies, as well as the efficacy and safety methods and outcomes associated with each. A systematic search of electronic databases helped in identifying relevant, included studies. Reference lists of included studies were further screened to identify supplemental studies for inclusion. In total, 107 articles were included in this review, and the following parameters were identified as influencing efficacy and safety outcomes: microbubbles, transducer frequency, peak-negative pressure, pulse characteristics, and the dosing of ultrasound applications. Current protocols and parameters achieving ultrasound-mediated BBB disruption, as well as their associated efficacy and safety outcomes, are identified and summarised. Greater standardisation of protocols and parameters in future preclinical and clinical studies is required to inform robust clinical translation.
... Since this initial study [16], FUS+MB treatments have been used to successfully deliver a variety of agents across the BBB, resulting in functional benefits. Chemotherapeutics delivered to the brain using FUS+MBs resulted in increased concentrations of drug within brain tumours, reduced tumour growth, and improved median survival in rodents [25][26][27][28][29]. Antibodies have been delivered to the brain parenchyma using this technique [30,31], leading to functional effects such as β-amyloid plaque reduction [32]. FUS+MB treatments have also been used to deliver cells [33,34], neurotrophic factors [35], nucleic acids [36], viral vectors [37][38][39], non-viral genes encapsulated in liposomes [40], and brain-penetrating nanoparticles [41,42]. ...
... Histological methods have been used extensively to characterize the impact of FUS+MB treatments on both BBB permeability and tissue health (Figure 3). Extravasation of dyes, such as trypan blue and Evans blue [30,46], are sensitive tools for confirming changes in BBB permeability in preclinical studies. Immunohistochemistry and immunoblot analyses of extravasated exogenous [30,118,119] and endogenous antibodies [32,118,120] have also been used for this purpose. ...
... Extravasation of dyes, such as trypan blue and Evans blue [30,46], are sensitive tools for confirming changes in BBB permeability in preclinical studies. Immunohistochemistry and immunoblot analyses of extravasated exogenous [30,118,119] and endogenous antibodies [32,118,120] have also been used for this purpose. Significant increases in the levels of IgG and IgM in the brain parenchyma have been observed from one hour to four days after FUS+MB treatment in mice and rabbit brains [32,118,120]. ...
Article
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Introduction: Treatment of several diseases of the brain are complicated by the presence of the skull and the blood-brain barrier (BBB). Focused ultrasound (FUS) and microbubble (MB)-mediated BBB treatment is a minimally invasive method to transiently increase the permeability of blood vessels in targeted brain areas. It can be used as a general delivery system to increase the concentration of therapeutic agents in the brain parenchyma. Areas covered: Over the past two decades, the safety of using FUS+MBs to deliver agents across the BBB has been interrogated through various methods of imaging, histology, biochemical assays, and behavior analyses. Here we provide an overview of the factors that affect the safety profile of these treatments, describe methods by which FUS+MB treatments are controlled, and discuss data that have informed the assessment of treatment risks. Expert opinion: There remains a need to assess the risks associated with clinically relevant treatment strategies, specifically repeated FUS+MB treatments, with and without therapeutic agent delivery. Additionally, efforts to develop metrics by which FUS+MB treatments can be easily compared across studies would facilitate a more rapid consensus on the risks associated with this intervention.
... Microbubbles of 1-5 µm (reF. 278 ) oscillate during exposure to Fus, resulting in transient opening of the BBB at Fus-targeted sites, allowing blood-borne therapeutics and imaging agents to enter the brain at the targeted regions [279][280][281] . Fus parameters, such as burst duration and repetition frequency, pressure amplitude and exposure time, have to be optimized to achieve transient BBB opening and minimize Fus-MB-related brain damage 275,281 . ...
... Fus parameters, such as burst duration and repetition frequency, pressure amplitude and exposure time, have to be optimized to achieve transient BBB opening and minimize Fus-MB-related brain damage 275,281 . Fus-MB-induced BBB opening lasts approximately 1-4 hours and is recovered after about 24 hours 278 . in mice, Fus-MB allowed molecules of 3-2,000 kDa -such as doxorubicin, trastuzumab, rNa molecules and contrast imaging agents -to enter the brain 279,280,[282][283][284][285] , and was also found to increase transcytosis and fenestration of BBB cells in sonicated regions 286 . a pilot study using transcranial Mri-guided Fus (MrgFus) for site-specific BBB opening in patients with brain tumours intravenously infused with doxorubicin or temozolomide 287 showed MrgFus was well tolerated without adverse events. ...
... this trial was based on promising preclinical results 289,290 , and the results are yet to be published. safety is a major concern for Fus-MB-induced BBB opening as Fus has been shown to cause microhaemorrhages in the mouse brain 279,280,291 , and mechanical effects caused by Fus forces and microbubbles are known to damage blood vessels 292 . Furthermore, dark neurons and microvacuolated sites were observed 30 minutes after sonication, as were the functional effects of reduced blood oxygen level-dependent responses and functional connectivity in sonicated regions 291 . ...
Article
Achieving sufficient delivery across the blood–brain barrier is a key challenge in the development of drugs to treat central nervous system (CNS) disorders. This is particularly the case for biopharmaceuticals such as monoclonal antibodies and enzyme replacement therapies, which are largely excluded from the brain following systemic administration. In recent years, increasing research efforts by pharmaceutical and biotechnology companies, academic institutions and public–private consortia have resulted in the evaluation of various technologies developed to deliver therapeutics to the CNS, some of which have entered clinical testing. Here we review recent developments and challenges related to selected blood–brain barrier-crossing strategies — with a focus on non-invasive approaches such as receptor-mediated transcytosis and the use of neurotropic viruses, nanoparticles and exosomes — and analyse their potential in the treatment of CNS disorders. The blood–brain barrier is a perennial challenge for the delivery of therapeutics to the central nervous system. In their Review, Terstappen and colleagues discuss non-invasive approaches to brain delivery, particularly for biopharmaceuticals, some of which are now in clinical testing.
... Microbubbles of 1-5 µm (reF. 278 ) oscillate during exposure to Fus, resulting in transient opening of the BBB at Fus-targeted sites, allowing blood-borne therapeutics and imaging agents to enter the brain at the targeted regions [279][280][281] . Fus parameters, such as burst duration and repetition frequency, pressure amplitude and exposure time, have to be optimized to achieve transient BBB opening and minimize Fus-MB-related brain damage 275,281 . ...
... Fus parameters, such as burst duration and repetition frequency, pressure amplitude and exposure time, have to be optimized to achieve transient BBB opening and minimize Fus-MB-related brain damage 275,281 . Fus-MB-induced BBB opening lasts approximately 1-4 hours and is recovered after about 24 hours 278 . in mice, Fus-MB allowed molecules of 3-2,000 kDa -such as doxorubicin, trastuzumab, rNa molecules and contrast imaging agents -to enter the brain 279,280,[282][283][284][285] , and was also found to increase transcytosis and fenestration of BBB cells in sonicated regions 286 . a pilot study using transcranial Mri-guided Fus (MrgFus) for site-specific BBB opening in patients with brain tumours intravenously infused with doxorubicin or temozolomide 287 showed MrgFus was well tolerated without adverse events. ...
... this trial was based on promising preclinical results 289,290 , and the results are yet to be published. safety is a major concern for Fus-MB-induced BBB opening as Fus has been shown to cause microhaemorrhages in the mouse brain 279,280,291 , and mechanical effects caused by Fus forces and microbubbles are known to damage blood vessels 292 . Furthermore, dark neurons and microvacuolated sites were observed 30 minutes after sonication, as were the functional effects of reduced blood oxygen level-dependent responses and functional connectivity in sonicated regions 291 . ...
Article
Full-text available
Achieving sufficient delivery across the blood–brain barrier is a key challenge in the development of drugs to treat central nervous system (CNS) disorders. This is particularly the case for biopharmaceuticals such as monoclonal antibodies and enzyme replacement therapies, which are largely excluded from the brain following systemic administration. In recent years, increasing research efforts by pharmaceutical and biotechnology companies, academic institutions and public–private consortia have resulted in the evaluation of various technologies developed to deliver therapeutics to the CNS, some of which have entered clinical testing. Here we review recent developments and challenges related to selected blood–brain barrier- crossing strategies — with a focus on non- invasive approaches such as receptor- mediated transcytosis and the use of neurotropic viruses, nanoparticles and exosomes — and analyse their potential in the treatment of CNS disorders.
... Delivery of therapeutic agents to the brain has proven a particularly attractive target for the US plus cavitation nuclei approach. In such studies, US has been used in conjunction with intravenously (iv.) administered microbubbles (typically US contrast agents [UCAs]) to temporarily open the blood-brain barrier (BBB) of a nontumor bearing animal and allow for extravasation of large macromolecules such as antibodies [22][23][24][25]. Kinoshita et al. showed that trastuzumab could be quantified in the brain in sites exposed to sonication with PRFPs of 0.6-0.8 ...
... Recent murine studies using an oncolytic virus provided an elegant demonstration of how alterations to systemic blood pressure (with exercise or anesthesia) can have an impact on tumor uptake [64], although standardizing and clinically translating such observations could be a challenge. Endeavors to assist tumoral drug uptake using external stimuli to generate heat or cavitation events have also gained much research focus [10,23,27,[65][66][67]. Although the targeted heating of tumors at depth can be hard to control and may require the deposition of large amounts of energy and therefore expensive and scarce equipment [68], instigation of US-mediated cavitation events is potentially more broadly and cheaply applicable. ...
Article
Aims: Testing ultrasound-mediated cavitation for enhanced delivery of the therapeutic antibody cetuximab to tumors in a mouse model. Methods: Tumors with strong EGF receptor expression were grown bilaterally. Cetuximab was coadministered intravenously with cavitation nuclei, consisting of either the ultrasound contrast agent Sonovue or gas-stabilizing nanoscale SonoTran Particles. One of the two tumors was exposed to focused ultrasound. Passive acoustic mapping localized and monitored cavitation activity. Both tumors were then excised and cetuximab concentration was quantified. Results: Cavitation increased tumoral cetuximab concentration. When nucleated by Sonovue, a 2.1-fold increase (95% CI 1.3- to 3.4-fold) was measured, whereas SonoTran Particles gave a 3.6-fold increase (95% CI 2.3- to 5.8-fold). Conclusions: Ultrasound-mediated cavitation, especially when nucleated by nanoscale gas-entrapping particles, can noninvasively increase site-specific delivery of therapeutic antibodies to solid tumors.
... In addition to delivery of anticancer chemotherapeutic agents, MB-ultrasound BBB opening has been shown to successfully deliver therapeutic proteins or monoclonal antibodies for brain tumor treatment. BBB opening can be used to deliver large molecular therapeutic monoclonal antibodies including herceptin [166], dopamine D-4 receptortargeting antibody [167], bevacizumab [168], and trastuzumab [169]. Kinoshita et al. demonstrated the possibility of delivering various large molecular monoclonal antibodies with penetration through the BBB and into the brain region, including dopamine D-4 receptor-targeting antibody [166] and humanized anti-human epidermal growth factor receptor 2 (HER2/c-erbB2) monoclonal antibody [167]. ...
... BBB opening can be used to deliver large molecular therapeutic monoclonal antibodies including herceptin [166], dopamine D-4 receptortargeting antibody [167], bevacizumab [168], and trastuzumab [169]. Kinoshita et al. demonstrated the possibility of delivering various large molecular monoclonal antibodies with penetration through the BBB and into the brain region, including dopamine D-4 receptor-targeting antibody [166] and humanized anti-human epidermal growth factor receptor 2 (HER2/c-erbB2) monoclonal antibody [167]. In addition, it has been reported that enhanced bevacizumab delivery with BBB opening led to a significant increase in animal survival compared to the bevacizumab treatment group alone [168]. ...
Article
Full-text available
Ultrasonic technologies show great promise for diagnostic imaging and drug delivery in theranostic applications. The development of functional and molecular ultrasound imaging is based on the technical breakthrough of high frame–rate ultrasound. The evolution of shear wave elastography, high-frequency ultrasound imaging, ultrasound contrast imaging, and super-resolution blood flow imaging are described in this review. Recently, the therapeutic potential of the interaction of ultrasound with microbubble cavitation or droplet vaporization has become recognized. Microbubbles and phase-change droplets not only provide effective contrast media, but also show great therapeutic potential. Interaction with ultrasound induces unique and distinguishable biophysical features in microbubbles and droplets that promote drug loading and delivery. In particular, this approach demonstrates potential for central nervous system applications. Here, we systemically review the technological developments of theranostic ultrasound including novel ultrasound imaging techniques, the synergetic use of ultrasound with microbubbles and droplets, and microbubble/droplet drug-loading strategies for anticancer applications and disease modulation. These advancements have transformed ultrasound from a purely diagnostic utility into a promising theranostic tool.
... Tremendous effort has been invested in improving antibody delivery into the brain [161][162][163][164]. For instance, Kinoshita et al. introduced a technique to increase antibody brain delivery through transiently disrupting BBB by ultrasound, opening up tight cellular junctions, and facilitating the antibody penetration in the brain [164]. ...
... Tremendous effort has been invested in improving antibody delivery into the brain [161][162][163][164]. For instance, Kinoshita et al. introduced a technique to increase antibody brain delivery through transiently disrupting BBB by ultrasound, opening up tight cellular junctions, and facilitating the antibody penetration in the brain [164]. RMT-based antibody delivery has gained momentum as a viable method to treat central nervous system (CNS) disorders [78,[165][166][167][168]. ...
Article
Full-text available
With more than 90 approved drugs by 2020, therapeutic antibodies have played a central role in shifting the treatment landscape of many diseases, including autoimmune disorders and cancers. While showing many therapeutic advantages such as long half-life and highly selective actions, therapeutic antibodies still face many outstanding issues associated with their pharmacokinetics (PK) and pharmacodynamics (PD), including high variabilities, low tissue distributions, poorly-defined PK/PD characteristics for novel antibody formats, and high rates of treatment resistance. We have witnessed many successful cases applying PK/PD modeling to answer critical questions in therapeutic antibodies’ development and regulations. These models have yielded substantial insights into antibody PK/PD properties. This review summarized the progress, challenges, and future directions in modeling antibody PK/PD and highlighted the potential of applying mechanistic models addressing the development questions.
... Early studies show the transducer frequency, ultrasound intensity and microbubble size and dosage can affect the extent of BBB disruption and its safety profile [26]. At 1.5 MHz, grade 3 hemorrhage with apoptosis and tissue damage was detected with ultrasound pressure of 2.4 MPa at 690 kHz, and grade 2 with 1.6 MPa [27,28]. ...
... Journal of Controlled Release 309 (2019)[25][26][27][28][29][30][31][32][33][34][35][36] ...
Article
The blood-brain barrier (BBB), while fundamental in maintaining homeostasis in the central nervous system, is a bottleneck to achieving efficacy for numerous therapeutics. Improved brain penetration is also desirable for reduced dose, cost, and systemic side effects. Transient disruption of the BBB with focused ultrasound (FUS) can facilitate drug delivery noninvasively and with precise spatial and temporal specificity. FUS technology is transcranial and effective without further drug modifications, key advantages that will accelerate adoption and translation of existing therapeutic pipelines. In this review, we performed a comprehensive literature search to build a database and provide a synthesis of ultrasound parameters and drug characteristics that influence the safety and efficacy profile of FUS to enhance drug delivery.
... Multi-parametric, long-term studies have shown that repeated US-induced disruption of the BBB was clinically well tolerated in primates [26][27][28]. A large variety of antineoplastic molecules and agents have been shown to have enhanced delivery to the brain after BBBD including small molecular weight drugs [29], monoclonal antibodies [30], enzymes [31], neurotrophic factors [32], DNA [33], and cells [34]. Both tumor control and increased survival times have been achieved in preclinical tumor models of glioma after US-induced opening of the BBB and systemic injection of drugs [35][36][37]. ...
... Multiple other studies have also shown that this strategy can enhance the delivery of intravenously-injected antibodies to the brain. For example, Dopamine D4 receptor-targeting antibodies were delivered to the hippocampus and basal ganglia in mice after BBB opening, whereas no antibodies were detected in the contralateral and unsonicated brain regions [30]. The same team was able to deliver trastuzumab, a humanized antihuman epidermal growth factor receptor 2 monoclonal antibody, to mouse brain parenchyma, with the amount of the antibody delivered directly correlating with the MRI signal change after gadolinium injection, allowing for imaging assessment of delivery [49]. ...
Article
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IntroductionOpening of the blood–brain barrier (BBB) by pulsed low intensity ultrasound has been developed during the last decade and is now recognized as a safe technique to transiently and repeatedly open the BBB. This non- or minimally invasive technique allows for a targeted and uniform dispersal of a wide range of therapeutic substances throughout the brain, including immune cells and antibodies.Methods In this review article, we summarize pre-clinical studies that have used BBB-opening by pulsed low intensity ultrasound to enhance the delivery of immune therapeutics and effector cell populations, as well as several recent clinical studies that have been initiated. Based on this analysis, we propose immune therapeutic strategies that are most likely to benefit from this strategy. The literature review and trial data research were performed using Medline/Pubmed databases and clinical trial registry www.clinicaltrials.gov. The reference lists of all included articles were searched for additional studies.ResultsA wide range of immune therapeutic agents, including small molecular weight drugs, antibodies or NK cells, have been safely and efficiently delivered to the brain with pulsed low intensity ultrasound in preclinical models, and both tumor control and increased survival have been demonstrated in different types of brain tumor models in rodents. Ultrasound-induced BBB disruption may also stimulate innate and cellular immune responses.Conclusions Ultrasound BBB opening has just recently entered clinical trials with encouraging results, and the association of this strategy with immune therapeutics creates a new field of brain tumor treatment.
... Thus, by applying different acoustic pressures, the two target areas showed comparable degrees of BBBD, comparable patterns of DTI parameters, and histological results. Since the FUS parameter in group-0.65 MPa has been widely used in the rat brain to safely disrupt the BBB and deliver therapeutics in several previous studies (Kinoshita et al., 2006b;Cho et al., 2016;Jung et al., 2019), we selected the FUS parameter to determine whether water transport could be facilitated in the BBBD brain area. We additionally applied higher sonication conditions in group-0.80 ...
Article
Full-text available
Interstitial solutes can be removed by various overlapping clearance systems, including blood–brain barrier (BBB) transport and glymphatic clearance. Recently, focused ultrasound (FUS)-induced BBB disruption (BBBD) has been applied to visualize glymphatic transport. Despite evidence that FUS–BBBD might facilitate glymphatic transport, the nature of fluid movement within the sonication region is yet to be determined. In this study, we sought to determine whether FUS–BBBD may facilitate the local movement of water molecules. Two different FUS conditions (0.60–0.65 MPa and 0.75–0.80 MPa) were used to induce BBBD in the caudate-putamen and thalamus regions of healthy Sprague–Dawley rats. The water diffusion caused by FUS–BBBD was analyzed using the apparent diffusion coefficient (ADC), axial diffusivity, radial diffusivity (RD), and fractional anisotropy, obtained at 5 min, 24 and 48 h, as well as the water channel expression of aquaporin-4 (AQP-4) immunostaining at 48 h after FUS-induced BBBD. In addition, hematoxylin and eosin histopathology and Fluoro-Jade C (FJC) immunostaining were performed to analyze brain damage. The signal changes in ADC and RD in the sonication groups showed significant and transient reduction at 5 min, with subsequent increases at 24 and 48 h after FUS-induced BBBD. When we applied higher sonication conditions, the ADC and RD showed enhancement until 48 h, and became comparable to contralateral values at 72 h. AQP-4 expression was upregulated after FUS-induced BBBD in both sonication conditions at 48 h. The results of this study provide preliminary evidence on how mechanical forces from FUS alter water dynamics through diffusion tensor imaging (DTI) measures and AQP4 expression.
... Ultrasound-based blood-brain barrier disruption (US-BBBD) allows for increased penetration of systemically administered small and large molecule drug therapies into the brain [13][14][15]. Strikingly, US-BBBD alone has been shown to reduce β-amyloid and tau pathologies, stimulate neurogenesis, and improve cognitive performance in mouse AD models [16][17][18] and can be further coupled with drug therapies to improve their brain penetration and efficacy [13,19,20]. ...
Article
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Background Temporary disruption of the blood-brain barrier (BBB) using pulsed ultrasound leads to the clearance of both amyloid and tau from the brain, increased neurogenesis, and mitigation of cognitive decline in pre-clinical models of Alzheimer’s disease (AD) while also increasing BBB penetration of therapeutic antibodies. The goal of this pilot clinical trial was to investigate the safety and efficacy of this approach in patients with mild AD using an implantable ultrasound device. Methods An implantable, 1-MHz ultrasound device (SonoCloud-1) was implanted under local anesthesia in the skull (extradural) of 10 mild AD patients to target the left supra-marginal gyrus. Over 3.5 months, seven ultrasound sessions in combination with intravenous infusion of microbubbles were performed twice per month to temporarily disrupt the BBB. ¹⁸ F-florbetapir and ¹⁸ F-fluorodeoxyglucose positron emission tomography (PET) imaging were performed on a combined PET/MRI scanner at inclusion and at 4 and 8 months after the initiation of sonications to monitor the brain metabolism and amyloid levels along with cognitive evaluations. The evolution of cognitive and neuroimaging features was compared to that of a matched sample of control participants taken from the Alzheimer’s Disease Neuroimaging Initiative (ADNI). Results A total of 63 BBB opening procedures were performed in nine subjects. The procedure was well-tolerated. A non-significant decrease in amyloid accumulation at 4 months of − 6.6% (SD = 7.2%) on ¹⁸ F-florbetapir PET imaging in the sonicated gray matter targeted by the ultrasound transducer was observed compared to baseline in six subjects that completed treatments and who had evaluable imaging scans. No differences in the longitudinal change in the glucose metabolism were observed compared to the neighboring or contralateral regions or to the change observed in the same region in ADNI participants. No significant effect on cognition evolution was observed in comparison with the ADNI participants as expected due to the small sample size and duration of the trial. Conclusions These results demonstrate the safety of ultrasound-based BBB disruption and the potential of this technology to be used as a therapy for AD patients. Research of this technique in a larger clinical trial with a device designed to sonicate larger volumes of tissue and in combination with disease-modifying drugs may further enhance the effects observed. Trial registration ClinicalTrials.gov, NCT03119961
... Another method for delivering drugs to the brain is the disruption of the BBB. This method can be applied in different ways; disruption by osmotic pressure, MRI-guided focused ultrasound the BBB, application of bradykininanalog [81][82][83]. All these methods are expensive, require hospitalization, and are non-friendly patients. ...
Article
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The blood-brain barrier is one of the most complicated barrier to pass for therapeutic drugs. Because of the structure of the blood-brain barrier, only a few small molecules with appropriate lipophilicity, molecular weight, and charge can penetrate through the blood-brain barrier and pass in the central nervous system. Because of this unique property, blood-brain barrier is still a major problem for the treatment of central nervous system diseases. In the last decades, many strategies to overcome this barrier have been investigated. Compared to other drug delivery strategies, due to the reduced side effects and no requirement for surgical operations, brain targeted nanoparticle is one the most promising and popular strategy used do deliver drugs to the brain. Many in vitro and in vivo preclinical studies have been conducted to determine optimum brain targeted nanoparticles. These studies were reported that characteristics of nanoparticles such as particle size, zeta potential, and targeting ligand are critical to achieving the goals. In this review, first of all, the structure of the blood-brain barrier and possible causes of blood-brain barrier disruption were summarized. Later, previous strategies of brain targeted drug delivery and characteristic prosperities for optimized brain-targeted nanoparticles were evaluated. Moreover, different strategies, such as focus ultrasound, which can increase the effectiveness of nanoparticular system applications, are mentioned.
... The flexibility of using FUS+MB exposures to enhance the delivery of a variety of therapeutic agents to targeted brain regions has been demonstrated under numerous experimental conditions [1][2][3][4][5][6]. By transiently increasing the permeability of the BBB [7], drugs can be delivered systemically and extravasate in the targeted locations to exert a therapeutic effect. ...
Article
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Rationale: Clinical trials are currently underway to test the safety and efficacy of delivering therapeutic agents across the blood-brain barrier (BBB) using focused ultrasound and microbubbles (FUS+MBs). While acoustic feedback control strategies have largely minimized the risk of overt tissue damage, transient induction of inflammatory processes have been observed following sonication in preclinical studies. The goal of this work was to explore the potential of post-sonication dexamethasone (DEX) administration as a means to mitigate treatment risk. Vascular permeability, inflammatory protein expression, blood vessel growth, and astrocyte activation were assessed. Methods: A single-element focused transducer (transmit frequency = 580 kHz) and DefinityTM microbubbles were used to increase BBB permeability unilaterally in the dorsal hippocampi of adult male rats. Sonicating pressure was calibrated based on ultraharmonic emissions. Dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) was used to quantitatively assess BBB permeability at 15 min (baseline) and 2 hrs following sonication. DEX was administered following baseline imaging and at 24 hrs post-FUS+MB exposure. Expression of key inflammatory proteins were assessed at 2 days, and astrocyte activation and blood vessel growth were assessed at 10 days post-FUS+MB exposure. Results: Compared to saline-treated control animals, DEX administration expedited the restoration of BBB integrity at 2 hrs, and significantly limited the production of key inflammation-related proteins at 2 days, following sonication. Indications of FUS+MB-induced astrocyte activation and vascular growth were diminished at 10 days in DEX-treated animals, compared to controls. Conclusions: These results suggest that DEX provides a means of modulating the duration of BBB permeability enhancement and may reduce the risk of inflammation-induced tissue damage, increasing the safety profile of this drug-delivery strategy. This effect may be especially relevant in scenarios for which the goal of treatment is to restore or preserve neural function and multiple sonications are required.
... MR-guided focused ultrasound (MRgFUS) is a non-invasive ultrasound-based strategy for BBBD (49)(50)(51)(52). A single-arm Phase I trial demonstrated BBBD by dynamic contrast-enhanced MRI (DCE-MRI) using MRgFUS in high-grade glioma (HGG) patients (53). ...
Article
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The blood-brain barrier (BBB) presents a formidable challenge in the development of effective therapeutics in neuro-oncology. This has fueled several decades of efforts to develop strategies for disrupting the BBB, but progress has not been satisfactory. As such, numerous drug- and device-based methods are currently being investigated in humans. Through a focused assessment of completed, active, and pending clinical trials, our first aim in this review is to outline the scientific foundation, successes, and limitations of the BBBD strategies developed to date. Among 35 registered trials relevant to BBBD in neuro-oncology in the ClinicalTrials.gov database, mannitol was the most common drug-based method, followed by RMP-7 and regadenoson. MR-guided focused ultrasound was the most common device-based method, followed by MR-guided laser ablation, ultrasound, and transcranial magnetic stimulation. While most early-phase studies focusing on safety and tolerability have met stated objectives, advanced-phase studies focusing on survival differences and objective tumor response have been limited by heterogeneous populations and tumors, along with a lack of control arms. Based on shared challenges among all methods, our second objective is to discuss strategies for confirmation of BBBD, choice of systemic agent and drug design, alignment of BBBD method with real-world clinical workflow, and consideration of inadvertent toxicity associated with disrupting an evolutionarily-refined barrier. Finally, we conclude with a strategic proposal to approach future studies assessing BBBD.
... Low-intensity focal ultrasound has been shown to be able to open the BBB in a noninvasive way. [83][84][85][86] This reversible opening time can be modulated by the intensity of the ultrasound. [87] Targeted ultrasound has been used to increase delivery of anti Aβeta antibodies to transgenic mice, [88] and this has demonstrated decrease in the Aβ load in the treated mice. ...
Article
Alzheimer's disease (AD) and other forms of dementia can have a large impact on patients, their families, and for the society as a whole. Current medical treatments have not shown enough potential in treating or altering the course of the disease. Deep brain stimulation (DBS) has shown great neuromodulatory potential in Parkinson's disease, and there is a growing body of evidence for justifying its use in cognitive disorders. At the same time there is mounting interest at less invasive and alternative modes of neuromodulation for the treatment of AD. This manuscript is a brief review of the infrastructure of memory, the current understanding of the pathophysiology of AD, and the body of preclinical and clinical evidence for noninvasive and invasive neuromodulation modalities for the treatment of cognitive disorders and AD in particular.
... Low intensity FUS with circulating MB is a form of mechanical FUS energy deposition that has been shown to elicit transient opening of tight junctions, sonoporation of vascular endothelium, and enhanced capacity for transcytosis; these effects have been exploited for blood brain barrier (BBB) and/or blood-tumor barrier (BTB) disruption [17]. This disruption can facilitate delivery of various agents to the brain, including chemotherapies, drug and gene-bearing polymeric nanoparticles, and antibodies [18][19][20][21][22][23][24][25][26]. ...
Article
Full-text available
Immunotherapy holds tremendous promise as a strategy for eradicating solid tumors. However, poor T cell infiltration and persistence within most solid tumor microenvironments, as well as mechanisms of adaptive resistance, continue to severely limit the accessibility of most immunotherapies to a broad patient population. This limitation perpetuates the demand for allied therapeutic strategies. Among such strategies is focused ultrasound (FUS), a non-invasive, non-ionizing technique for precisely targeted acoustic energy deposition into tissues. FUS has gained remarkable attention over recent years as a modality for elicitation of immune mechanisms in cancer and other pathologies. In 2017, we published a comprehensive review paper detailing existing evidence for immune modulation and therapy with FUS, as well as impending challenges and opportunities of consideration for the field. Over the last two years, a multitude of clinical trials have come online to explore safety, feasibility, and efficacy of FUS for cancers of the brain and periphery - including the first clinical trial to combine FUS with immunotherapy. Moreover, the last two years have seen a surge in FUS immunotherapy presentations at therapeutic ultrasound scientific meetings. Given the burst of activity in this field, we submit that an update on FUS immunotherapy progress is timely. In this review, we offer an updated overview and perspectives on scientific and clinical development in the FUS immunotherapy domain.
... In this scenario, minimally invasive techniques have attracted attention as alternatives to lumpectomy, such as radiofrequency ablation, cryosurgery, and high-intensity focused ultrasound (HIFU) [7]. Among these techniques, magnetic resonance-guided focused ultrasound surgery (MRgFUS) is a HIFU method that has been used to facilitate targeted drug delivery and to treat uterine fibroids, essential tremor, and desmoid tumors [8][9][10]. In 2016, Peek et al. evaluated US-guided HIFU for benign tumors based on the change in tumor volume [11]. ...
Article
Full-text available
Objective This report describes the first clinical experience with magnetic resonance imaging-guided focused ultrasound surgery (MRgFUS) using the ExAblate 2100 system for non-invasive breast cancer. Methods Two women with non-invasive breast cancer underwent MRgFUS treatment. One week after the MRgFUS treatment, US-guided vacuum-assisted biopsy was performed for the ablated lesions at the same time as breast-conserving surgery. Results The patients experienced good cosmetic outcomes and did not experience any severe adverse events, such as skin burns. Pathological examination of the surgical specimens revealed a few degenerated intraductal lesions around the breast biopsy markers. Conclusion Performing MRgFUS with the new ExAblate 2100 system appears to be safe and feasible. The histopathological results revealed that adequate ultrasound energy in the appropriate location can induce tumor necrosis.
... FUS in conjunction with pre-formed MBs has been studied for decades as a means to facilitate the delivery of a wide range of therapeutic agents, such as antibodies [9], neurotrophic factors [10], proteins [11], viral vectors [12], stem cells [13], nanoparticles [14], and chemotherapeutics [15], through the BBB. Following a large body of promising pre-clinical work, a number of clinical trials are currently in progress, aiming to efficiently and safely open the BBB in patients with Alzheimer's disease [16], glioblastoma [17], and amyotrophic lateral sclerosis [18]. ...
Article
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 the fractional anisotropy from $0.21 \pm 0.02$ to $0.38 \pm 0.03$ ( $82.6 \pm 5.2\%$ change) within the targeted area one hour after BBB opening. Enhanced DTI contrast overlapped by $77.22 \pm 9.2\%$ with hyper-intense areas of gadolinium-enhanced T1-weighted scans, indicating diffusion anisotropy enhancement only within the BBB opening volume. Diffusion enhancement 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 place of contrast-enhanced T1 mapping for detecting BBB opening during focused-ultrasound treatment or evaluating BBB integrity in brain-related pathologies.
... Ultrasound-based blood-brain barrier disruption (US-BBBD) allows for increased penetration of systemically administered small and large molecule drug therapies into the brain (8-10). Strikingly, US-BBBD alone has been shown to reduce β-amyloid and tau pathologies, stimulate neurogenesis, and improve cognitive performance in mouse AD models (11)(12)(13) and can be further coupled with drug therapies to improve their brain penetration and e cacy (8, 14,15). ...
Preprint
Full-text available
Background: Temporary disruption of the blood-brain barrier (BBB) using pulsed ultrasound leads to the clearance of both amyloid and tau from the brain, increased neurogenesis, and mitigation of cognitive decline in pre-clinical models of Alzheimer’s disease (AD) while also increasing BBB penetration of therapeutic antibodies. The goal of this pilot clinical trial was to investigate the safety and the efficacy of this approach in patients with mild AD using an implantable ultrasound device. Methods: An implantable, 1 MHz ultrasound device (SonoCloud-1) was extradurally implanted under local anesthesia in the skull of 10 mild AD patients to target the left supra-marginal gyrus. Over 3.5 months, seven ultrasound sessions in combination with intravenous infusion of microbubbles were performed twice per month to temporarily disrupt the BBB. ¹⁸F-Florbetapir and ¹⁸F-fluorodeoxyglucose positron emission tomography (PET) imaging were performed on a combined PET/MRI scanner at inclusion and at four and eight months after initiation of sonications to monitor brain metabolism and amyloid levels along with cognitive evaluations. Evolution of cognitive and neuroimaging features were compared to that of a matched sample of control participants. Results: A total of 63 BBB opening procedures were performed in nine subjects. The procedure was well-tolerated. A non-significant decrease in amyloid accumulation at four months of -6.6% (SD=7.2%) on ¹⁸F-Florbetapir PET imaging in the sonicated gray matter targeted by the ultrasound transducer was observed compared to baseline in six subjects that completed treatments and who had evaluable imaging scans. No differences in longitudinal change in glucose metabolism were observed compared to neighboring or contralateral regions or to the change observed in the same region in ADNI participants. No significant effect on cognition evolution was observed in comparison to the ADNI participants as expected due to the small sample size and duration of the trial. Conclusions: These results demonstrate the safety of ultrasound-based BBB disruption and potential of this technology to be used as a therapy for AD patients. They support further research of this technique in a larger clinical trial with a device designed to sonicate larger volumes of tissue and in combination with disease modifying drugs to further enhance the effects observed. Trial registration: NCT03119961 (clinicaltrials.gov)
... An alternative technique for BBB opening is based on systemic injection of ultrasound contrast agents. This method was successfully used to increase the delivery of Herceptin to brain parenchyma in a mice model (Hynynen et al., 2001Kinoshita et al., 2006). ...
Chapter
Blood brain barrier (BBB) is formed by the brain microvascular endothelial cells (BMVECs) lining the wall of brain capillaries. Its integrity is regulated by multiple mechanisms, including up/downregulation of tight junction proteins or adhesion molecules, altered Ca2 + homeostasis, remodeling of cytoskeleton, that are confined at the level of BMVECs. Beside the contribution of BMVECs to BBB permeability changes, other cells, such as pericytes, astrocytes, microglia, leukocytes or neurons, etc. are also exerting direct or indirect modulatory effects on BBB. Alterations in BBB integrity play a key role in multiple brain pathologies, including neurological (e.g. epilepsy) and neurodegenerative disorders (e.g. Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis etc.). In this review, the principal Ca2 + signaling pathways in brain microvascular endothelial cells are discussed and their contribution to BBB integrity is emphasized. Improving the knowledge of Ca2 + homeostasis alterations in BMVECa is fundamental to identify new possible drug targets that diminish/prevent BBB permeabilization in neurological and neurodegenerative disorders.
... Enhanced drug delivery via MB-assisted FUS-BBB opening is widely established. Herceptin (150 kDa) and D4 receptor antibodies (150 kDa) have been successfully delivered into mouse brain (Kinoshita et al., 2006). Methotrexate (545 Da) has also been delivered into normal rat brains in the FUS-assisted model at significantly higher concentrations than in control rats (Mei et al., 2009). ...
Article
Full-text available
Focused Ultrasound (FUS) in combination with gaseous microbubbles has emerged as a potential new means of effective drug delivery to the brain. Recent research has shown that, under burst-type energy exposure with the presence of microbubbles, this modality can transiently permeate the blood-brain barrier (BBB). The bioavailability of therapeutic agents is site-specifically augmented only in the zone where the FUS energy is targeted. The non-invasiveness of this approach makes FUS-induced BBB opening a novel and attractive means to perform localized CNS therapeutic agent delivery. Over the past decade, FUS-BBB opening has been preclinically confirmed to successfully enhance CNS penetration of therapeutic agents including chemotherapeutic agents, therapeutic peptides, monoclonal antibodies, and nanoparticles. Recently, a number of clinical human trials have begun to explore clinical utility. This review article, explores this technology through its physical mechanisms, summarizes the existing preclinical findings (including current medical device designs and technical approaches), and summarizes current ongoing clinical trials.
... With this technique, microbubbles consisting of a lipid or protein shell and a heavy gas core (diameter 1-10 μm) are injected into the bloodstream along with the therapeutic agents of interest. Pulses of ultrasound are then applied to the targeted brain region, driving the microbubbles to oscillate, which ultimately allows the delivery of drugs across the BBB and into the brain [24,25]. In vivo studies have demonstrated that liposomes can be delivered into the brain using this focused ultrasound technique [26][27][28][29][30][31][32][33][34][35][36][37][38][39][40]. ...
Article
Full-text available
Liposomes are clinically used drug carriers designed to improve the delivery of drugs to specific tissues while minimising systemic distribution. However, liposomes are unable cross the blood-brain barrier (BBB) and enter the brain, mostly due to their large size (ca. 100 nm). A noninvasive and localised method of delivering liposomes across the BBB is to intravenously inject microbubbles and apply long pulses of ultrasound (pulse length: >1 ms) to a targeted brain region. Recently, we have shown that applying rapid short pulses (RaSP) (pulse length: 5 μs) can deliver drugs with an improved efficacy and safety profile. However, this was tested with a relatively smaller 3-kDa molecule (dextran). In this study, we examine whether RaSP can deliver liposomes to the murine brain in vivo. Fluorescent DiD-PEGylated liposomes were synthesized and injected intravenously alongside microbubbles. The left hippocampus of mice was then sonicated with either a RaSP sequence (5 μs at 1.25 kHz in groups of 10 ms at 0.5 Hz) or a long pulse sequence (10 ms at 0.5 Hz), with each pulse having a 1-MHz centre frequency (0.35 and 0.53 MPa). The delivery and distribution of the fluorescently-labelled liposomes were assessed by fluorescence imaging of the brain sections. The safety profile of the sonicated brains was assessed by histological staining. RaSP was shown to locally deliver liposomes across the BBB at 0.53 MPa with a more diffused and safer profile compared to the long pulse ultrasound sequence. Cellular uptake of liposomes was observed in neurons and microglia, while no uptake within astrocytes was observed in both RaSP and long pulse-treated brains. This study shows that RaSP allows a targeted and safe delivery of liposomal drugs into the murine brain with potential to deliver drugs into neuronal and glial targets.
... Especially intriguing is the ability to deliver neural stem cells (125) or natural killer cells (69), which may provide a platform for a variety of therapeutic approaches. Other therapeutic agents delivered include antiamyloid beta antibodies (126), antidopamine receptor D4 antibodies (127), Herceptin (128), interleukin-12 (129), erythropoietin (130), and brain-derived neurotrophic factor (131). In addition to characterizing the delivery and efficacy of FUS+MB-mediated therapeutic agent delivery, further work in the field should focus on evaluating the movement of substances that have crossed the BBB. ...
Article
Specialized features of vasculature in the central nervous system greatly limit therapeutic treatment options for many neuropathologies. Focused ultrasound, in combination with circulating microbubbles, can be used to transiently and noninvasively increase cerebrovascular permeability with a high level of spatial precision. For minutes to hours following sonication, drugs can be administered systemically to extravasate in the targeted brain regions and exert a therapeutic effect, after which permeability returns to baseline levels. With the wide range of therapeutic agents that can be delivered using this approach and the growing clinical need, focused ultrasound and microbubble (FUS+MB) exposure in the brain has entered human testing to assess safety. This review outlines the use of FUS+MB-mediated cerebrovascular permeability enhancement as a drug delivery technique, details several technical and biological considerations of this approach, summarizes results from the clinical trials conducted to date, and discusses the future direction of the field. Expected final online publication date for the Annual Review of Biomedical Engineering, Volume 23 is June 2021. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.
... This strategy allows drugs to flow directly from the systemic circulation into the brain by BBB disruption using different methodologies. The most important are: (1) osmotic disruption, due to the administration of hypertonic solutions (e.g., mannitol) causing cells shrinking based on cell dehydration [37]; (2) ultrasound methods, which rely on transcranial delivery of low-frequency ultrasound waves resulting in the opening of tight junctions [38]; and (3) pharmacological agents, such as bradykinin-like compounds (e.g., histamine, bradykinin) that disrupt tight junctions by stimulating B 2 receptors presented in endothelial cells and transiently increasing cytosolic Ca 2+ [39]. The costs, anesthetic administration, and hospitalization are significant drawbacks for all these approaches. ...
Article
Full-text available
The incidence of brain metastases (BM) in cancer patients is increasing. After diagnosis, overall survival (OS) is poor, elicited by the lack of an effective treatment. Monoclonal antibody (mAb)-based therapy has achieved remarkable success in treating both hematologic and non-central-nervous system (CNS) tumors due to their inherent targeting specificity. However, the use of mAbs in the treatment of CNS tumors is restricted by the blood-brain barrier (BBB) that hinders the delivery of either small-molecules drugs (sMDs) or therapeutic proteins (TPs). To overcome this limitation, active research is focused on the development of strategies to deliver TPs and increase their concentration in the brain. Yet, their molecular weight and hydrophilic nature turn this task into a challenge. The use of BBB peptide shuttles is an elegant strategy. They explore either receptor-mediated transcytosis (RMT) or adsorptive-mediated transcytosis (AMT) to cross the BBB. The latter is preferable since it avoids enzymatic degradation, receptor saturation, and competition with natural receptor substrates, which reduces adverse events. Therefore, the combination of mAbs properties (e.g., selectivity and long half-life) with BBB peptide shuttles (e.g., BBB translocation and delivery into the brain) turns the therapeutic conjugate in a valid approach to safely overcome the BBB and efficiently eliminate metastatic brain cells.
... Focused ultrasound techniques concentrate acoustic energy on a focal spot in the brain measuring a few millimeters in diameter and, combined with magnetic resonance imaging (MRI), have been used in the clinic for thermal coagulation of tumors in human patients with real-time monitoring [44,45]. MRI-guided focused ultrasound (MRgFUS) used to induce cavitation of intravenously (IV) administered microbubbles has been reported to reliably cause temporary physical disruption of the BBB in small animal models and has been used for delivery of BBB-impermeably compounds such as antibody drugs [46,47]. Airan et al. delivered nanoemulsions encapsulating a BBB-permeable small molecule anesthetic using MRgFUS to enable potent intracranial drug delivery with high spatiotemporal control in an acute rate seizure model without brain parenchymal damage [48], demonstrating the fine level of control that can be exerted with this neuromodulatory technique. ...
Article
Glioblastoma is one of the deadliest forms of primary adult tumors, with median survival of 14.6 months post-diagnosis despite aggressive standard of care treatment. This grim prognosis for glioblastoma patients has changed little in the past two decades, necessitating novel treatment modalities. One potential treatment modality is cancer immunotherapy, which has shown remarkable progress in slowing disease progression or even potentially curing certain solid tumors. However, the transport barriers posed by the blood–brain barrier and the immune privileged status of the central nervous system pose drug delivery obstacles that are unique to brain tumors. In this review, we provide an overview of the various physiological, immunological, and drug delivery barriers that must be overcome for effective glioblastoma treatment. We discuss chemical modification strategies to enable nanomedicines to bypass the blood–brain barrier and reach intracranial tumors. Finally, we highlight recent advances in biomaterial-based strategies for cancer immunotherapy that can be adapted to glioblastoma treatment.
... Focused ultrasound (FUS)-mediated disruption of the BBB is an example of a therapeutic strategy that could be influenced by such findings, given that this application relies on the presence of intravascular MBs. FUS BBB opening is rapidly advancing as a non-invasive, safe, and repeatable strategy 26 to treat a wide variety of neurological pathologies [25][26][27][28][29][30][31][32][33][34][35][36][37][38] . ...
Article
Full-text available
Abstract Microbubbles (MB) are widely used as contrast agents to perform contrast-enhanced ultrasound (CEUS) imaging and as acoustic amplifiers of mechanical bioeffects incited by therapeutic-level ultrasound. The distribution of MBs in the brain is not yet fully understood, thereby limiting intra-operative CEUS guidance or MB-based FUS treatments. In this paper we describe a robust platform for quantification of MB distribution in the human brain, allowing to quantitatively discriminate between tumoral and normal brain tissues and we provide new information regarding real-time cerebral MBs distribution. Intraoperative CEUS imaging was performed during surgical tumor resection using an ultrasound machine (MyLab Twice, Esaote, Italy) equipped with a multifrequency (3–11 MHz) linear array probe (LA332) and a specific low mechanical index (MI
... Une étude d'ouverture de la BHE pour le traitement de la sclérose amyotrophique latérale est aussi menée à Toronto (NCT03321487). Dans le traitement de la maladie de Parkinson et de la schizophrénie, l'utilisation d'un anticorps anti-domaine extracellulaire du récepteur à la dopamine D4 a montré une augmentation de pénétration cérébrale après ouverture de la BHE par US (Kinoshita et al., 2006c D'autre part, il a été montré que ABCE1 était un régulateur de la prolifération V / C 274 et de la migration de cellules tumorales en régulant la dynamique du cytosquelette d'actine (Yu et al., 2017). Cette fonction lui confère un rôle de point de contrôle de l'entrée en mitose dans le cycle cellulaire (Jiao et al., 2017). ...
Thesis
Le glioblastome (GBM) est le cancer du cerveau le plus fréquent et grave chez l’adulte. Son pronostic sombre est en partie dû à la résistance de ces tumeurs aux chimiothérapies. L’une des principales causes de ces résistances est l’incapacité des chimiothérapies à pénétrer dans le cerveau depuis le sang à cause de la barrière hémato-encéphalique (BHE), une spécificité des vaisseaux sanguins cérébraux. Par l’injection de microbulles et l’envoi d’ultrasons dans le cerveau, il est possible d’ouvrir cette BHE pour permettre à des chimiothérapies d’entrer dans le cerveau. Nous avons montré quela chimiothérapie carboplatine gagnait en efficacité lorsqu’elle était injectée après ce procédé. Les GBM peuvent aussi montrer une résistance aux chimiothérapies par des mécanismes génétiques intrinsèques à la tumeur. Nous avons étudié l’expression et l’impact sur le pronostique des patients atteints de GBM des gènes de la famille des transporteurs ABC, dont le membre ABCA13 s’est avéré important.
... The delivery of large molecular drugs across the BBB has also been demonstrated, including IL-12 (70 kDa) [67], dopamine D-4 receptor-targeting antibody [68], and HER2/c-erbB2 (humanized antihuman epidermal growth factor receptor 2) monoclonal antibody [69]. We have demonstrated the enhanced delivery of bevacizumab (149 kDa) in a mouse glioma model [45]. ...
Article
Full-text available
The blood–brain barrier (BBB) protects the central nervous system (CNS) from invasive pathogens and maintains the homeostasis of the brain. Penetrating the BBB has been a major challenge in the delivery of therapeutic agents for treating CNS diseases. Through a physical acoustic cavitation effect, focused ultrasound (FUS) combined with microbubbles achieves the local detachment of tight junctions of capillary endothelial cells without inducing neuronal damage. The bioavailability of therapeutic agents is increased only in the area targeted by FUS energy. FUS with circulating microbubbles is currently the only method for inducing precise, transient, reversible, and noninvasive BBB opening (BBBO). Over the past decade, FUS-induced BBBO (FUS-BBBO) has been preclinically confirmed to not only enhance the penetration of therapeutic agents in the CNS, but also modulate focal immunity and neuronal activity. Several recent clinical human trials have demonstrated both the feasibility and potential advantages of using FUS-BBBO in diseased patients. The promising results support adding FUS-BBBO as a multimodal therapeutic strategy in modern CNS disease management. This review article explores this technology by describing its physical mechanisms and the preclinical findings, including biological effects, therapeutic concepts, and translational design of human medical devices, and summarizes completed and ongoing clinical trials.
... The BBB is very challenging for the clinical diagnosis and treatment of brain tumors. To overcome this problem, focused ultrasound sonication (FUS) with drug-loaded microbubble, opens the rigid protective barrier of the brain and allows the penetration of therapeutics agent across the BBB into the brain [112,113]. The focused ultrasonic waves break the microbubbles in nanosized, which is more effective in the therapeutic delivery of medicine (Fig. 14). ...
Article
Objective: We previously developed short burst, phase keying (SBPK) focused ultrasound (FUS) to mitigate standing waves in the human vertebral canal. Here, we show microbubble emissions from these pulses can be detected through the human vertebral arch and that these pulses are effective for blood-spinal cord barrier (BSCB) opening. Methods: At $\text{f}_0$ =514kHz, circulating microbubbles were sonicated through ex vivo human vertebrae(60kPa-1MPa) using a dual-aperture approach and SBPK exposures engineered to incorporate pulse inversion (PI). Signals from a 250kHz receiver were analyzed using PI, short-time Fourier analysis and the maximum projection over the pulse train. In rats (n=14), SBPK FUS+microbubbles was applied to 3 locations/spinal cord at fixed pressures (~0.20-0.47MPa). MRI and histology were used to assess opening and tissue damage. Results: In human vertebrae between 0.2-0.4MPa, PI amplified the microbubble/baseline ratio at $\text{f}_0$ /2 and 2 $\text{f}_0$ by 202±40%(132-291%). This was maximal at 0.4MPa, coinciding with the onset of broadband emissions. In vivo, opening was achieved at 40/42 locations, with mean MRI enhancement of 46±32%(16%-178%). Using PI, $\text{f}_0$ /2 was detected at 14/40 opening locations. At the highest pressures ( $\text{f}_0$ /2 present) histology showed widespread bleeding throughout the focal region. At the lowest pressures, opening was achieved without bleeding. Conclusion: This study confirmed that PI can increase sensitivity to transvertebral detection of microbubble signals. Preliminary in vivo investigations show that SBPK FUS can increase BSCB permeability without tissue damage. Significance: SBPK is a clinically relevant pulse scheme and, in combination with PI, provides a means of mediating and monitoring BSCB opening noninvasively.
Article
Focused ultrasound (FUS), in combination with microbubble contrast agents, can be used to transiently open the blood-brain barrier (BBB) to allow intravascular agents to cross into the brain. Often, FUS is carried out in conjunction with magnetic resonance imaging (MRI) to evaluate BBB opening to gadolinium-based MRI contrast agents. Although MRI allows direct visualization of the distribution of gadolinium-based contrast agents in the brain parenchyma, it does not allow measurements of the distribution of other molecules crossing the BBB. Therapeutic molecules (e.g., monoclonal antibodies) are much different in size than MRI contrast agents and have been found to have different distributions in the brain after FUS-mediated BBB opening. In the work described here, we combined in vivo MRI and ex vivo multispectral fluorescence imaging to compare the distributions of MRI contrast and dextran molecules of different molecular weights (3, 70 and 500 kDa) after FUS-mediated BBB opening through a range of ultrasound pressures (0.18-0.46 MPa) in laboratory mice. The volume of brain exposed was calculated from the MRI and fluorescence images and was significantly dependent on both molecular weight and ultrasound pressure. Diffusion coefficients of the different-molecular-weight dextran molecules in the brain parenchyma were also calculated from the fluorescence images and were negatively correlated with the molecular weight of the dextran molecules. The results of this work build on a body of knowledge that is critically important for the FUS technique to be used in clinical delivery of therapeutics to the brain.
Article
Background and Purpose: The present study aimed to explore the feasibility and efficacy of the targeted non-invasive implantation of MSCs by low-intensity ultrasound-targeted microbubble destruction (LI-UTMD) assisted blood brain barrier opening and its improvement on neurobehavioral outcomes in brain ischemic rats. Methods: A transcranial irradiation of low-intensity ultrasound by diagnostic devices was performed, and lipid microbubbles and MSCs were simultaneously infused. Then, the MSC transmigration from brain vessels to parenchyma was demonstrated, and MSCs were statistically analyzed on day 1, 4, 7 and 14. Behavioral function was statistically analyzed. Results: The extra-vascular leakage of lanthanum and EB was observed at the brain ischemic area receiving ultrasound. MSCs were observed at the ultrasound irradiated brain hemisphere, and the number of MSCs in LI-UTMD assisted MSCs group was significantly higher than that in the MSCs group (P < 0.01). The attachment, traversing and trans-migration of MSCs across the blood brain barrier were recorded. Neuro-behavioral function improved with this approach. Conclusion: The transcranial irradiation approach of low intensity ultrasound targeted microbubble destruction on brain ischemic rats might be a safe and efficient blood brain barrier opening approach to prompt the successful delivery of MSCs into the targeted area of brain ischemia, and ameliorate neurological function
Chapter
As a well-established method for clinical imaging, ultrasound can also be used as source of focused waves to selectively affect tissues by activating drug carriers and/or improving drug uptake in tumors providing a novel strategy for targeted treatment [1]. In this chapter we describe the design of nanoparticles that can respond to effects induced by focused ultrasound that can be used in cancer therapy.
Article
The blood-brain barrier (BBB) prevents many drugs from entering the brain. Yet, conventional methods that open the BBB are technically demanding, poorly reversible, and can be associated with long-term adverse effects. In comparison, carbogen, which is introduced nearly a century ago as a treatment for psychiatric disorders, is easy to administer and readily available to many labs and hospitals. Here, we show that carbogen inhalation opened the BBB in rats, as indicated by the extravasation of an intravenous protein tracer. When the tracer was injected immediately or hours after carbogen inhalation, less tracer was detected in the rat brains, suggesting at least partial reversibility of this response after carbogen exhalation. Despite marked increase in BBB permeability, inhalation of carbogen for 30-90 min had no acute effect on the level of neuroinflammation or apoptosis in the brain, and had no long-term effect on body weight, food intake, locomotor activity, or learning and memory performance. Our study demonstrated that carbogen inhalation is a safe method to open the BBB.
Article
Full-text available
Magnetic resonance-guided focused ultrasound (MRgFUS) is an emerging new technology with considerable potential to treat various neurological diseases. With refinement of ultrasound transducer technology and integration with magnetic resonance imaging guidance, transcranial sonication of precise cerebral targets has become a therapeutic option. Intensity is a key determinant of ultrasound effects. High-intensity focused ultrasound can produce targeted lesions via thermal ablation of tissue. MRgFUS-mediated stereotactic ablation is non-invasive, incision-free, and confers immediate therapeutic effects. Since the US Food and Drug Administration approval of MRgFUS in 2016 for unilateral thalamotomy in medication-refractory essential tremor, studies on novel indications such as Parkinson's disease, psychiatric disease, and brain tumors are underway. MRgFUS is also used in the context of blood-brain barrier (BBB) opening at low intensities, in combination with intravenously-administered microbubbles. Preclinical studies show that MRgFUS-mediated BBB opening safely enhances the delivery of targeted chemotherapeutic agents to the brain and improves tumor control as well as survival. In addition, BBB opening has been shown to activate the innate immune system in animal models of Alzheimer's disease. Amyloid plaque clearance and promotion of neurogenesis in these studies suggest that MRgFUS-mediated BBB opening may be a new paradigm for neurodegenerative disease treatment in the future. Here, we review the current status of preclinical and clinical trials of MRgFUS-mediated thermal ablation and BBB opening, described their mechanisms of action, and discuss future prospects.
Thesis
Glioblastoma (GBM) is the most frequent and the most aggressive primary brain cancer in adults. The current standard of care of newly diagnosed GBM includes maximal safe surgery followed by concurrent temozolomide (TMZ) and radiotherapy followed by adjuvant TMZ. Despite this intensive therapeutic regimen, the median overall survival is below 18 months. One of the main limitations of current anti-tumor treatments is the blood-brain barrier and the local immunosuppressed microenvironment in the setting of brain cancers.This thesis is focused on overcoming the blood-brain barrier to modulate local immunity to better anti-tumor treatments. Firstly, we reviewed the role of ABC proteins superfamily members expressed in the BBB cells and their role in GBM cells' resistance to chemotherapy. Secondly, we have studied the expression of immune checkpoint proteins CD80 and CD86 and their impacts on outcome in newly diagnosed glioblastoma patients treated with the standard of care. CD86 overexpression is associated with a better prognosis. Finally, we have shown dramatic efficacy of immune checkpoint inhibitors targeting PLD1 when combined with ultrasound-mediated BBB opening in glioblastoma-bearing mice compared to anti-PDL1 alone. BBB and immunity modulations are promising strategies to improve our therapeutic arsenal against glioblastoma.
Article
Physically triggered systems hold promise for improving drug delivery by enhancing the controllability of drug accumulation and release, lowering non-specific toxicity, and facilitating clinical translation. Several external physical stimuli including ultrasound, light, electric fields and magnetic fields have been used to control drug delivery and they share some common features such as spatial targeting, spatiotemporal control, and minimal invasiveness. At the same time, they possess several distinctive features in terms of interactions with biological entities and/or the extent of stimulus response. Here, we review the key advances of such systems with a focus on discussing their physical mechanisms and the design rationales.
Article
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A long-standing goal of translational neuroscience is the ability to noninvasively deliver therapeutic agents to specific brain regions with high spatiotemporal resolution. Focused ultrasound (FUS) is an emerging technology that can noninvasively deliver energy up the order of 1 kW/cm2 with millimeter and millisecond resolution to any point in the human brain with Food and Drug Administration-approved hardware. Although FUS is clinically utilized primarily for focal ablation in conditions such as essential tremor, recent breakthroughs have enabled the use of FUS for drug delivery at lower intensities (i.e., tens of watts per square centimeter) without ablation of the tissue. In this review, we present strategies for image-guided FUS-mediated pharmacologic neurointerventions. First, we discuss blood–brain barrier opening to deliver therapeutic agents of a variety of sizes to the central nervous system. We then describe the use of ultrasound-sensitive nanoparticles to noninvasively deliver small molecules to millimeter-sized structures including superficial cortical regions and deep gray matter regions within the brain without the need for blood–brain barrier opening. We also consider the safety and potential complications of these techniques, with attention to temporal acuity. Finally, we close with a discussion of different methods for mapping the ultrasound field within the brain and describe future avenues of research in ultrasound-targeted drug therapies.
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Focused ultrasound and microbubble (FUS + MB)-mediated blood–brain barrier (BBB) permeability enhancement can facilitate targeted brain-drug delivery. While controlling the magnitude of BBB permeability enhancement is necessary to limit tissue damage, little work has attempted to decouple these concepts. This work investigated the relationship between BBB permeability enhancement and the relative transcription of inflammatory mediators 4 h following sonication. Three microbubble formulations, Definity, BG8774, and MSB4, were compared, with the dose of each formulation normalized to gas volume. While changes in the transcription of key proinflammatory mediators, such as Il1b, Ccl2, and Tnf, were correlated to the magnitude of BBB permeability enhancement, these correlations were not independent of microbubble formulation; microbubble size distribution may play an important role, as linear regression analyses of BBB permeability magnitude versus differential gene expression for these proinflammatory mediators revealed significantly greater slopes for MSB4, a monodisperse microbubble with mean diameter of 4 μm, compared to Definity or BG8774, both polydisperse microbubbles with mean diameters below 2 μm. Additionally, the function of an acoustic feedback control algorithm, based on the detection threshold of ultraharmonic emissions, was assessed. While this control strategy was effective in limiting both wideband emissions and red blood cell extravasation, microbubble formulation was found to influence the magnitude of BBB leakage and correlations to acoustic emissions. This work demonstrates that while the initial magnitude of FUS + MB-mediated BBB permeability enhancement has a clear influence on the subsequent inflammatory responses, microbubble characteristics influence these relationships and must also be considered.
Chapter
Although great progress has been made in recent years and more than 7000 small-molecule drugs are available, few effective treatments and no cures of the central nervous system (CNS) diseases are currently available. This is mainly due to the impermeability of the blood-brain barrier (BBB) that allows only 5% of those drugs to diffuse to the brain parenchyma thereby allowing treatment of only a tiny fraction of these diseases. Safe and localized opening of the BBB has been proven to pose an equally significant challenge. Focused ultrasound (FUS), in conjunction with microbubbles, remains the sole technique that can induce localized BBB opening noninvasively. In this chapter, we demonstrate how the microbubble diameter and peak negative pressure can be optimized in order to dictate the BBB opening volume and permeability in small and large animals. We subsequently demonstrate that neuroprotection and neurorestoration in the dopaminergic neurons in the nigrostriatal pathway at the early stages of Parkinson’s disease as well as amyloid and tau reduction at the early stages of Alzheimer’s disease can be achieved at therapeutic levels safely.
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Phagocytic immunotherapies such as CD47 blockade have emerged as promising strategies for glioblastoma (GB) therapy, but the blood brain/tumor barriers (BBB/BTB) pose a persistent challenge for mCD47 delivery that can be overcome by focused ultrasound (FUS)-mediated BBB/BTB disruption. We here leverage immuno-PET imaging to determine how timing of [⁸⁹Zr]-mCD47 injection relative to FUS impacts antibody penetrance into orthotopic murine gliomas. We then design and implement a rational paradigm for combining FUS and mCD47 for glioma therapy. We demonstrate that timing of antibody injection relative to FUS BBB/BTB disruption is a critical determinant of mCD47 access, with post-FUS injection conferring superlative antibody delivery to gliomas. We also show that mCD47 delivery across the BBB/BTB with repeat sessions of FUS can significantly constrain tumor outgrowth and extend survival in glioma-bearing mice. This study generates provocative insights for ongoing pre-clinical and clinical evaluations of FUS-mediated antibody delivery to brain tumors. Moreover, our results confirm that mCD47 delivery with FUS is a promising therapeutic strategy for GB therapy.
Chapter
Extracranial ultrasound assessment of the cervical arteries provides real-time, bedside evaluation of blood flow and vessel walls. Ultrasound examination is a non-invasive imaging modality of atherosclerosis in the extracranial arteries. For better yield of microembolic signals, power motion Doppler is superior to single-channel transcranial Doppler ultrasonography (TCD). TCD yields good-to-excellent results in detecting steno-occlusive intracranial lesions in the setting of acute cerebral ischemia when compared against computed tomographic angiography (CTA) or magnetic resonance angiography (MRA). Waveform pattern yields more information about thrombus location, hemodynamic significance of occlusion, and resistance in the distal vessels compared to velocity difference by itself. TCD is more sensitive than transthoracic echocardiogram for right-to-left shunt detection. Despite initial promising results of sonothrombolysis, the efficacy of ultrasound enhancement of thrombolysis has not been proven. Novel applications of ultrasound are molecular imaging, opening of the blood-brain barrier, and targeted drug delivery.
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Background Focused ultrasound combined with microbubbles has been used in clinical studies for blood-brain barrier (BBB) opening in conjunction with MRI. However, the impact of the static magnetic field generated by an MRI scanner on the BBB opening outcome has not been evaluated. Purpose To determine the relationship of the static magnetic field of an MRI scanner on focused ultrasound combined with microbubble-induced BBB opening. Materials and Methods Thirty wild-type mice were divided into four groups. Mice from different groups were sonicated with focused ultrasound in different static magnetic fields (approximately 0, 1.5, 3.0, and 4.7 T), with all other experimental parameters kept the same. Focused ultrasound sonication was performed after intravenous injection of microbubbles. Microbubble cavitation activity, the fundamental -physical mechanism underlying focused ultrasound BBB opening, was monitored with passive cavitation detection. After sonication, contrast-enhanced T1-weighted MRI was performed to assess BBB opening outcome. Intravenously injected Evans blue was used as a model agent to evaluate trans-BBB delivery efficiency. Results The microbubble cavitation dose decreased by an average of 2.1 dB at 1.5 T (P = .05), 2.9 dB at 3.0 T (P = .01), and 3.0 dB at 4.7 T (P = .01) compared with that outside the magnetic field (approximately 0 T). The static magnetic field of an MRI scanner decreased BBB opening volume in mice by 3.2-fold at 1.5 T (P < .001), 4.5-fold at 3.0 T (P < .001), and 11.6-fold at 4.7 T (P <.001) compared with mice treated outside the magnetic field. It also decreased Evans blue trans-BBB delivery 1.4-fold at 1.5 T (P = .009), 1.6-fold at 3.0 T (P < .001), and 1.9-fold at 4.7 T (P < .001). Conclusion Static magnetic fields dampened microbubble cavitation activity and decreased trans-blood-brain barrier (BBB) delivery by focused ultrasound combined with microbubble-induced BBB opening. © RSNA, 2021.
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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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Brain tumours have a poor prognosis and lack effective treatments. The blood–brain barrier (BBB) represents a major hurdle to drug delivery to brain tumours. In some locations in the tumour, the BBB may be disrupted to form the blood–brain tumour barrier (BBTB). This leaky BBTB enables diagnosis of brain tumours by contrast enhanced magnetic resonance imaging; however, this disruption is heterogeneous throughout the tumour. Thus, relying on the disrupted BBTB for achieving effective drug concentrations in brain tumours has met with little clinical success. Because of this, it would be beneficial to design drugs and drug delivery strategies to overcome the ‘normal’ BBB to effectively treat the brain tumours. In this review, we discuss the role of BBB/BBTB in brain tumour diagnosis and treatment highlighting the heterogeneity of the BBTB. We also discuss various strategies to improve drug delivery across the BBB/BBTB to treat both primary and metastatic brain tumours. Recognizing that the BBB represents a critical determinant of drug efficacy in central nervous system tumours will allow a more rapid translation from basic science to clinical application. A more complete understanding of the factors, such as BBB–limited drug delivery, that have hindered progress in treating both primary and metastatic brain tumours, is necessary to develop more effective therapies. Abstract
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Survival outcomes for patients with glioblastoma multiforme (GBM) have remained poor for the past 15 years, reflecting a clear challenge in the development of more effective treatment strategies. The efficacy of systemic therapies for GBM is greatly limited by the presence of the blood-brain barrier (BBB), which prevents drug penetration and accumulation in regions of infiltrative tumour, as represented in a consistent portion of GBM lesions. Focused ultrasound (FUS) – a technique that uses low-frequency ultrasound waves to induce targeted temporary disruption of the BBB – promises to improve survival outcomes by enhancing drug delivery and accumulation to infiltrating tumour regions. In this review we discuss the current state of preclinical investigations using FUS to enhance delivery of systemic therapies to intracranial neoplasms. We highlight critical methodological inconsistencies that are hampering clinical translation of FUS and we provide guiding principles for future preclinical studies. Particularly, we focus our attention on the importance of the selection of clinically relevant animal models and to the standardization of methods for FUS delivery, which will be paramount to the successful clinical translation of this promising technology for treatment in GBM patients. We also discuss how preclinical FUS research can benefit the development of GBM immunotherapies.
Chapter
The tightly regulated permeance of the blood-brain barrier (BBB) greatly limits the range of therapeutic treatment options for central nervous system (CNS) diseases. The use of focused ultrasound (FUS), in conjunction with circulating microbubbles, is a unique approach whereby the transcranial application of acoustic energy, focused within targeted brain areas, can be used to induce a noninvasive, transient, and targeted increase in BBB permeability. This can provide an avenue for the delivery of therapeutic agents from the systemic circulation into the brain. While this approach continues to show great promise and has entered clinical testing, there remains a need for preclinical research to investigate the long-term effects of single and repeated FUS treatment on cerebrovascular health and neurological function, as well the pharmacokinetics of specific drugs following FUS. Additionally, there is a need for improved monitoring strategies that can precisely predict resulting bio-effects. This will allow the continued development of control algorithms that can further increase the safety profile of FUS. Here we will describe two approaches to study FUS-mediated increases in BBB permeability in rodent models: MRI-guided FUS and in vivo two-photon fluorescence microscopy FUS experiments. The goal of this chapter is to outline each procedure, present options for experimental design, and highlight important considerations for the collection and interpretation of data.
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To test the feasibility of noninvasive magnetic resonance (MR) imaging-guided focused ultrasound surgery (FUS) of benign fibroadenomas in the breast. Eleven fibroadenomas in nine patients under local anesthesia were treated with MR imaging-guided FUS. Based on a T2-weighted definition of target volumes, sequential sonications were delivered to treat the entire target. Temperature-sensitive phase-difference-based MR imaging was performed during each sonication to monitor focus localization and tissue temperature changes. After the procedure, T2-weighted and contrast material-enhanced T1-weighted MR imaging were performed to evaluate immediate and long-term effects. Thermal imaging sequences were improved over the treatment period, with 82% (279 of 342) of the hot spots visible in the last seven treatments. The MR imager was used to measure temperature elevation (12.8 degrees -49.9 degrees C) from these treatments. Eight of the 11 lesions treated demonstrated complete or partial lack of contrast material uptake on posttherapy T1-weighted images. Three lesions showed no marked decrease of contrast material uptake. This lack of effective treatment was most likely due to a lower acoustic power and/or patient movement that caused misregistration. No adverse effects were detected, except for one case of transient edema in the pectoralis muscle 2 days after therapy. MR imaging-guided FUS can be performed to noninvasively coagulate benign breast fibroadenomas.
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To determine if focused ultrasound beams can be used to locally open the blood-brain barrier without damage to surrounding brain tissue and if magnetic resonance (MR) imaging can be used to monitor this procedure. The brains of 18 rabbits were sonicated (pulsed sonication) in four to six locations, with temporal peak acoustic power ranging from 0.2 to 11.5 W. Prior to each sonication, a bolus of ultrasonographic (US) contrast agent was injected into the ear vein of the rabbit. A series of fast or spoiled gradient-echo MR images were obtained during the sonications to monitor the temperature elevation and potential tissue changes. Contrast material-enhanced MR images obtained minutes after sonications and repeated 1-48 hours later were used to depict blood-brain barrier opening. Whole brain histologic evaluation was performed. Opening of the blood-brain barrier was confirmed with detection of MR imaging contrast agent at the targeted locations. The lowest power levels used produced blood-brain barrier opening without damage to the surrounding neurons. Contrast enhancement correlated with the focal signal intensity changes in the magnitude fast spoiled gradient-echo MR images. The blood-brain barrier can be consistently opened with focused ultrasound exposures in the presence of a US contrast agent. MR imaging signal intensity changes may be useful in the detection of blood-brain barrier opening during sonication.
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Getting drugs and genes into the brain is a tall order. This is because the presence of the blood-brain barrier prevents many molecules from crossing into the brain. Overcoming this problem will have a profound effect on the treatment of many neurological disorders, allowing larger water-soluble molecules to pass into the brain. Transport vectors, such as endogenous peptides, modified proteins or peptidomimetic monoclonal antibodies, are one way of tricking the brain into allowing these molecules to pass. This article will review such molecular Trojan Horses, and the progress that has been made in the delivery of drugs and genes to the brain.
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Brain drug development of either small molecule or large molecule (recombinant proteins, gene medicines) neurotherapeutics has been limited, owing to the restrictive transport properties of the brain microvasculature, which forms the blood-brain barrier (BBB) in vivo. Widespread drug delivery to the brain, while not feasible via craniotomy and intracerebral injection, is possible if the drug is delivered to brain via the transvascular route through the BBB. Novel brain drug delivery and drug targeting strategies can be developed from an understanding of the molecular and cellular biology of the brain microvascular and BBB transport processes.
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The feasibility and safety of magnetic resonance (MR) imaging-guided focused ultrasound surgery for uterine leiomyomas is reported. Sequential sonications were delivered to nine targets. Temperature-sensitive phase-difference MR imaging monitored the location of the focus and measured tissue temperature elevations, ensuring therapeutic dose. MR images and hysterectomy specimens were evaluated. Six leiomyomas received full therapeutic doses, and 98.5% of the sonications were visualized. MR thermometry was successful in all sonications and cases. Focal necrotic lesions were seen in all cases at MR, and five were pathologically confirmed. MR imaging-guided focused ultrasound causes thermocoagulation and necrosis in uterine leiomyomas and is feasible and safe, without serious consequences.
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Blood-brain barrier (BBB) genomics enables the rapid discovery of novel transporters that are expressed at the brain capillary endothelium. The BBB transporters are potential conduits to the brain that therapeutic drugs may use to gain passage across the BBB. Due to the small volume of brain occupied by the endothelium (10(-3) parts of the brain), it is necessary to build a BBB genomics program that is separate from a whole-brain genomics analysis. It is estimated that approximately 15% of all genes selectively expressed at the BBB encode for transporter proteins, and that only approximately 50% of BBB transporters are currently known. The development of a BBB genomics program and the discovery of novel BBB transporters could lead to the invention of new approaches to solving the BBB drug delivery problem.
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One of the most clinically advanced forms of experimental disease-modifying treatment for Alzheimer disease is immunization against the amyloid beta protein (Abeta), but how this may prevent cognitive impairment is unclear. We hypothesized that antibodies to Abeta could exert a beneficial action by directly neutralizing potentially synaptotoxic soluble Abeta species in the brain. Intracerebroventricular injection of naturally secreted human Abeta inhibited long-term potentiation (LTP), a correlate of learning and memory, in rat hippocampus in vivo but a monoclonal antibody to Abeta completely prevented the inhibition of LTP when injected after Abeta. Size fractionation showed that Abeta oligomers, not monomers or fibrils, were responsible for inhibiting LTP, and an Abeta antibody again prevented such inhibition. Active immunization against Abeta was partially effective, and the effects correlated positively with levels of antibodies to Abeta oligomers. The ability of exogenous and endogenous antibodies to rapidly neutralize soluble Abeta oligomers that disrupt synaptic plasticity in vivo suggests that treatment with such antibodies might show reversible cognitive deficits in early Alzheimer disease.
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Most lymphomas that involve the central nervous system are B-cell neoplasms that express the cell surface molecule CD20. After intravenous administration, rituximab can be reproducibly measured in the cerebrospinal fluid (CSF) in patients with primary central nervous system lymphoma; however, the CSF levels of rituximab are approximately 0.1% of serum levels associated with therapeutic activity in patients with systemic non-Hodgkin lymphoma. Because lymphomatous meningitis is a frequent complication of non-Hodgkin lymphoma, we have conducted an analysis of the safety and pharmacokinetics of direct intrathecal administration of rituximab using cynomolgus monkeys. No significant acute or delayed toxicity, neurologic or otherwise, was detected. Pharmacokinetic analysis suggests that drug clearance from the CSF is biphasic, with a terminal half-life of 4.96 hours. A phase 1 study to investigate the safety and pharmacokinetics of intrathecal rituximab in patients with recurrent lymphomatous meningitis will be implemented based on these findings.
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Delivery of therapeutic proteins into tissues and across the blood-brain barrier is severely limited by the size and biochemical properties of the proteins. Here it is shown that intraperitoneal injection of the 120-kilodalton β-galactosidase protein, fused to the protein transduction domain from the human immunodeficiency virus TAT protein, results in delivery of the biologically active fusion protein to all tissues in mice, including the brain. These results open new possibilities for direct delivery of proteins into patients in the context of protein therapy, as well as for epigenetic experimentation with model organisms.
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The aim of this study was to determine the safety and efficacy of intraarterial chemotherapy with osmotic opening of the blood-brain barrier (BBB) for the treatment of malignant brain tumors when administered across multiple centers. Patients with primary central nervous system lymphoma (PCNSL), primitive neuroectodermal tumor (PNET), germ cell tumor, cancer metastasis to the brain, or low or high grade glioma were eligible. Prior to entry, magnetic resonance imaging or computed tomography brain scan, medical history, neurologic status, and Karnofsky performance status were reviewed at the coordinating center. Standardized anesthesia and intraarterial catheterization guidelines were followed by a multidisciplinary team at each center. Between March 1994 and November 1997, 5 universities treated 221 adult patients with intraarterial chemotherapy with or without osmotic opening of the BBB (2464 procedures). Of evaluable patients with PCNSL, 40 of 53 (75%) achieved complete response (CR). All evaluable patients with PNET (n = 17), metastatic disease (n = 12), or germ cell tumor (n = 4) achieved stable disease (SD) or better. Of 57 evaluable patients with glioblastoma multiforme, 45 (79%) achieved SD or better. Asymptomatic subintimal tear occurred in 11of 221 patients (5%), pulmonary embolism in 6 of 221 (2.7%), and renal toxicity in 4 of 221 (1.8%). One patient with extensive glioma expired within 48 hours after treatment. Using standard guidelines and protocols, intraarterial chemotherapy with or without osmotic opening of the BBB is feasible across multiple centers with a low incidence of catheter-related complications. In patients with chemotherapy-sensitive tumors, such as PCNSL, PNET, germ cell tumor, and cancer metastasis to the central nervous system, enhanced delivery results in a high degree of tumor response, with an efficacy profile that is reproducible across multiple centers. Cancer 2000;88:637–47.
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In this study, the threshold for subharmonic emission during in vivo sonication of rabbit brain was investigated. In addition, the histologic effects of pulsed sonication above this threshold were studied. Two spherically curved focused ultrasound transducers with a diameter of 80 mm and a radius of curvature of 70 mm were used in the sonications. The operating frequencies of the transducers were 0.936 and 1.72 MHz. The sonication duration was varied between 0.001 and 1 s and the repetition frequency between 0.1 and 5 Hz. The threshold for subharmonic emission at the frequency of 0.936 MHz was found to be approximately 2000 W cm-2 and 3600 W cm-2 for pulse durations of 1 s and 0.001 s, respectively. The threshold was approximately 1.5-fold as high at a frequency of 1.72 MHz. However, there was considerable variation from experiment to experiment. The multiple pulse experiments at a frequency of 1.72 MHz and an intensity of 7000 W cm-2 showed that the histologic effects ranged from no observable damage of the tissue, to blood-brain barrier breakage, to local haemorrhagia, to local destruction of the tissue, to gross hemorrhage resulting in the death of the animal. The severity of the tissue damage increased as the pulse duration, number of pulses and their repetition frequency increased. The results indicate that the end point of the tissue damage may be controlled by selecting the sonication parameters. Such control over tissue effects can have several different applications when brain disorders are treated.
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In 1996, we are half-way through the Decade of the Brain, yet we still have few effective treatments for major disorders of the central nervous system. These include affective disorders, epilepsy, neurodegenerative disorders, brain tumours, infections and HIV encephalopathy; sufferers far outnumber the morbidity of cancer or heart disease. Increased understanding of the pharmacology of the brain and its blood supply, and methods for rational drug design, are leading to potential new drug therapies based on highly specific actions on particular target sites, such as neurotransmitter receptors and uptake systems. These methods are capable of reducing the side effects that are common with more general treatments. However, all these treatments and potential treatments meet a formidable obstacle--the blood-brain barrier. In this article, we review the properties of this barrier that complicate drug delivery to the brain, and some of the most hopeful strategies for overcoming or bypassing the barrier in humans.
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The distribution of the dopaminergic D4 receptor in rat brain was studied employing site directed polyclonal antibodies. Antisera were raised in rabbits to two oligopeptides corresponding to amino acids 160-172 of the second extracellular loop (P1) and amino acids 260-273 of the third intracellular loop (P2) of the D4 receptor sequence. Affinity-purified antibodies (anti-P1 and anti-P2) specifically recognized two major bands of 42-45 and 95 kDa in Western blots of denatured preparations of various rat brain areas. Immunocyto-chemistry studies showed that D4 receptor is widely distributed in rat central nervous system (CNS) showing higher labelling in the hippocampus (CA1, CA2, CA3 and dentate gyrus) frontal cortex, entorhinal cortex, caudate putamen, nucleus accumbens, olfactory tubercle, cerebellum, supraoptic nucleus and sustancia nigra pars compacta. In addition, anti-P1 decreased the binding of the antagonist [3H]YM-09151-2 selective for D2, D3 and D4 receptors but did not modify the binding of [3H]raclopride an antagonist selective for D2 and D3, in striatal synaptosomes. Anti-P2 did not modify the binding of these ligands. These results confirm the selectivity of the antibodies towards the D4 receptor and suggest that the binding site for the antagonists might be located at or close to the second extracellular loop of the protein sequence. D4 receptor protein is mainly expressed in plasma membranes and in the peripheral cytoplasm of neurons and is more widely distributed than was originally proposed based on mRNA localization, since it is present both in limbic, diencephalic and motor areas of rat brain.
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To determine the feasibility of the use of temperature-sensitive magnetic resonance (MR) imaging for the detection of local temperature elevations at the focus of a low-power ultrasound beam in the brain. The brains in 28 rabbits were sonicated at acoustic power levels of 3.5-17.5 W. Four to five different locations were sonicated at different acoustic power levels in each rabbit. MR images were obtained 2 hours, 48 hours, 10 days, and 23 days after the sonications, depending on when the animals were sacrificed. Histologic evaluation of whole brain was performed. Forty of 43 (93%) of the lowest-power (3.5-W) sonications were visible on temperature-sensitive MR images and did not result in any short- or long-term histologic or MR imaging evidence of tissue damage. A contrast-to-noise ratio of approximately 6 and a temperature elevation of 7 degrees-8 degrees C were observed. Temperature elevations induced by means of focused ultrasound exposures that do not cause damage in the in vivo rabbit brain can be detected at temperature-sensitive MR imaging.
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We investigated regional therapy of recurrent malignant brain tumors with transferrin-CRM107, a conjugate of human transferrin (Tf) and a genetic mutant of diphtheria toxin (CRM107) that lacks native toxin binding. Physiological barriers to delivering proteins to tumor and surrounding infiltrated brain were circumvented with high-flow interstitial microinfusion. At least a 50% reduction in tumor volume on magnetic resonance imaging (MRI) occurred in 9 of 15 patients who could be evaluated (60%), including two complete responses. Peritumoral toxicity developed 1-4 weeks after treatment in three of three patients at 1.0 microg/ml, but in zero of nine patients treated at lower concentrations. No symptomatic systemic toxicity occurred. Regional perfusion with Tf-CRM107 produces tumor responses without systemic toxicity in patients with malignant brain tumors refractory to conventional therapy. Direct interstitial infusion can be used successfully to distribute a large protein in the tumor and infiltrated brain surrounding the tumor.
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This article reviews historical aspects of the blood-brain barrier (BBB) and recent advances in mechanisms to deliver therapeutic agents across the BBB for the treatment of intracerebral tumors and other neurological diseases. The development of the osmotic BBB disruption procedure as a clinically useful technique is described. Osmotic BBB disruption is contrasted with alternative methods for opening or bypassing the BBB, including pharmacological modification of the BBB with bradykinin and direct intracerebral infusion. Laboratory studies have played a fundamental role in advancing our understanding of the BBB and delivery of agents to brain. Preclinical animal studies will continue to serve an integral function in our efforts to improve the diagnosis and treatment of a number of neurological disorders. Techniques involving the modification of the BBB and/or blood-tumor barrier to increase delivery of therapeutic agents have been advanced to clinical trials in patients with brain tumors with very favorable results. Improving delivery of agents to the brain will play a major role in the therapeutic outcome of brain neoplasms. As techniques for gene therapy are advanced, manipulation of the BBB also may be important in the treatment of central nervous system genetic disorders.
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Significant advances have been made in the application of monoclonal antibody-based therapies to the treatment of patients with lymphoma. The most promising areas appear to be the use of unconjugated monoclonal antibodies and the use of radiolabeled monoclonal antibodies. The recent approval by the US Food and Drug Administration (FDA) of rituximab (Rituxan), an unconjugated chimeric antibody against the CD20 antigen for the treatment of relapsed low-grade or follicular B-cell non-Hodgkin's lymphoma marked a milestone in the development of these antibody-based treatments. Other new drug applications to the FDA are pending using both unconjugated and radiolabeled monoclonal antibodies, and it is anticipated that further new treatment options based on monoclonal antibody technology will soon be available for the treatment of patients with non-Hodgkin's lymphoma. Forthcoming clinical trial results combining these new agents with current therapies are needed to determine if the addition of these new biologic agents to our armamentarium against lymphoma will alter the natural history of this disease for our patients. The most promising of these treatments and the comparison of these strategies are reviewed here.
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Delivery of therapeutic proteins into tissues and across the blood-brain barrier is severely limited by the size and biochemical properties of the proteins. Here it is shown that intraperitoneal injection of the 120-kilodalton beta-galactosidase protein, fused to the protein transduction domain from the human immunodeficiency virus TAT protein, results in delivery of the biologically active fusion protein to all tissues in mice, including the brain. These results open new possibilities for direct delivery of proteins into patients in the context of protein therapy, as well as for epigenetic experimentation with model organisms.
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The aim of this study was to determine the safety and efficacy of intraarterial chemotherapy with osmotic opening of the blood-brain barrier (BBB) for the treatment of malignant brain tumors when administered across multiple centers. Patients with primary central nervous system lymphoma (PCNSL), primitive neuroectodermal tumor (PNET), germ cell tumor, cancer metastasis to the brain, or low or high grade glioma were eligible. Prior to entry, magnetic resonance imaging or computed tomography brain scan, medical history, neurologic status, and Karnofsky performance status were reviewed at the coordinating center. Standardized anesthesia and intraarterial catheterization guidelines were followed by a multidisciplinary team at each center. Between March 1994 and November 1997, 5 universities treated 221 adult patients with intraarterial chemotherapy with or without osmotic opening of the BBB (2464 procedures). Of evaluable patients with PCNSL, 40 of 53 (75%) achieved complete response (CR). All evaluable patients with PNET (n = 17), metastatic disease (n = 12), or germ cell tumor (n = 4) achieved stable disease (SD) or better. Of 57 evaluable patients with glioblastoma multiforme, 45 (79%) achieved SD or better. Asymptomatic subintimal tear occurred in 11 of 221 patients (5%), pulmonary embolism in 6 of 221 (2.7%), and renal toxicity in 4 of 221 (1.8%). One patient with extensive glioma expired within 48 hours after treatment. Using standard guidelines and protocols, intraarterial chemotherapy with or without osmotic opening of the BBB is feasible across multiple centers with a low incidence of catheter-related complications. In patients with chemotherapy-sensitive tumors, such as PCNSL, PNET, germ cell tumor, and cancer metastasis to the central nervous system, enhanced delivery results in a high degree of tumor response, with an efficacy profile that is reproducible across multiple centers.
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Using polyclonal antibody against dopamine D4 receptor we investigated cortical distribution of D4 receptors, with the special emphasis on regions of the prefrontal cortex. Prefrontal cortex is regarded as a target for neuroleptic drugs, and engaged in the regulation of the psychotic effects of various substances used in the experimental modeling of schizophrenia. Western blot analysis performed on samples from the rat cingulate, parietal, piriform cortices and also striatum revealed that antibody recognized one main band of approximately 40 kD, which corresponds to the predicted molecular weight of D4 receptor protein. In immunocytochemical studies we found D4 receptor-positive neurons in all regions of prefrontal cortex (cingulate, agranular/insular and orbital cortices) and all cortical regions adjacent to prefrontal cortex, such as frontal, parietal and piriform cortex. Substantial number of D4 receptor-positive neurons has also been observed within the striatum and nucleus accumbens. In general, a clear stratification of the D4 receptor-positive neurons was observed in the cortex with the highest density seen in layers II/III and V/VI. D4 immunopositive material was also found in the dendritic processes, particularly clearly visible in the layer II/III. At the cellular level D4 receptor immunoreactivity was seen predominantly on the periphery of the cell body, but a certain population of neurons with clear cytoplasmatic localization was also identified. In addition to cortical distribution of D4 receptor-positive neurons we tried also to define types of neurons expressing D4 receptor protein. In double-labeling experiments, D4 receptor protein was found in nonphosphorylated neurofilament H-positive, calbindin-D28k-positive, as well as parvalbumin-positive cells. Since, used proteins are markers of certain populations of pyramidal neurons and GABA-ergic interneurons, respectively, our data indicate that D4 receptors are located on cortical pyramidal output neurons and their dendritic processes as well as on interneurons. Above localization indicates that D4 receptors are not only directly influencing excitability of cortical inter- and output neurons but also might be engaged in dendritic spatial and temporal integration, required for the generation of axonal messages. Additionally, our data show that D4 receptors are widely distributed throughout the cortex of rat brain, and that their cortical localization exceeds the localization of dopaminergic terminals.
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Much evidence indicates that abnormal processing and extracellular deposition of amyloid-beta peptide (A beta), a proteolytic derivative of the beta-amyloid precursor protein (betaAPP), is central to the pathogenesis of Alzheimer's disease (reviewed in ref. 1). In the PDAPP transgenic mouse model of Alzheimer's disease, immunization with A beta causes a marked reduction in burden of the brain amyloid. Evidence that A beta immunization also reduces cognitive dysfunction in murine models of Alzheimer's disease would support the hypothesis that abnormal A beta processing is essential to the pathogenesis of Alzheimer's disease, and would encourage the development of other strategies directed at the 'amyloid cascade'. Here we show that A beta immunization reduces both deposition of cerebral fibrillar A beta and cognitive dysfunction in the TgCRND8 murine model of Alzheimer's disease without, however, altering total levels of A beta in the brain. This implies that either a approximately 50% reduction in dense-cored A beta plaques is sufficient to affect cognition, or that vaccination may modulate the activity/abundance of a small subpopulation of especially toxic A beta species.
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Herceptin (trastuzumab), an anti-HER2 monoclonal antibody, is the first oncogene-targeted therapy to be developed for the treatment of metastatic breast cancer. The Herceptin clinical trial program has demonstrated that treatment with Herceptin provides substantial clinical benefits when used either as monotherapy or in combination with a number of chemotherapeutic agents. Of note, accurate assessment of HER2 status is essential to ensure that the patients most likely to benefit from Herceptin are identified: patients with immunohistochemistry (IHC) 3+ or fluorescence in-situ hybridization (FISH)-positive disease gain the greatest clinical benefits. In addition, clinical benefits appear to be greater the earlier Herceptin is used, although there is currently no direct clinical evidence to indicate whether an initial strategy of combination therapy is better than monotherapy or vice versa. Herceptin has been shown to be generally well tolerated. The most severe adverse events are rare serious infusion-related reactions and cardiotoxicity. These adverse events can be managed by standard care and patients at risk can often be identified prior to the initiation of Herceptin treatment. Currently, Herceptin should be given until disease progression, but there could be benefit in continuing treatment beyond disease progression.
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MRI-guided focused ultrasound was tested in the brains of rhesus monkeys. Locations up to 4.8 cm deep were targeted. Focal heating was observed in all cases with MRI-derived temperature imaging. Subthreshold heating was observed at the focus when the ultrasound beam was targeted with low power sonications, and in the ultrasound beam path during high-power exposures. Lethal temperature values and histologically confirmed tissue damage were confined to the focal zone (e.g., not in the ultrasound beam path), except when the focus was close to the bone. In that case, damage to the neighboring brain tissue was observed. Focal lesions were observed on histological examination and, in some cases, in MR images acquired immediately after the ultrasound exposures. The capabilities demonstrated in this study will be of benefit for clinical ultrasound therapies in the brain.
Article
The authors review the preclinical and clinical results of the ligand-targeted toxin conjugate Transferrin-CRM107 (Tf-CRM107), for the treatment of malignant gliomas. Tf-CRM107 is a conjugate protein of diphtheria toxin with a point mutation (CRM107) linked by a thioester bond to human transferrin (Tf). This conjugate exhibits potent cytotoxicity in vitro against mammalian cells expressing the transferrin receptor with activity at picomolar concentrations. Phase I clinical trial results demonstrated that Tf-CRM107, delivered via a high-flow convection method utilizing stereotactically placed catheters, produced tumor response in patients with malignant brain tumors refractory to conventional therapy without severe neurologic or systemic toxicity. The results of a Phase II study are also summarized. Tf-CRM107 treatment results in complete and partial tumor response without severe toxicity in 35% of the evaluable patients. These data warrant a Phase III study as well as continued research in the field of targeted toxin therapy. Future directions of research include optimizing Tf-CRM107 delivery to targeted brain regions, and improving the treatment efficacy by combining with other toxin conjugates targeted to different receptors.
Article
Blood brain barrier (BBB) is a major barrier for delivering therapeutic agents in the brain. In this study we investigated the feasibility of open the BBB by using focused ultrasound. Rabbit brains were exposed to pulsed focused ultrasound while injecting ultrasound contrast agent containg microbubbles intravenously. The BBB opening was measured after the sonications by injecting MRI contrast agent i.v. and evaluating the local enhancement in the brain. Low ultrasound powers and pressure amplitudes were found to cause focal enhancement. Before sacrificing the animals trypan blue was also injected i.v.. After the sacrifice of the animals blue spots were found in the brain in the sonicated locations. This method may have potential for targeted delivery of macromolecules in the brain.
Article
The ability of MRI-derived thermometry to predict thermally induced tissue changes in the brain was tested, and the thermal thresholds for blood-brain barrier (BBB) disruption and brain tissue damage were estimated. In addition, the ability of standard MRI to detect threshold-level effects was confirmed. These safety thresholds are being investigated to provide guidelines for clinical thermal ablation studies in the brain. MRI-monitored focused ultrasound heating was delivered to 63 locations in 26 rabbits. Tissue changes were detected in T(2)-weighted imaging and T(1)-weighted imaging (with and without contrast) and with light microscopy. The probability for tissue damage as a function of the accumulated thermal dose, the peak temperature achieved, the applied acoustic energy, and the peak acoustic power was estimated with probit regression. The discriminative abilities of these parameters were compared using the areas under the receiver operator characteristic (ROC) curves. In MRI, BBB disruption was observed in contrast-enhanced T(1)-weighted imaging shortly after the ultrasound exposures, sometimes accompanied by changes in T(2)-weighted imaging. Two days later, changes in T(2)-weighted imaging were observed, sometimes accompanied by changes in T(1)-weighted imaging. In histology, tissue damage was seen at every location where MRI changes were observed, ranging from small (diameter <1.0 mm) areas of tissue necrosis to severe vascular damage and associated hemorrhagic infarct. In one location, small (diameter: 0.8 mm) damage was not detected in MRI. The thermal dose and peak temperature thresholds were between 12.3-40.1 equivalent min at 43 degrees C and 48.0-50.8 degrees C, respectively, and values of 17.5 equivalent min at 43 degrees C and 48.4 degrees C were estimated to result in tissue damage with 50% probability. Thermal dose and peak temperature were significantly better predictors than the applied acoustic energy and peak acoustic power (P < 0.01). BBB disruption was always accompanied by tissue damage. The temperature information was better than the applied acoustic power or energy for predicting the damage than the ultrasound parameters. MRI was sensitive in detecting threshold-level damage.
Article
Local blood-brain barrier (BBB) opening is an advantageous approach for targeted drug delivery to the brain. Recently, it has been shown that focused ultrasound (US) exposures (sonications), when applied in the presence of preformed gas bubbles, caused magnetic-resonance (MR) proven reversible opening of the BBB in targeted locations. The cellular mechanisms of such transient barrier disruption are largely unknown. We investigated US-induced changes in endothelial cell fine morphology that resulted in the BBB opening in rabbits. To obtain evidence for the passage of blood-borne macromolecules through the opened transvascular routes, an immunocytochemical procedure for endogenous immunoglobulinG (IgG) was performed, in addition to the routine electron microscopy. An increased number of vesicles and vacuoles, fenestration and channel formation, as well as opening of some tight junctions, were seen in capillaries after low-power (0.55 W) sonication. Immunosignals presented in some of the vesicles and vacuoles, in the cytoplasmic channels and, so rarely, in intercellular clefts; immunosignals could also be seen in neuropil around the blood vessels. Damage to the cellular ultrastructure was not seen in these areas. However, cell destruction and leakage of IgG through defects of the endothelial lining took place at 3 W sonications. The data reveals that several mechanisms of transcapillary passage are possible after such sonications: 1. transcytosis; 2. endothelial cell cytoplasmic openings - fenestration and channel formation; 3. opening of a part of tight junctions; and 4. free passage through the injured endothelium (with the higher power sonications). These findings could be considered in further development of the strategy for drug delivery to brain parenchyma. (E-mail: [email protected] /* */)
Article
The purpose of this study was to test the hypothesis that burst ultrasound in the presence of an ultrasound contrast agent can disrupt the blood-brain barrier (BBB) with acoustic parameters suitable for completely noninvasive exposure through the skull. The 10-ms exposures were targeted in the brains of 22 rabbits with a frequency of 690 kHz, a repetition frequency of 1 Hz, and peak rarefactional pressure amplitudes up to 3.1 MPa. The total exposure (sonication) time was 20 s. Prior to each sonication, a bolus of ultrasound contrast agent was injected intravenously. Contrast-enhanced MR images were obtained after the sonications to detect localized BBB disruption via local enhancement in the brain. Brain sections were stained with H&E, TUNEL, and vanadium acid fuchsin (VAF)-toluidine blue staining. In addition, horseradish peroxidase (HRP) was injected into four rabbits prior to sonications and transmission electron microscopy was performed. The MRI contrast enhancement demonstrated BBB disruption at pressure amplitudes starting at 0.4 MPa with approximately 50%; at 0.8 MPa, 90%; and at 1.4 MPa, 100% of the sonicated locations showed enhancement. The histology findings following 4 h survival indicated that brain tissue necrosis was induced in approximately 70-80% of the sonicated locations at a pressure amplitude level of 2.3 MPa or higher. At lower pressure amplitudes, however, small areas of erythrocyte extravasation were seen. The electron microscopy findings demonstrated HRP passage through vessel walls via both transendothelial and paraendothelial routes. These results demonstrate that completely noninvasive focal disruption of the BBB is possible.
Article
Short interfering RNA (siRNA) has attracted much attention for clinical use in various diseases. However, its delivery, especially through the cell membrane, continues to present a challenge. Advances in ultrasound- and ultrasound contrast-agent technologies have made it possible to change transiently the permeability of the cell membrane and, using a focused ultrasound transducer, to narrow and focus the ultrasound energy on a small target, thereby avoiding damage to surrounding tissue. In this in vitro study, we demonstrate that it is possible to deliver siRNA intracellularly via microbubble-enhanced focused ultrasound. Although further optimization is necessary, our novel method for siRNA transduction represents a powerful tool for using siRNA in vivo and possibly in the clinical setting.
Drug and gene targeting to the brain with molecular Trojan horses
• Pardridge