Intravoxel incoherent motion (IVIM)-derived perfusion fraction mapping for the visual evaluation of MR-guided high intensity focused ultrasound (MR-HIFU) ablation of uterine fibroids.

BACKGROUND: A method for periprocedural contrast agent-free visualization of uterine fibroid perfusion could potentially shorten magnetic resonance-guided high intensity focused ultrasound (MR-HIFU) treatment times and improve outcomes. Our goal was to test feasibility of perfusion fraction mapping by intravoxel incoherent motion (IVIM) modeling using diffusion-weighted MRI as method for visual evaluation of MR-HIFU treatment progression. METHODS: Conventional and T2-corrected IVIM-derived perfusion fraction maps were retrospectively calculated by applying two fitting methods to diffusion-weighted MRI data (b = 0, 50, 100, 200, 400, 600 and 800 s/mm2 at 1.5 T) from forty-four premenopausal women who underwent MR-HIFU ablation treatment of uterine fibroids. Contrast in perfusion fraction maps between areas with low perfusion fraction and surrounding tissue in the target uterine fibroid immediately following MR-HIFU treatment was evaluated. Additionally, the Dice similarity coefficient (DSC) was calculated between delineated areas with low IVIM-derived perfusion fraction and hypoperfusion based on CE-T1w. RESULTS: Average perfusion fraction ranged between 0.068 and 0.083 in areas with low perfusion fraction based on visual assessment, and between 0.256 and 0.335 in surrounding tissues (all p < 0.001). DSCs ranged from 0.714 to 0.734 between areas with low perfusion fraction and the CE-T1w derived non-perfused areas, with excellent intraobserver reliability of the delineated areas (ICC 0.97). CONCLUSION: The MR-HIFU treatment effect in uterine fibroids can be visualized using IVIM perfusion fraction mapping, in moderate concordance with contrast enhanced MRI. IVIM perfusion fraction mapping has therefore the potential to serve as a contrast agent-free imaging method to visualize the MR-HIFU treatment progression in uterine fibroids.

Ultrasound and Microbubbles Mediated Bleomycin Delivery in Feline Oral Squamous Cell Carcinoma-An In Vivo Veterinary Study.

To investigate the feasibility and tolerability of ultrasound and microbubbles (USMB)-enhanced chemotherapy delivery for head and neck cancer, we performed a veterinary trial in feline companion animals with oral squamous cell carcinomas. Six cats were treated with a combination of bleomycin and USMB therapy three times, using the Pulse Wave Doppler mode on a clinical ultrasound system and EMA/FDA approved microbubbles. They were evaluated for adverse events, quality of life, tumour response and survival. Furthermore, tumour perfusion was monitored before and after USMB therapy using contrast-enhanced ultrasound (CEUS). USMB treatments were feasible and well tolerated. Among 5 cats treated with optimized US settings, 3 had stable disease at first, but showed disease progression 5 or 11 weeks after first treatment. One cat had progressive disease one week after the first treatment session, maintaining a stable disease thereafter. Eventually, all cats except one showed progressive disease, but each survived longer than the median overall survival time of 44 days reported in literature. CEUS performed immediately before and after USMB therapy suggested an increase in tumour perfusion based on an increase in median area under the curve (AUC) in 6 out of 12 evaluated treatment sessions. In this small hypothesis-generating study, USMB plus chemotherapy was feasible and well-tolerated in a feline companion animal model and showed potential for enhancing tumour perfusion in order to increase drug delivery. This could be a forward step toward clinical translation of USMB therapy to human patients with a clinical need for locally enhanced treatment.

Microbubble-Assisted Ultrasound for Drug Delivery to the Retina in an Ex Vivo Eye Model.

Drug delivery to the retina is one of the major challenges in ophthalmology due to the biological barriers that protect it from harmful substances in the body. Despite the advancement in ocular therapeutics, there are many unmet needs for the treatment of retinal diseases. Ultrasound combined with microbubbles (USMB) was proposed as a minimally invasive method for improving delivery of drugs in the retina from the blood circulation. This study aimed to investigate the applicability of USMB for the delivery of model drugs (molecular weight varying from 600 Da to 20 kDa) in the retina of ex vivo porcine eyes. A clinical ultrasound system, in combination with microbubbles approved for clinical ultrasound imaging, was used for the treatment. Intracellular accumulation of model drugs was observed in the cells lining blood vessels in the retina and choroid of eyes treated with USMB but not in eyes that received ultrasound only. Specifically, 25.6 ± 2.9% of cells had intracellular uptake at mechanical index (MI) 0.2 and 34.5 ± 6.0% at MI 0.4. Histological examination of retinal and choroid tissues revealed that at these USMB conditions, no irreversible alterations were induced at the USMB conditions used. These results indicate that USMB can be used as a minimally invasive targeted means to induce intracellular accumulation of drugs for the treatment of retinal diseases.

Focused Ultrasound and RadioTHERapy for non-invasive palliative pain treatment in patients with bone metastasis: a study protocol for the three armed randomized controlled FURTHER trial.

BACKGROUND: Cancer-induced bone pain (CIBP), caused by bone metastases, is a common complication of cancer and strongly impairs quality of life (QoL). External beam radiotherapy (EBRT) is the current standard of care for treatment of CIBP. However, approximately 45% of patients have no adequate pain response after EBRT. Magnetic resonance image-guided high-intensity focused ultrasound (MR-HIFU) may improve pain palliation in this patient population. The main objective of this trial was to compare MR-HIFU, EBRT, and MR-HIFU + EBRT for the palliative treatment of bone metastases. METHODS/DESIGN: The FURTHER trial is an international multicenter, three-armed randomized controlled trial. A total of 216 patients with painful bone metastases will be randomized in a 1:1:1 ratio to receive EBRT only, MR-HIFU only, or combined treatment with EBRT followed by MR-HIFU. During a follow-up period of 6 months, patients will be contacted at eight time points to retrieve information about their level of pain, QoL, and the occurrence of (serious) adverse events. The primary outcome of the trial is pain response at 14 days after start of treatment. Secondary outcomes include pain response at 14 days after trial enrolment, pain scores (daily until the 21st day and at 4, 6, 12 and 24 weeks), toxicity, adverse events, QoL, and survival. Cost-effectiveness and cost-utility analysis will be conducted. DISCUSSION: The FURTHER trial aims to evaluate the effectiveness and cost-effectiveness of MR-HIFU-alone or in combination with EBRT-compared to EBRT to relieve CIBP. The trial will be performed in six hospitals in four European countries, all of which are partners in the FURTHER consortium. TRIAL REGISTRATION: The FURTHER trial is registered under the Netherlands Trials Register number NL71303.041.19 and ClinicalTrials.gov registration number NCT04307914. Date of trial registration is 13-01-2020.

Ultrasound-directed enzyme-prodrug therapy (UDEPT) using self-immolative doxorubicin derivatives.

Background: Enzyme-activatable prodrugs are extensively employed in oncology and beyond. Because enzyme concentrations and their (sub)cellular compartmentalization are highly heterogeneous in different tumor types and patients, we propose ultrasound-directed enzyme-prodrug therapy (UDEPT) as a means to increase enzyme access and availability for prodrug activation locally. Methods: We synthesized ß-glucuronidase-sensitive self-immolative doxorubicin prodrugs with different spacer lengths between the active drug moiety and the capping group. We evaluated drug conversion, uptake and cytotoxicity in the presence and absence of the activating enzyme ß-glucuronidase. To trigger the cell release of ß-glucuronidase, we used high-intensity focused ultrasound to aid in the conversion of the prodrugs into their active counterparts. Results: More efficient enzymatic activation was observed for self-immolative prodrugs with more than one aromatic unit in the spacer. In the absence of ß-glucuronidase, the prodrugs showed significantly reduced cellular uptake and cytotoxicity compared to the parent drug. High-intensity focused ultrasound-induced mechanical destruction of cancer cells resulted in release of intact ß-glucuronidase, which activated the prodrugs, restored their cytotoxicity and induced immunogenic cell death. Conclusion: These findings shed new light on prodrug design and activation, and they contribute to novel UDEPT-based mechanochemical combination therapies for the treatment of cancer.

The Effect of Microbubble-Assisted Ultrasound on Molecular Permeability across Cell Barriers.

The combination of ultrasound and microbubbles (USMB) has been applied to enhance drug permeability across tissue barriers. Most studies focused on only one physicochemical aspect (i.e., molecular weight of the delivered molecule). Using an in vitro epithelial (MDCK II) cell barrier, we examined the effects of USMB on the permeability of five molecules varying in molecular weight (182 Da to 20 kDa) and hydrophilicity (LogD at pH 7.4 from 1.5 to highly hydrophilic). Treatment of cells with USMB at increasing ultrasound pressures did not have a significant effect on the permeability of small molecules (molecular weight 259 to 376 Da), despite their differences in hydrophilicity (LogD at pH 7.4 from -3.2 to 1.5). The largest molecules (molecular weight 4 and 20 kDa) showed the highest increase in the epithelial permeability (3-7-fold). Simultaneously, USMB enhanced intracellular accumulation of the same molecules. In the case of the clinically relevant anti- C-X-C Chemokine Receptor Type 4 (CXCR4) nanobody (molecular weight 15 kDa), USMB enhanced paracellular permeability by two-fold and increased binding to retinoblastoma cells by five-fold. Consequently, USMB is a potential tool to improve the efficacy and safety of the delivery of drugs to organs protected by tissue barriers, such as the eye and the brain.

Synthetic CT for the planning of MR-HIFU treatment of bone metastases in pelvic and femoral bones: a feasibility study.

OBJECTIVES: Visualization of the bone distribution is an important prerequisite for MRI-guided high-intensity focused ultrasound (MRI-HIFU) treatment planning of bone metastases. In this context, we evaluated MRI-based synthetic CT (sCT) imaging for the visualization of cortical bone. METHODS: MR and CT images of nine patients with pelvic and femoral metastases were retrospectively analyzed in this study. The metastatic lesions were osteolytic, osteoblastic or mixed. sCT were generated from pre-treatment or treatment MR images using a UNet-like neural network. sCT was qualitatively and quantitatively compared to CT in the bone (pelvis or femur) containing the metastasis and in a region of interest placed on the metastasis itself, through mean absolute difference (MAD), mean difference (MD), Dice similarity coefficient (DSC), and root mean square surface distance (RMSD). RESULTS: The dataset consisted of 3 osteolytic, 4 osteoblastic and 2 mixed metastases. For most patients, the general morphology of the bone was well represented in the sCT images and osteolytic, osteoblastic and mixed lesions could be discriminated. Despite an average timespan between MR and CT acquisitions of 61 days, in bone, the average (± standard deviation) MAD was 116 ± 26 HU, MD - 14 ± 66 HU, DSC 0.85 ± 0.05, and RMSD 2.05 ± 0.48 mm and, in the lesion, MAD was 132 ± 62 HU, MD - 31 ± 106 HU, DSC 0.75 ± 0.2, and RMSD 2.73 ± 2.28 mm. CONCLUSIONS: Synthetic CT images adequately depicted the cancellous and cortical bone distribution in the different lesion types, which shows its potential for MRI-HIFU treatment planning. KEY POINTS: • Synthetic computed tomography was able to depict bone distribution in metastatic lesions. • Synthetic computed tomography images intrinsically aligned with treatment MR images may have the potential to facilitate MR-HIFU treatment planning of bone metastases, by combining visualization of soft tissues and cancellous and cortical bone.

Workflow for automatic renal perfusion quantification using ASL-MRI and machine learning.

PURPOSE: Clinical applicability of renal arterial spin labeling (ASL) MRI is hampered because of time consuming and observer dependent post-processing, including manual segmentation of the cortex to obtain cortical renal blood flow (RBF). Machine learning has proven its value in medical image segmentation, including the kidneys. This study presents a fully automatic workflow for renal cortex perfusion quantification by including machine learning-based segmentation. METHODS: Fully automatic workflow was achieved by construction of a cascade of 3 U-nets to replace manual segmentation in ASL quantification. All 1.5T ASL-MRI data, including M0 , T1 , and ASL label-control images, from 10 healthy volunteers was used for training (dataset 1). Trained cascade performance was validated on 4 additional volunteers (dataset 2). Manual segmentations were generated by 2 observers, yielding reference and second observer segmentations. To validate the intended use of the automatic segmentations, manual and automatic RBF values in mL/min/100 g were compared. RESULTS: Good agreement was found between automatic and manual segmentations on dataset 1 (dice score = 0.78 ± 0.04), which was in line with inter-observer variability (dice score = 0.77 ± 0.02). Good agreement was confirmed on dataset 2 (dice score = 0.75 ± 0.03). Moreover, similar cortical RBF was obtained with automatic or manual segmentations, on average and at subject level; with 211 ± 31 mL/min/100 g and 208 ± 31 mL/min/100 g (P < .05), respectively, with narrow limits of agreement at -11 and 4.6 mL/min/100 g. RBF accuracy with automated segmentations was confirmed on dataset 2. CONCLUSION: Our proposed method automates ASL quantification without compromising RBF accuracy. With quick processing and without observer dependence, renal ASL-MRI is more attractive for clinical application as well as for longitudinal and multi-center studies.

Ultrasound-Mediated Drug Delivery With a Clinical Ultrasound System: In Vitro Evaluation.

Chemotherapy efficacy is often reduced by insufficient drug uptake in tumor cells. The combination of ultrasound and microbubbles (USMB) has been shown to improve drug delivery and to enhance the efficacy of several drugs in vitro and in vivo, through effects collectively known as sonopermeation. However, clinical translation of USMB therapy is hampered by the large variety of (non-clinical) US set-ups and US parameters that are used in these studies, which are not easily translated to clinical practice. In order to facilitate clinical translation, the aim of this study was to prove that USMB therapy using a clinical ultrasound system (Philips iU22) in combination with clinically approved microbubbles (SonoVue) leads to efficient in vitro sonopermeation. To this end, we measured the efficacy of USMB therapy for different US probes (S5-1, C5-1 and C9-4) and US parameters in FaDu cells. The US probe with the lowest central frequency (i.e. 1.6 MHz for S5-1) showed the highest USMB-induced intracellular uptake of the fluorescent dye SYTOX™ Green (SG). These SG uptake levels were comparable to or even higher than those obtained with a custom-built US system with optimized US parameters. Moreover, USMB therapy with both the clinical and the custom-built US system increased the cytotoxicity of the hydrophilic drug bleomycin. Our results demonstrate that a clinical US system can be used to perform USMB therapy as efficiently as a single-element transducer set-up with optimized US parameters. Therefore, future trials could be based on these clinical US systems, including validated US parameters, in order to accelerate successful translation of USMB therapy.

Ultrasound and Microbubbles for the Treatment of Ocular Diseases: From Preclinical Research towards Clinical Application.

The unique anatomy of the eye and the presence of various biological barriers make efficacious ocular drug delivery challenging, particularly in the treatment of posterior eye diseases. This review focuses on the combination of ultrasound and microbubbles (USMB) as a minimally invasive method to improve the efficacy and targeting of ocular drug delivery. An extensive overview is given of the in vitro and in vivo studies investigating the mechanical effects of ultrasound-driven microbubbles aiming to: (i) temporarily disrupt the blood-retina barrier in order to enhance the delivery of systemically administered drugs into the eye, (ii) induce intracellular uptake of anticancer drugs and macromolecules and (iii) achieve targeted delivery of genes, for the treatment of ocular malignancies and degenerative diseases. Finally, the safety and tolerability aspects of USMB, essential for the translation of USMB to the clinic, are discussed.

AAPM Task Group 241: A medical physicist's guide to MRI-guided focused ultrasound body systems.

Magnetic resonance-guided focused ultrasound (MRgFUS) is a completely non-invasive technology that has been approved by FDA to treat several diseases. This report, prepared by the American Association of Physicist in Medicine (AAPM) Task Group 241, provides background on MRgFUS technology with a focus on clinical body MRgFUS systems. The report addresses the issues of interest to the medical physics community, specific to the body MRgFUS system configuration, and provides recommendations on how to successfully implement and maintain a clinical MRgFUS program. The following sections describe the key features of typical MRgFUS systems and clinical workflow and provide key points and best practices for the medical physicist. Commonly used terms, metrics and physics are defined and sources of uncertainty that affect MRgFUS procedures are described. Finally, safety and quality assurance procedures are explained, the recommended role of the medical physicist in MRgFUS procedures is described, and regulatory requirements for planning clinical trials are detailed. Although this report is limited in scope to clinical body MRgFUS systems that are approved or currently undergoing clinical trials in the United States, much of the material presented is also applicable to systems designed for other applications.

Interleaved water and fat MR thermometry for monitoring high intensity focused ultrasound ablation of bone lesions.

PURPOSE: To demonstrate that interleaved MR thermometry can monitor temperature in water and fat with adequate temporal resolution. This is relevant for high intensity focused uUltrasounds (HIFU) treatment of bone lesions, which are often found near aqueous tissues, as muscle, or embedded in adipose tissues, as subcutaneous fat and bone marrow. METHODS: Proton resonance frequency shift (PRFS)-based thermometry scans and T1 -based 2D variable flip angle (2D-VFA) thermometry scans were acquired alternatingly over time. Temperature in water was monitored using PRFS thermometry, and in fat by 2D-VFA thermometry with slice profile effect correction. The feasibility of interleaved water/fat temperature monitoring was studied ex vivo in porcine bone during MR-HIFU sonication. Precision and stability of measurements in vivo were evaluated in a healthy volunteer under non-heating conditions. RESULTS: The method allowed observing temperature change over time in muscle and fat, including bone marrow, during MR-HIFU sonication, with a temporal resolution of 6.1 s. In vivo, the apparent temperature change was stable on the time scale of the experiment: In 7 min the systematic drift was <0.042°C/min in muscle (PRFS after drift correction) and <0.096°C/min in bone marrow (2D-VFA). The SD of the temperature change averaged over time was 0.98°C (PRFS) and 2.7°C (2D-VFA). CONCLUSIONS: Interleaved MR thermometry allows temperature measurements in water and fat with a temporal resolution high enough for monitoring HIFU ablation. Specifically, combined fat and water thermometry provides uninterrupted information on temperature changes in tissue close to the bone cortex.

Rapid 2D variable flip angle method for accurate and precise T1 measurements over a wide range of T1 values.

PURPOSE: To perform dynamic T1 mapping using a 2D variable flip angle (VFA) method, a correction for the slice profile effect is needed. In this work we investigated the impact of flip angle selection and excitation RF pulse profile on the performance of slice profile correction when applied to T1 mapping over a range of T1 values. METHODS: A correction of the slice profile effect is proposed, based on Bloch simulation of steady-state signals. With this correction, Monte Carlo simulations were performed to assess the accuracy and precision of 2D VFA T1 mapping in the presence of noise, for RF pulses with time-bandwidth products of 2, 3 and 10 and with flip angle pairs in the range [1°-90°]. To evaluate its performance over a wide range of T1 , maximum errors were calculated for six T1 values between 50 ms and 1250 ms. The method was demonstrated using in vitro and in vivo experiments. RESULTS: Without corrections, 2D VFA severely underestimates T1 . Slice profile errors were effectively reduced with the correction based on simulations, both in vitro and in vivo. The precision and accuracy of the method depend on the nominal T1 values, the FA pair, and the RF pulse shape. FA pairs leading to <5% errors in T1 can be identified for the common RF shapes, for T1 values between 50 ms and 1250 ms. CONCLUSIONS: 2D VFA T1 mapping with Bloch-simulation-based correction can deliver T1 estimates that are accurate and precise to within 5% over a wide T1 range.

Combining radiotherapy and focused ultrasound for pain palliation of cancer induced bone pain; a stage I/IIa study according to the IDEAL framework.

BACKGROUND: Cancer induced bone pain (CIBP) strongly interferes with patient's quality of life. Currently, the standard of care includes external beam radiotherapy (EBRT), resulting in pain relief in approximately 60% of patients. Magnetic Resonance guided High Intensity Focused Ultrasound (MR-HIFU) is a promising treatment modality for CIBP. METHODS: A single arm, R-IDEAL stage I/IIa study was conducted. Patients presenting at the department of radiation oncology with symptomatic bone metastases in the appendicular skeleton, as well as in the sacrum and sternum were eligible for inclusion. All participants underwent EBRT, followed by MR-HIFU within 4 days. Safety and feasibility were assessed, and pain scores were monitored for 4 weeks after completing the combined treatment. RESULTS: Six patients were enrolled. Median age was 67 years, median lesion diameter was 56,5 mm. In all patients it was logistically possible to plan and perform the MR-HIFU treatment within 4 days after EBRT. All patients tolerated the combined procedure well. Pain response was reported by 5 out of 6 patients at 7 days after completion of the combined treatment, and stabilized on 60% at 4 weeks follow up. No treatment related serious adverse events occurred. CONCLUSION: This is the first study to combine EBRT with MR-HIFU. Our results show that combined EBRT and MR-HIFU in first-line treatment of CIBP is safe and feasible, and is well tolerated by patients. Superiority over standard EBRT, in terms of (time to) pain relief and quality of life need to be evaluated in comparative (randomized) study.

Exploring label dynamics of velocity-selective arterial spin labeling in the kidney.

PURPOSE: Velocity-selective arterial spin labeling (VSASL) has been proposed for renal perfusion imaging to mitigate planning challenges and effects of arterial transit time (ATT) uncertainties. In VSASL, label generation may shift in the vascular tree as a function of cutoff velocity. Here, we investigate label dynamics and especially the ATT of renal VSASL and compared it with a spatially selective pulsed arterial spin labeling technique, flow alternating inversion recovery (FAIR). METHODS: Arterial spin labeling data were acquired in 7 subjects, using free-breathing dual VSASL and FAIR with five postlabeling delays: 400, 800, 1200, 2000, and 2600 ms. The VSASL measurements were acquired with cutoff velocities of 5, 10, and 15 cm/s, with anterior-posterior velocity-encoding direction. Cortical perfusion-weighted signal, temporal SNR, quantified renal blood flow, and arterial transit time were reported. RESULTS: In contrast to FAIR, renal VSASL already showed fairly high signal at the earliest postlabeling delays, for all cutoff velocities. The highest VSASL signal and temporal SNR was obtained with a cutoff velocity of 10 cm/s at postlabeling delay = 800 ms, which was earlier than for FAIR at 1200 ms. Fitted ATT on VSASL was ≤ 0 ms, indicating ATT insensitivity, which was shorter than for FAIR (189 ± 79 ms, P < .05). Finally, the average cortical renal blood flow measured with cutoff velocities of 5 cm/s (398 ± 84 mL/min/100 g) and 10 cm/s (472 ± 160 mL/min/100 g) were similar to renal blood flow measured with FAIR (441 ± 84 mL/min/100 g) (P > .05) with good correlations on subject level. CONCLUSION: Velocity-selective arterial spin labeling in the kidney reduces ATT sensitivity compared with the recommended pulsed arterial spin labeling method, as well as if cutoff velocity is increased to reduce spurious labeling due to motion. Thus, VSASL has potential as a method for time-efficient, single-time-point, free-breathing renal perfusion measurements, despite lower tSNR than FAIR.

Synthesis, characterization, and imaging of radiopaque bismuth beads for image-guided transarterial embolization.

Current therapy for hypervascular cancers, e.g., hepatocellular carcinoma, includes occlusion of the tumor blood supply by arterial infusion of embolic microspheres (beads) suspended in iodine-based contrast under fluoroscopic guidance. Available radiopaque, imageable beads use iodine as the radiopacifier and cannot be differentiated from contrast. This study aimed to synthesize and characterize imageable beads using bismuth as the radiopacifier that could be distinguished from iodine contrast based upon the difference in the binding energy of k-shell electrons (k-edge). Radiodense bismuth beads were successfully synthesized some with uniform bismuth distribution across the beads. The beads were spherical and could be infused through clinical microcatheters. The bismuth beads could be imaged with clinical dual-energy computed tomography (CT), where iodine-based contrast could be distinguished from the microspheres. The ability to separate iodine from bismuth may enhance the diagnostic information acquired on follow-up CT, identifying the distribution of the embolic beads separately from the contrast. Furthermore, with sequential use of iodine- and bismuth-based beads, the two radiopaque beads could be spatially distinguished on imaging, which may enable the development of dual drug delivery and dual tracking.

Phase I feasibility study of Magnetic Resonance guided High Intensity Focused Ultrasound-induced hyperthermia, Lyso-Thermosensitive Liposomal Doxorubicin and cyclophosphamide in de novo stage IV breast cancer patients: study protocol of the i-GO study.

INTRODUCTION: In breast cancer, local tumour control is thought to be optimised by administering higher local levels of cytotoxic chemotherapy, in particular doxorubicin. However, systemic administration of higher dosages of doxorubicin is hampered by its toxic side effects. In this study, we aim to increase doxorubicin deposition in the primary breast tumour without changing systemic doxorubicin concentration and thus without interfering with systemic efficacy and toxicity. This is to be achieved by combining Lyso-Thermosensitive Liposomal Doxorubicin (LTLD, ThermoDox, Celsion Corporation, Lawrenceville, NJ, USA) with mild local hyperthermia, induced by Magnetic Resonance guided High Intensity Focused Ultrasound (MR-HIFU). When heated above 39.5°C, LTLD releases a high concentration of doxorubicin intravascularly within seconds. In the absence of hyperthermia, LTLD leads to a similar biodistribution and antitumour efficacy compared with conventional doxorubicin. METHODS AND ANALYSIS: This is a single-arm phase I study in 12 chemotherapy-naïve patients with de novo stage IV HER2-negative breast cancer. Previous endocrine treatment is allowed. Study treatment consists of up to six cycles of LTLD at 21-day intervals, administered during MR-HIFU-induced hyperthermia to the primary tumour. We will aim for 60 min of hyperthermia at 40°C-42°C using a dedicated MR-HIFU breast system (Profound Medical, Mississauga, Canada). Afterwards, intravenous cyclophosphamide will be administered. Primary endpoints are safety, tolerability and feasibility. The secondary endpoint is efficacy, assessed by radiological response.This approach could lead to optimal loco-regional control with less extensive or even no surgery, in de novo stage IV patients and in stage II/III patients allocated to receive neoadjuvant chemotherapy. ETHICS AND DISSEMINATION: This study has obtained ethical approval by the Medical Research Ethics Committee Utrecht (Protocol NL67422.041.18, METC number 18-702). Informed consent will be obtained from all patients before study participation. Results will be published in an academic peer-reviewed journal. TRIAL REGISTRATION NUMBERS: NCT03749850, EudraCT 2015-005582-23.

New Developments in Imaging for Sentinel Lymph Node Biopsy in Early-Stage Oral Cavity Squamous Cell Carcinoma.

Sentinel lymph node biopsy (SLNB) is a diagnostic staging procedure that aims to identify the first draining lymph node(s) from the primary tumor, the sentinel lymph nodes (SLN), as their histopathological status reflects the histopathological status of the rest of the nodal basin. The routine SLNB procedure consists of peritumoral injections with a technetium-99m [99mTc]-labelled radiotracer followed by lymphoscintigraphy and SPECT-CT imaging. Based on these imaging results, the identified SLNs are marked for surgical extirpation and are subjected to histopathological assessment. The routine SLNB procedure has proven to reliably stage the clinically negative neck in early-stage oral squamous cell carcinoma (OSCC). However, an infamous limitation arises in situations where SLNs are located in close vicinity of the tracer injection site. In these cases, the hotspot of the injection site can hide adjacent SLNs and hamper the discrimination between tracer injection site and SLNs (shine-through phenomenon). Therefore, technical developments are needed to bring the diagnostic accuracy of SLNB for early-stage OSCC to a higher level. This review evaluates novel SLNB imaging techniques for early-stage OSCC: MR lymphography, CT lymphography, PET lymphoscintigraphy and contrast-enhanced lymphosonography. Furthermore, their reported diagnostic accuracy is described and their relative merits, disadvantages and potential applications are outlined.

A Doxorubicin-Glucuronide Prodrug Released from Nanogels Activated by High-Intensity Focused Ultrasound Liberated ß-Glucuronidase.

The poor pharmacokinetics and selectivity of low-molecular-weight anticancer drugs contribute to the relatively low effectiveness of chemotherapy treatments. To improve the pharmacokinetics and selectivity of these treatments, the combination of a doxorubicin-glucuronide prodrug (DOX-propGA3) nanogel formulation and the liberation of endogenous ß-glucuronidase from cells exposed to high-intensity focused ultrasound (HIFU) were investigated in vitro. First, a DOX-propGA3-polymer was synthesized. Subsequently, DOX-propGA3-nanogels were formed from this polymer dissolved in water using inverse mini-emulsion photopolymerization. In the presence of bovine ß-glucuronidase, the DOX-propGA3 in the nanogels was quantitatively converted into the chemotherapeutic drug doxorubicin. Exposure of cells to HIFU efficiently induced liberation of endogenous ß-glucuronidase, which in turn converted the prodrug released from the DOX-propGA3-nanogels into doxorubicin. ß-glucuronidase liberated from cells exposed to HIFU increased the cytotoxicity of DOX-propGA3-nanogels to a similar extend as bovine ß-glucuronidase, whereas in the absence of either bovine ß-glucuronidase or ß-glucuronidase liberated from cells exposed to HIFU, the DOX-propGA3-nanogels hardly showed cytotoxicity. Overall, DOX-propGA3-nanogels systems might help to further improve the outcome of HIFU-related anticancer therapy.

Potential Use of Extracellular Vesicles Generated by Microbubble-Assisted Ultrasound as Drug Nanocarriers for Cancer Treatment.

Extracellular vesicles (EVs)-carrying biomolecules derived from parental cells have achieved substantial scientific interest for their potential use as drug nanocarriers. Ultrasound (US) in combination with microbubbles (MB) have been shown to trigger the release of EVs from cancer cells. In the current study, the use of microbubbles-assisted ultrasound (USMB) to generate EVs containing drug cargo was investigated. The model drug, CellTracker™ green fluorescent dye (CTG) or bovine serum albumin conjugated with fluorescein isothiocyanate (BSA FITC) was loaded into primary human endothelial cells in vitro using USMB. We found that USMB loaded CTG and BSA FITC into human endothelial cells (HUVECs) and triggered the release of EVs containing these compounds in the cell supernatant within 2 h after treatment. The amount of EV released seemed to be correlated with the increase of US acoustic pressure. Co-culturing these EVs resulted in uptake by the recipient tumour cells within 4 h. In conclusion, USMB was able to load the model drugs into endothelial cells and simultaneously trigger the release of EVs-carrying model drugs, highlighting the potential of EVs as drug nanocarriers for future drug delivery in cancer.