With cavitational therapies continuing to be adopted clinically, the ability to accurately quantify and compare treatments is essential to ensure consistent efficacy and patient safety. Passive Acoustic Mapping (PAM) can reconstruct quantitative maps of cavitational energy density in real time, providing a powerful platform for cavitation dosimetry. However, the quality and accuracy of PAM images can be significantly affected by the measurement setup, physical properties of the monitoring device, and choice of computational parameters. To obtain reliable cavitation doses that can be compared across indications and experiments, compensations for the device, position, and frequency dependent point spread function as well as attenuation and array element sensitivity and directivity must be applied. The selection of beamforming parameters, such as data acquisition length and windowing, voxel size, and array element decimation, can profoundly affect the accuracy of PAM-derived cavitation doses. This work will present a step-by-step guide to mapping cavitational signals, using practical examples of in vitro, ex vivo, and in vivo experimental data covering a range of device geometries and measurement setups to illustrate potential pitfalls. Trade-offs between image quality, accuracy, and computation time will be explored for each application, toward energy preserving, setup independent dosimetry of cavitational treatments.

In cavitation-mediated focused ultrasound therapies, it is common to use separate devices for therapy beam transmission and bubble activity monitoring. In the interest of reducing complexity and cost of instrumentation, we present several examples demonstrating the use of conventional handheld arrays for all required functions: cavitation generation, passive acoustic mapping, and structural imaging. Strategies for transmit and receive beamforming, aperture use, and real time cavitation dose control are presented with three different array geometries and data sets spanning small animal invivo and perfused human organ deployments. We further discuss limitations, solutions, and prospects for handheld array deployments in clinical treatment scenarios.

3549 Background: Effective treatment of colorectal liver metastases is limited by poor drug delivery, particularly for monoclonal antibodies, the large molecular weight of which, together with abnormal tumor vasculature, elevated interstitial pressure and dense stromal architecture, restricts penetration and distribution into solid tumors. This first-in-human study evaluated a novel platform combining intravenously infused sub-micron gas-stabilizing sonosensitive particles with novel handheld ultrasound probe to cause cavitation that enhances the transport of systemically co-administered therapeutics (i.e. cetuximab/irinotecan) from the vasculature into tumors, without the need to modify these therapeutics. Methods: The study comprised 3 patient cohorts. Cohort 1 evaluated the safety of ultrasound activation of intravenously infused sonosensitive particles alone, without drug. With safety confirmed, Cohort 2 assessed intratumoural drug concentration in patients undergoing surgical resection after receiving subtherapeutic irinotecan and cetuximab, with or without ultrasound-activated particles. Cohort 3 evaluated early efficacy in patients receiving standard of care FOLFIRI and cetuximab, with or without cavitation in repeated treatment cycles. Primary endpoints were grade 3 or higher adverse events in Cohort 1, intratumoural drug concentrations in Cohort 2, and radiological tumor response in Cohort 3. Results: In Cohort 2, seven evaluable patients underwent resection, with 4 in the control arm and 3 in the ultrasound-mediated-cavitation arm. Mean cetuximab concentrations were modestly higher in the tumour centre (3.6 vs 3.2 ng/mL) and mid radius (4.1 vs 3.7 ng/mL), and substantially higher at the tumour periphery (7.1 vs 3.7 ng/mL) in the ultrasound-mediated-cavitation arm, while liver margin levels were comparable. There was no appreciable beneficial difference in the mean tumour concentrations of irinotecan and its metabolites between the two arms. In Cohort 3, 10 evaluable participants completed at least 3 treatment cycles (control n=3; cavitation n=7). Objective response rate was 66.7% in the control arm versus 85.7 % with cavitation. Conclusions: Cavitation with sonosensitive particle infusion demonstrated a favourable safety profile, increased intratumoural antibody delivery, and showed early signals of enhanced anti-tumour activity, supporting further clinical development. Clinical trial information: ISRCTN17598292. Cohort Number treated Primary endpoint Key findings 1 9 Safety No grade 3 or higher events; sustained cavitation in the target region. 2 7 Drug levels Increased cetuximab concentrations in the tumours treated with cavitation (7.1 vs 3.7 ng/mL) 3 10 Radiological response Enhanced radiological response (ORR of 85.7% in patients treated with cavitation) ORR = objective response rate based on FDG PET-CT.

The amygdala shows abnormal metabolism in depression, a disorder marked by altered emotion, motivation, and learning. Yet its causal role in these processes remains unclear because non-invasive, reversible perturbation in humans has not been possible. We used transcranial focused ultrasound stimulation (TUS) to modulate basolateral amygdala (BLA) activity. In separate sessions, healthy volunteers received offline TUS to bilateral BLA, mid-insula, or sham before completing a novel emotional learning task validated online. 7T-resting-state connectivity and metabolite measures confirmed target engagement: BLA-TUS reduced the BLA’s connectivity fingerprint and lowered its excitation/inhibition balance. Behaviorally, BLA-TUS increased approach tendencies toward neutral, emotionally ambiguous faces in a stimulation-volume-dependent manner and slowed responses to neutral and happy faces. These effects were functionally and regionally specific and suggest a causal role for the amygdala in resolving emotional ambiguity. Our findings inform studies of mood disorders, where difficulty resolving ambiguity may contribute to emotional and learning biases.

Marine bioacousticians are increasingly interested in measurement of acoustic vector fields (particle motion) to investigate sound sensitivity, noise impacts, and use of soundscapes by fishes and invertebrates. These animals detect sounds by transducing particle motion, and only some sense acoustic pressure. Bioacousticians studying these taxa often ask: when is a single pressure sensor sufficient to estimate particle motion versus measurement of the 3-D vector field? While measuring directionality or polarization properties requires a vector sensor, amplitude (particle motion level) can sometimes be obtained from a single hydrophone. Past and current empirical studies with compact hydrophone arrays in a variety of underwater habitats will be contrasted to answer this question. Focus regions include Caribbean coral reefs, the U.S. Mid and South Atlantic continental slope, and offshore Gulf of Maine waters. Together with recent related literature, these studies highlight that the ratio of measured particle motion amplitude to that estimated from single hydrophones depends on frequency, depth, near versus far field, time and frequency averaging, and ambient noise versus discrete sound sources. Such factors can lead to high variability of these ratios within and across datasets. We will provide recommendations for marine bioacousticians as they seek to quantify particle motion amplitudes.

The ability to quantify and compare different cavitational treatments is essential given their accelerating clinical adoption. Passive acoustic mapping (PAM) can reconstruct quantitative maps of cavitational radiated energy density (CRED) in real time, providing a powerful platform for cavitation dosimetry. However, significant imaging artefacts caused by a large and variable point spread function (PSF) limit the spatial resolvability of PAM-derived cavitation doses. The Lucy-Richardson deconvolution (LRD) can significantly reduce imaging artefacts, correct for gross overestimates of CRED values, and reduce setup-dependence of CRED estimates when applied to PAM images. However, the LRD condenses source energy densities into the central axis of the image, obscuring the underlying source distribution. Typically, a single central PSF is used to perform deconvolution, which additionally preserves biases in energy estimation due to PSF variation. This work investigates the underlying causes of these errors and presents four deconvolution approaches designed to preserve source localisation and improve spatial accuracy of PAM images. Each algorithm is tested using numerical simulations and experimental cavitational data monitoring cavitation in vitro as well as in normothermically perfused porcine liver, and show varied accuracy, robustness, and computational load. Modelling PSF variations over space to enable a shift-variant implementation of the LRD (svLRD-PAM) improves the spatial accuracy of cavitation doses and reduces imaging artefacts by 4-5 fold with a modest and parallelisable 36% increase in computation time relative to the standard delay-sum-integrate beamformer. svLRD-PAM shows great promise as the next step towards accurate, spatially resolvable cavitation dosimetry.

Orthopaedic knee implant-associated infections caused by biofilm-forming bacteria present clinical and economic challenges due to poor antibiotic penetration and immune evasion. Shock wave (SW)-based therapies are a promising non-invasive approach to biofilm disruption, and acoustic models of the complex propagation environment would aid translational efforts. We treated S taphylococcus aureus biofilms grown on ∼12-mm diameter stainless steel, titanium, and hydroxyapatite discs with either 50 SWs (focused on the centre of the disc; 3.5 MPa peak negative pressure (PNP), 0.5 Hz; 70 mJ incident energy) or 6 SWs (scanned over the disc; 3.5 MPa PNP; 5 mJ incident energy). Passive cavitation detection (PCD) characterized bubble cloud dynamics, including collapse time. The 50 centredSWs produced 3.8-log (95% CI: 3.1–4.5) reductions in biofilm-associated bacteria, and the six scanned SWs produced 2.5-log (95% CI: 1.8–3.2) reductions, demonstrating that shock waves have the potential to remove biofilm with relatively low energy. Optical profilometry was carried out on the disc to assess surface damage. Three-dimensional simulations were conducted using CT-derived anatomy for knee implants, identifying regions where SW energy can be effectively delivered to the knee joint. These results suggest that non-invasiveSW therapy can be used to address implant-related infections.

Orthopaedic implant infections caused by biofilm-forming bacteria pose significant clinical and economic challenges due to high revision rates and increasing antibiotic resistance. Biofilms limit antibiotic penetration, rendering many conventional treatments ineffective. Cavitation, either spontaneous or facilitated by cavitation nuclei, can mechanically disrupt biofilms, offering a promising alternative. This study evaluates the efficacy of therapeutic ultrasound (US) and shockwaves (SW) in treating Staphylococcus aureus biofilms on clinically relevant implant materials, both independently and combined with antibiotics. In vitro biofilms were cultured on stainless steel, hydroxyapatite, and titanium discs (10–12 mm). Treatments included scanning US (0.95 MHz, 2.5 MPa PNP, 500 Hz pulse repetition frequency, 10% duty cycle, 10 s per location across nine positions) with in-house protein-based cavitation nuclei and SW (3.5 MPa PNP, 0.5 Hz PRF, 50 pulses). Passive cavitation detection monitored emissions during treatment. Biofilm disruption was quantified by fluorescence imaging and bacterial colony-forming unit counts. Both US and SW significantly reduced biofilm burden, with substrate-specific effects and distinct spatial disruption patterns (US: 1.5±0.8 and SW: 4.0±1.1 log reduction). Ongoing work is assessing whether detached aggregates exhibit increased antibiotic susceptibility. These findings support cavitation-based methods as minimally invasive adjuncts to enhance antibiotic efficacy and reduce revision surgeries.

The acoustic emissions generated by oscillating gas bubbles (cavitation) provide an extremely useful means of treatment monitoring in ultrasound-mediated therapy. Metrics such as the relative proportions of harmonics and broadband noise in the acoustic emissions frequency spectra are widely used to categorize cavitation, e.g., as “inertial” or “stable.” It is unclear, however, whether these simple categories can adequately describe the wide range of bubble dynamics that may occur or the effects that these may produce in tissue. We have previously shown through simultaneous capture of high-speed video footage and acoustic radiation that even for single bubbles there is no simple correlation between different types of bubble behavior and the frequency content of the acoustic emissions. This suggests that measures such as the onset of broadband noise, or the appearance of sub- or ultra-harmonics are not appropriate for use as universal thresholds in therapeutic ultrasound. Binary categories such as stable and inertial cavitation, or the use of spectral characteristics to infer these, should similarly be avoided. In this talk, the implications of these findings for defining a cavitation dose and applying it to different types of therapy will be discussed.

A number of factors impede the pursuit of reliable and meaningful dosimetry for cavitation-mediated therapies. This presentation will review major technical challenges in the measurement of biomedical cavitation, along with the resulting uncertainties in cavitation dose calculation across several treatment modalities. Mitigation methods for propagation path, receiver, and processing related effects will then be summarized. Finally, an optimistic outlook for this field will be highlighted based on progress made to date and lessons learned from other areas of medical dosimetry.

Back pain from spinal disc degeneration affects over 600 million people worldwide. Treatments range from conservative approaches like physiotherapy to invasive procedures such as spinal fusion. Injectable engineered hydrogels offer a promising minimally invasive alternative; however, conventional self-curing lacks control, and optical curing is limited by poor tissue penetration. In contrast, ultrasound-triggered implant formation enables precise spatiotemporal control of gelation at clinically relevant depths. We developed a cavitation-based ultrasound guidance protocol using broadband cavitation emissions as a real-time metric of hydrogel gelation and stiffening. In vitro experiments with hydrogel precursors containing calcium-loaded liposomes showed a rapid drop in broadband cavitation activity within 100–150 s, reflecting gel stiffening and inhibited bubble dynamics. Building on this, an automated feedback system was implemented where ultrasound was modulated based on the running median of broadband power relative to a dynamic threshold, enabling on/off cycling until cavitation ceased, signaling treatment completion. Passive thermometry confirmed safe temperature control. Validation in nine ex vivo bovine spinal units (six treated, three control) confirmed effective ultrasound-triggered gelation without material herniation and partial restoration of biomechanical function. This cavitation-guided approach represents a key advancement in non-invasive, adaptive monitoring for ultrasound-mediated spinal disc repair.

Passive Acoustic Mapping (PAM) is a leading approach for the quantification of cavitation energy and can inform the safe and effective use of cavitation in medicine. To achieve quantitative measurements, the sensitivity and directivity of each element in a PAM array must be calibrated, and any issues in the calibration can propagate into poor resolution, incorrect magnitudes, and location errors in all subsequent PAM images. Accurate array calibrations are therefore crucial in the pursuit of a “cavitation dose” to correlate with bioeffects. The most commonly used calibration method, reported by Gray and Coussios in 2018, is a substitution approach using a fine wire scatterer to approximate a point source. Despite its apparent simplicity, this experiment features a number of complicating factors that can prohibit successful measurements, particularly for newer acousticians. Here, we describe the calibration experiment in detail, along with the impacts, causes, and solutions of a range of issues identified through the authors’ hard-won experience. Special consideration will be given to the selection of sources and hydrophones, alignment, management of signal-to-noise ratio, and data processing. We will also discuss common assumptions and their validity, measurement uncertainty, comparing results to theory, and future directions for this area of ultrasound metrology.

Passive Acoustic Mapping (PAM) makes it possible to qualify and quantify the spatio-temporal distribution of cavitation activity during therapeutic ultrasound treatments, in addition to providing a valuable tool for real-time treatment monitoring and cavitation imaging. To enable meaningful correlation of cavitation activity with the safety and efficacy of associated bioeffects, a cavitation dose metric is required that is energy-preserving, device-independent and adequately normalized to the tissue volume being affected by the reported cavitation activity. We introduce cavitational radiated energy density (CRED), defined as the total energy of either narrowband or broadband radiated acoustic emissions over a volume, divided by that volume. We show that image blurring caused by the instrumentation-dependentpoint spread function introduces significant image artifacts and CRED errors when using existing conventional and adaptive beamformers, and propose a novel PAM algorithm that utilizes the Lucy–Richardson deconvolution (LRD) technique to compensate for the point-spread function and thus provide cavitation maps with reduced tail artifacts and improved energy estimates in a computationally efficient manner. Using a combination of modeling and experimental approaches, we demonstrate the quantitative significance of CRED and the potential of the LRD-PAM algorithm to enable energy-preserving quantitative real-time monitoring of cavitation-based therapies, independently of the instrumentation employed.

The ability to quantify and compare different cavitational treatments is essential given their accelerating clinical adoption. Passive acoustic mapping (PAM) can reconstruct quantitative maps of cavitational radiated energy density (CRED) in real time, providing a powerful platform for cavitation dosimetry. However, when compared to optical imaging, the relatively longer acoustic wavelengths and longitudinal nature of 2-D PAM result in greater variability and a more significant spatial impact of the point spread function (PSF). The associated imaging artifacts limit the spatial localization of cavitation dose and introduce errors in source energy estimation that vary across source parameters and between setups. Deconvolving with a constant PSF can reduce many of these errors and minimize the setup dependence of CRED estimates, but can introduce iteration-dependent errors in the estimated source location. This work explores the challenges of deconvolving images with large and variable PSFs using algorithms designed for optical applications and proposes strategies for mitigating errors. These approaches will be evaluated with respect to their quantitative performance, computation time, and robustness to noise both in silico and in vitro, with the aim of unlocking the full capabilities of PAM as a spatially resolved, energy preserving, setup independent platform for cavitational dosimetry.

As cavitation-based therapies continue to enter the clinic, there is growing demand for methods to quantify the energy released by cavitating bubbles. Passive Acoustic Mapping (PAM) can reconstruct the energy and distribution of cavitation from multi-sensor recordings of bubble emissions, but its accuracy is impaired in vitro by the aberrating presence of a sample container between the cavitating media (nuclei/cells/tissues) and detectors. To our knowledge, the effects of these vessels on PAM have never been studied. Additionally, the typical need for sterility and a large number of samples makes low cost essential for sample containers in cavitation experiments. Here, we characterize the effects of common laboratory vessels in the range 3–14 MHz via an acoustic reciprocity experiment, then describe the design and testing of a novel container with improved acoustic transparency. The new device reduced worst-case magnitude and phase errors by 13.7 dB and 6.6 radians respectively, compared to ordinary 2 ml centrifuge tubes. We will also present quantitative measures of container effects on PAM energy measurement and localization. The new containers are manufactured from 100 micron polymer film via vacuum forming, are quick and easy to make in any shape in a normal laboratory, and cost US$0.02 each.

Metastatic tumours in the brain now represent one of the leading causes of death from cancer. Current treatments are largely ineffective owing to the combination of late diagnosis and poor delivery of therapies across the blood-brain barrier (BBB). Conjugating magnetic resonance imaging (MRI) contrast agents with a monoclonal antibody for VCAM-1 (anti-VCAM1) has been shown to enable detection of micrometastases, two to three orders of magnitude smaller in volume than those currently detectable clinically. The aim of this study was to exploit this targeting approach to enable localised and temporary BBB opening at the site of early-stage metastases using functionalised microbubbles and ultrasound. Methods: Microbubbles functionalised with anti-VCAM1 were synthesised and shown to bind to VCAM-1-expressing cells in vitro. Experiments were then conducted in vivo in a unilateral breast cancer brain metastasis mouse model using Gadolinium-DTPA (Gd-DTPA) enhanced MRI to detect BBB opening. Following injection of Gd-DTPA and targeted microbubbles, the whole brain volume was simultaneously exposed to ultrasound (0.5 MHz, 10% duty cycle, 0.7 MPa peak negative pressure, 2 min treatment time). T1-weighted MRI was then performed to identify BBB opening, followed by histological confirmation via immunoglobulin G (IgG) immunohistochemistry. Results: In mice treated with targeted microbubbles and ultrasound, statistically significantly greater extravasation of Gd-DTPA and IgG was observed in the left tumour-bearing hemisphere compared to the right hemisphere 5 min after treatment. No acute adverse effects were observed. There was no investigation of longer term bioeffects owing to the nature of the study. Conclusion: The results demonstrate the feasibility of using targeted microbubbles in combination with low intensity ultrasound to localise opening of the BBB to metastatic sites in the brain. This approach has potential application in the treatment of metastatic tumours whose location cannot be established a priori with conventional imaging methods.

IR780 iodide is a lipophilic cation heptamethine dye that has emerged as a potential fluorescent probe for in vivo tumor imaging. Previous work has shown it to be a sono- and photo-sensitizer (sound- and light-activated molecule) suitable for use in sonodynamic therapy (SDT) due to its proposed ability to produce ROS when ‘activated' by ultrasound. This study evaluated IR780 iodide as an SDT agent in vitro using A549s, HeLas, and HeLa S3 cell lines, a broad range of ultrasound and light parameters, multiple ultrasound and light systems, and with and without cavitation nuclei. Through temporal uncoupling of ultrasound application and compound administration in vitro and in vivo, evaluation of cavitation-only related cell death, assessment of the dark toxicity of IR780 iodide, and development of positive controls for cell permeabilization (sonoporation), this study shows that dark toxicity and cavitation play an important role in IR780 SDT-induced cell death. This potentially explains the high levels of cell death observed for comparatively low concentrations of ROS. Additionally, this study proposes a standard set of controls for SDT mechanistic studies, which were used to further help study the mechanisms of other SDT drugs including Rose Bengal, 5-aminolevulinic acid, and indocyanine green.

Ultrasound-mediated drug delivery is typically performed using transducers with center frequencies <= 1 MHz to promote acoustic cavitation. Such frequencies are not commonly used for diagnostic ultrasound due to limited spatial resolution. Therefore, delivery and monitoring of therapeutic ultrasound typically requires two transducers to enable both treatment and imaging. This study investigates the feasibility of using a single commercial ultrasound imaging transducer operating at 5 MHz for both drug delivery and real-time imaging. We compared a single-transducer system (STS) at 5 MHz with a conventional dual-transducer system (DTS) using a 1.1 MHz therapeutic transducer and an imaging probe. in vitro experiments demonstrated that the STS could achieve comparable extravasation depth and area as the DTS, with higher drug deposition observed at 5 MHz. Additionally, extravasation patterns were influenced by peak negative pressure (PNP) and duty cycle, with the narrower beam width at 5 MHz offering potential advantages for targeted drug delivery. in vivo experiments in a murine bladder cancer model confirmed the efficacy of the STS for real-time imaging and drug delivery, with cavitation dose correlating with drug deposition. The results suggest that a single-transducer approach may enhance the precision and efficiency of ultrasound-mediated drug delivery, potentially reducing system complexity and cost.

Motivation: Lower back pain (LBP) is one of the leading causes of global disability. LBP and related radicular leg pain are closely linked to intervertebral disc (IVD) degeneration, which accounts for approximately 40% of the estimated 619 million LBP cases worldwide. Currently, there are two diametrically opposed treatment options for this condition: conservative physiotherapy to provide temporary relief, or major surgical intervention. Neither has proven to provide suitable long-term outcomes. Emerging strategies focus on injectable biomaterials to provide structural support and facilitate tissue repair, although they are still largely experimental and face several limitations, including limited integration with native tissue. Moreover, the implant formation mechanism may be depth-limited (light curing) or time-constrained (self-curing). Aim: The objective of this research is to demonstrate a new option to restore spinal function through the use of extracorporeal ultrasound to remotely trigger in situ implant formation on demand, such that the clinician can control the process with the timing and location of their choosing. The system concept centers around an implant precursor material consisting of an anionic polysaccharide matrix seeded with thermally sensitive liposomes. Ultrasound-mediated heating on the order of 3-5 degrees above normal body temperature triggers the release of crosslinking species from the liposomes, thereby initiating hydrogel formation. Methods: Candidate polysaccharide and liposome formulations were evaluated for their injectability, loading efficiency, and post-gelation mechanical properties. Ultrasound parameters (frequency, pressure, duty cycle) were optimized for targeted heating efficiency. Techniques for treatment process monitoring and control were independently investigated using thermometry and acoustic cavitation emissions. The material constructs and ultrasound protocols were used together in a series of proof-of-concept experiments using ex vivo bovine IVD specimens, with biomechanical analysis across three states: intact, degenerated, and after ultrasound-triggered implant formation.Results: Extensive testing revealed an optimized implant precursor material consisting of sodium alginate (1.5 wt/v%) seeded with calcium-loaded liposomes (157±9 nm) to enable hyperthermia-triggered release and glass microspheres (6 wt/v%) to ensure preferential ultrasound absorption for safe heating. No significant difference was found between hydrogels heated with an incubator or ultrasound, suggesting comparable calcium release between both methods. Optimal ultrasound parameters for precursor gel heating were found to be 0.95 MHz, 1.6 MPa (peak negative), and 87% duty cycle. Automated treatment control using temperature or cavitation emission measurements both were successfully implemented, with cavitation being preferable for non-invasive implementation. Proof of concept experiments indicated partial restoration of biomechanical function in ex vivo bovine IVDs, with implant material well-integrated into the disc tissue, and without material herniation.Conclusion: We have demonstrated the feasibility of ultrasound-guided hydrogel gelation in situ. These results offer promise for treating spinal disc degeneration, with continued refinement of materials and protocols essential for achieving robust in-disc efficacy.