Boiling Histotripsy Treatments Research Articles

A comparative study of histotripsy parameters for the treatment of fibrotic ex-vivo human benign prostatic hyperplasia tissue

Histotripsy is a noninvasive focused ultrasound therapy that mechanically fractionates tissue to create well-defined lesions. In a previous clinical pilot trial to treat benign prostatic hyperplasia (BPH), histotripsy did not result in consistent objective improvements in symptoms, potentially because of the fibrotic and mechanically tough nature of this tissue. In this study, we aimed to identify the dosage required to homogenize BPH tissue by different histotripsy modalities, including boiling histotripsy (BH) and cavitation histotripsy (CH). A method for histotripsy lesion quantification via entropy (HLQE) analysis was developed and utilized to quantify lesion area of the respective treatments. These data were correlated to changes in mechanical stiffness measured by ultrasound shear-wave elastography before and after treatment with each parameter set and dose. Time points corresponding to histologically observed complete lesions were qualitatively evaluated and quantitatively measured. For the BH treatment, complete lesions occurred with > = 30 s treatment time, with a corresponding maximum reduction in stiffness of −90.9 ± 7.2(s.d.)%. High pulse repetition frequency (PRF) CH achieved a similar reduction to that of BH at 288 s (−91.6 ± 6.0(s.d.)%), and low-PRF CH achieved a (−82.1 ± 5.1(s.d.)%) reduction in stiffness at dose > = 144 s. Receiver operating characteristic curve analysis showed that a > ~ 75% reduction in stiffness positively correlated with complete lesions observed histologically, and can provide an alternative metric to track treatment progression.

A Comparative Study of Histotripsy Parameters for the Treatment of Fibrotic ex-vivo Human Benign Prostatic Hyperplasia Tissue.

Histotripsy is a noninvasive focused ultrasound therapy that mechanically fractionates tissue to create well-defined lesions. In a previous clinical pilot trial to treat benign prostatic hyperplasia (BPH), histotripsy did not result in consistent objective improvements in symptoms, potentially because of the fibrotic and mechanically tough nature of this tissue. In this study, we aimed to identify the dosage required to homogenize BPH tissue by different histotripsy modalities, including boiling histotripsy (BH) and cavitation histotripsy (CH). A method for histotripsy lesion quantification via entropy (HLQE) analysis was developed and utilized to quantify lesion area of the respective treatments. These data were correlated to changes in mechanical stiffness measured by ultrasound shear-wave elastography before and after treatment with each parameter set and dose. Time points corresponding to histologically observed complete lesions were qualitatively evaluated and quantitatively measured. For the BH treatment, complete lesions occurred with >=30s treatment time, with a corresponding maximum reduction in stiffness of -90.9±7.2(s.d.)%. High pulse repetition frequency (PRF) CH achieved a similar reduction to that of BH at 288s (-91.6±6.0(s.d.)%), and low-PRF CH achieved a (-82.1±5.1(s.d.)%) reduction in stiffness at dose >=144s. Receiver operating characteristic curve analysis showed that a >~75% reduction in stiffness positively correlated with complete lesions observed histologically, and can provide an alternative metric to track treatment progression.

Boiling histotripsy exhibits anti-fibrotic effects in animal models of liver fibrosis

Liver fibrosis is a hallmark of chronic liver disease which could lead to liver cirrhosis or liver cancer. However, there is currently lack of a direct treatment for liver fibrosis. Boiling histotripsy (BH) is an emerging non-invasive high-intensity focused ultrasound technique that can be employed to mechanically destruct solid tumour at the focus via acoustic cavitation without significant adverse effect on surrounding tissue. Here, we investigated whether BH can mechanically fractionate liver fibrotic tissue thereby exhibiting an anti-fibrotic effect in an animal model of liver fibrosis. BH-treated penumbra and its identical lobe showed reduced liver fibrosis, accompanied by increased hepatocyte specific marker expression, compared to the BH-untreated lobe. Furthermore, BH treatment improved serological liver function markers without notable adverse effects. The ability of BH to reduce fibrosis and promote liver regeneration in liver fibrotic tissue suggests that BH could potentially be an effective and reliable therapeutic approach against liver fibrosis.

Histology-based quantification of boiling histotripsy outcomes via ResNet-18 network: Towards mechanical dose metrics

This work was focused on the newly developed ultrasonic approach for non-invasive surgery – boiling histotripsy (BH) – recently proposed for mechanical ablation of tissues using pulsed high intensity focused ultrasound (HIFU). The BH lesion is known to depend in size and shape on exposure parameters and mechanical properties, structure and composition of tissue being treated. The aim of this work was to advance the concept of BH dose by investigating quantitative relationships between the parameters of the lesion, pulsing protocols, and targeted tissue properties. A HIFU focus of a 1.5 MHz 256-element array driven by power-enhanced Verasonics system was electronically steered along the grid within 12 × 4 × 12 mm volume to produce volumetric lesions in porcine liver (soft, with abundant collagenous structures) and bovine myocardium (stiff, homogenous cellular) ex vivo tissues with various pulsing protocols (1–10 ms pulses, 1–15 pulses per point). Quantification of the lesion size and completeness was performed through serial histological sectioning, and a computer vision approach using a combination of manual and automated detection of fully fractionated and residual tissue based on neural network ResNet-18 was developed. Histological sample fixation led to underestimation of BH ablation rate compared to the ultrasound-based estimations, and provided similar qualitative feedback as did gross inspection. This suggests that gross observation may be sufficient for qualitatively evaluating the BH treatment completeness. BH efficiency in liver tissue was shown to be insensitive to the changes in pulsing protocol within the tested parameter range, whereas in bovine myocardium the efficiency increased with either increasing pulse length or number of pulses per point or both. The results imply that one universal mechanical dose metric applicable to an arbitrary tissue type is unlikely to be established. The dose metric as a product of the BH pulse duration and the number of pulses per sonication point (BHD1) was shown to be more relevant for initial planning of fractionation of collagenous tissues. The dose metric as a number of pulses per point (BHD2) is more suitable for the treatment planning of softer targets primarily containing cellular tissue, allowing for significant acceleration of treatment using shorter pulses.

Pilot ex vivo study on non-thermal ablation of human prostate adenocarcinoma tissue using boiling histotripsy

Focused ultrasound technologies are of growing interest for noninvasive ablation of localized prostate cancer (PCa). Here we present the results of the first case study evaluating the feasibility of non-thermal mechanical ablation of human prostate adenocarcinoma tissue using the boiling histotripsy (BH) method on ex vivo tissue. High intensity focused ultrasound field was generated using a 1.5-MHz custom-made transducer with nominal F#=0.75. A sonication protocol of 734 W acoustic power, 10-ms long BH-pulses, 30 pulses per focal spot, 1 % duty cycle, and 1 mm distance between single foci was tested in an ex vivo human prostate tissue sample containing PCa. The protocol used here has been successfully applied in the previous BH studies for mechanical disintegration of ex vivo prostatic human tissue with benign hyperplasia. BH treatment was monitored using B-mode ultrasound. Post-treatment histologic analysis demonstrated BH produced liquefaction of the targeted tissue volume. BH treated benign prostate parenchyma and PCa had similar tissue fractionation into subcellular fragments. The results of the study demonstrated that PCa tumor tissue can be mechanically ablated using the BH method. Further studies will aim on optimizing protocol parameters to accelerate treatment while maintaining complete destruction of the targeted tissue volume into subcellular debris.

Initial Assessment of Boiling Histotripsy for Mechanical Ablation of Ex Vivo Human Prostate Tissue.

Boiling histotripsy (BH) is a focused ultrasound technology that uses millisecond-long pulses with shock fronts to induce mechanical tissue ablation. The pulsing scheme and mechanisms of BH differ from those of cavitation cloud histotripsy, which was previously developed for benign prostatic hyperplasia. The goal of the work described here was to evaluate the feasibility of using BH to ablate fresh ex vivo human prostate tissue as a proof of principle for developing BH for prostate applications. Fresh human prostate samples (N=24) were obtained via rapid autopsy (<24 h after death, institutional review board exempt). Samples were analyzed using shear wave elastography to ensure that mechanical properties of autopsy tissue were clinically representative. Samples were exposed to BH using 10- or 1-ms pulses with 1% duty cycle under real-time B-mode and Doppler imaging. Volumetric lesions were created by sonicating 1-4 rectangular planes spaced 1 mm apart, containing a grid of foci spaced 1-2 mm apart. Tissue then was evaluated grossly and histologically, and the lesion content was analyzed using transmission electron microscopy and scanning electron microscopy. Observed shear wave elastography characterization of ex vivo prostate tissue (37.9 ± 22.2 kPa) was within the typical range observed clinically. During BH, hyperechoic regions were visualized at the focus on B-mode, and BH-induced bubbles were also detected using power Doppler. As treatment progressed, hypoechoic regions of tissue appeared, suggesting successful tissue fractionation. BH treatment was twofold faster using shorter pulses (1 ms vs. 10 ms). Histological analysis revealed lesions containing completely homogenized cell debris, consistent with histotripsy-induced mechanical ablation. It was therefore determined that BH is feasible in fresh ex vivo human prostate tissue producing desired mechanical ablation. The study supports further work aimed at translating BH technology as a clinical option for prostate ablation.

Quantitative Assessment of Boiling Histotripsy Progression Based on Color Doppler Measurements.

Boiling histotripsy (BH) is a mechanical tissue liquefaction method that uses sequences of millisecond-long high intensity focused ultrasound (HIFU) pulses with shock fronts. The BH treatment generates bubbles that move within the sonicated volume due to acoustic radiation force. Since the velocity of the bubbles and tissue debris is expected to depend on the lesion size and liquefaction completeness, it could provide a quantitative metric of the treatment progression. In this study, the motion of bubble remnants and tissue debris immediately following BH pulses was investigated using high-pulse repetition frequency (PRF) plane-wave color Doppler ultrasound in ex vivo myocardium tissue. A 256-element 1.5 MHz spiral HIFU array with a coaxially integrated ultrasound imaging probe (ATL P4-2) produced 10 ms BH pulses to form volumetric lesions with electronic beam steering. Prior to performing volumetric BH treatments, the motion of intact myocardium tissue and anticoagulated bovine blood following isolated BH pulses was assessed as two limiting cases. In the liquid blood the velocity of BH-induced streaming at the focus reached over 200 cm/s, whereas the intact tissue was observed to move toward the HIFU array consistent with elastic rebound of tissue. Over the course of volumetric BH treatments tissue motion at the focus locations was dependent on the axial size of the forming lesion relative to the corresponding size of the HIFU focal area. For axially small lesions, the maximum velocity after the BH pulse was directed toward the HIFU transducer and monotonically increased over time from about 20-100 cm/s as liquefaction progressed, then saturated when tissue was fully liquefied. For larger lesions obtained by merging multiple smaller lesions in the axial direction, the high-speed streaming away from the HIFU transducer was observed at the point of full liquefaction. Based on these observations, the maximum directional velocity and its location along the HIFU propagation axis were proposed and evaluated as candidate metrics of BH treatment completeness.

Mechanical damage thresholds for hematomas near gas-containing bodies in pulsed HIFU fields

Objective. Boiling histotripsy (BH) is a novel high intensity focused ultrasound (HIFU) application currently being developed for non-invasive mechanical fractionation of soft tissues and large hematomas. In the context of development of BH treatment planning approaches for ablating targets adjacent to gas-containing organs, this study aimed at investigation of the ultrasound pressure thresholds of atomization-induced damage to the tissue-air interface and correlation of the danger zone dimensions with spatial structure of nonlinear HIFU field parameters. Approach. A flat interface with air of freshly clotted bovine blood was used as an ex vivo model due to its homogenous structure and higher susceptibility to ultrasound-induced mechanical damage compared to soft tissues. Three 1.5 MHz transducers of different F-numbers (0.77, 1 and 1.5) were focused at various distances before or beyond a flat clot surface, and a BH exposure was delivered either at constant, high-amplitude output level, or at gradually increasing level until a visible damage to the clot surface occurred. The HIFU pressure field parameters at the clot surface were determined through a combination of hydrophone measurements in water, forward wave propagation simulation using ‘HIFU beam’ software and an image source method to account for the wave reflection from the clot surface and formation of a standing wave. The iso-levels of peak negative pressure in the resulting HIFU field were correlated to the outlines of surface erosion to identify the danger zone around the BH focus. Main results. The outline of the danger zone was shown to differ from that of a typical BH lesion produced in a volume of clot material. In the prefocal area, the zone was confined within the 4 MPa contour of the incident peak-to-peak pressure; within the main focal lobe it was determined by the maximum BH lesion width, and in the postfocal area—by the transverse size of the focal lobe and position of the first postfocal pressure axial null. Significance. The incident HIFU pressure-based danger zone boundaries were outlined around the BH focus and can be superimposed onto in-treatment ultrasound image to avoid damage to adjacent gas-containing bodies.

Investigation of the long-term healing response of the liver to boiling histotripsy treatment in vivo

Boiling histotripsy (BH) is a promising High-Intensity Focused Ultrasound technique that can be employed to mechanically fractionate solid tumours. Whilst studies have shown the feasibility of BH to destroy liver cancer, no study has reported on the healing process of BH-treated liver tissue. We therefore extensively investigated the evolution of the healing response of liver to BH in order to provide an insight into the healing mechanisms. In the present study, 14 Sprague Dawley rats underwent the BH treatment and were sacrificed on days 0, 3, 7, 14, and 28 for morphological, histological, serological and qPCR analyses. The area of the treated region was 1.44 cm2 (1.2 cm × 1.2 cm). A well-defined BH lesion filled with coagulated blood formed on day 0. A week after the treatment, fibroblast activation was induced at the treatment site, leading to the formation of extracellular matrix structure (ECM). The ECM was then disrupted for 7 to 28 days. Regenerated normal hepatocytes and newly formed blood vessels were found within the BH region with the absence of hepatic fibrosis. No significant morphological, histological and genetic changes around the BH lesion occurred. These results suggest that BH could be a safe and promising therapeutic tool for treating solid tumours without inducing any significant adverse effect such as the formation of liver fibrosis.

MP04-02 ANALYSIS OF EX-VIVO HUMAN BPH TISSUE TREATED BY HISTOTRIPSY: DOSE AND PARAMETER EFFECTS

You have accessJournal of UrologyCME1 May 2022MP04-02 ANALYSIS OF EX-VIVO HUMAN BPH TISSUE TREATED BY HISTOTRIPSY: DOSE AND PARAMETER EFFECTS Zorawar Singh, Yashwanth N. Kumar, Yak-Nam Wang, Adam Maxwell, Matthew Bruce, Rishi Sekar, Eli Vlaisavljevich, Lawrence D. True, and George R. Schade Zorawar SinghZorawar Singh More articles by this author , Yashwanth N. KumarYashwanth N. Kumar More articles by this author , Yak-Nam WangYak-Nam Wang More articles by this author , Adam MaxwellAdam Maxwell More articles by this author , Matthew BruceMatthew Bruce More articles by this author , Rishi SekarRishi Sekar More articles by this author , Eli VlaisavljevichEli Vlaisavljevich More articles by this author , Lawrence D. TrueLawrence D. True More articles by this author , and George R. SchadeGeorge R. Schade More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000002521.02AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Histotripsy is a promising alternative to more invasive procedures for the treatment of benign prostatic hyperplasia (BPH). However, in a phase 1 clinical trial, histotripsy failed to provide objective improvements in urinary function. One explanation may be that the clinical trial pulse parameters, optimized in a canine model, do not translate to more fibrotic human prostates. We aimed to evaluate the efficacy of different histotripsy parameter settings to identify a more efficacious exposure in ex-vivo human BPH tissue (EVHP). METHODS: Deidentified EVHP tissue specimens (n=17) were treated with an 18 element 700kHz cavitation histotripsy transducer, which approximated the device used in the prior clinical trial, and a 1.5MHz boiling histotripsy (BH) transducer. Three different histotripsy exposures were compared: 1) a high pulse repetition frequency (PRF) (approximating the clinical system cavitation cloud histotripsy [CCH]), 2) low PRF CCH and 3) 1Hz PRF BH. Treatment effectiveness was evaluated by measuring pre vs. post treatment Young’s modulus through shear wave elastography (SWE) and histologic analysis (H&E, Masson’s Trichome) of the targeted area. The dose was standardized at 30s/mm and escalated as required to note a significant change in SWE. RESULTS: At the given dose of 30s/mm, BH treatments resulted in a significant 40.1% reduction in SWE Young’s modulus (Figure 1A) p < 0.001. At this dose, neither of the CCH parameter settings (500Hz and 10Hz) resulted in a significant reduction in the Young’s modulus with % changes of differences of 3.1% (p = .83) and -5% (p = .08), respectively. For CCH settings, greater exposure times (up to 90s) did result in significant stiffness reduction of 12.5% (p = 0.04) and 41.1% (p < 0.001) for 500Hz and 10Hz parameter settings, respectively. On histology (Figure 1B), at 30s/mm, the CCH parameter settings resulted in minimal intermittent damage while BH treatments caused near complete ablation. At increasing doses, CCH was also capable of causing complete liquefactive cavity formation. CONCLUSIONS: BH parameters applied in this study resulted in more complete ablation of EVH tissue in comparison to CCH parameters. Additionally, histotripsy treatment parameters similar to those applied with a previous clinical system did not produce consistent ablation of EVHP possibly explaining trial results. Source of Funding: None © 2022 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 207Issue Supplement 5May 2022Page: e56 Advertisement Copyright & Permissions© 2022 by American Urological Association Education and Research, Inc.MetricsAuthor Information Zorawar Singh More articles by this author Yashwanth N. Kumar More articles by this author Yak-Nam Wang More articles by this author Adam Maxwell More articles by this author Matthew Bruce More articles by this author Rishi Sekar More articles by this author Eli Vlaisavljevich More articles by this author Lawrence D. True More articles by this author George R. Schade More articles by this author Expand All Advertisement PDF DownloadLoading ...

The use of real-time passive cavitation imaging for monitoring boiling histotripsy treatments

Boiling histotripsy (BH) is a high intensity focused ultrasound method that induces mechanical tissue liquefaction and relies on the formation of a mm-sized vapor bubble at the focus. One challenge of transcutaneous BH is to confirm the presence of such bubble and to differentiate it from prefocal cavitation. Strong prefocal cavitation can shield the focus, prevent initiation of boiling, and lead to unsuccessful treatment, although an echogenic region would still be present on B-mode ultrasound images. Here, a methodology for real-time monitoring of different bubble activity during BH treatment is proposed. Custom BH systems of 1 and 1.5 MHz with inline ultrasound imaging probes connected to a Verasonics V1 system were used for BH treatments in pigs in vivo and in ex vivo tissues. Backscattered signals arriving to the imaging probe were recorded during the 10 ms BH pulses delivered at 1 Hz and passive cavitation imaging (PCI) reconstruction was performed and displayed on top of the B-mode images to identify and classify the regions of bubble activity. Spectral analysis of the PCI signals promoting prefocal cavitation and lack thereof was performed to identify the spectral features corresponding to prefocal cavitation and vapor bubble formation. [Work supported by NIH R01EB007643, R01GM122859, and R01EB25187.]

Doppler ultrasound observations and metrics of boiling histotripsy treatment progression

Boiling histotripsy (BH) is a mechanical tissue fractionation method that uses millisecond-long nonlinear HIFU pulses with shock fronts. The BH treatment generates bubbles and tissue debris that move within the liquified lesion due to acoustic radiation force and streaming. Since the velocity of the debris is expected to depend on the lesion size and fractionation completeness, it could provide a quantitative metric of the treatment progression. In this study, the motion of bubble remnants and tissue debris immediately following BH pulses was investigated by high-pulse repetition frequency (PRF) plane-wave color Doppler ultrasound (PRF 9–15 kHz, 50–100 pulses) in ex vivo bovine myocardium and porcine liver. A 256-element 1.5 MHz spiral HIFU array with a coaxially integrated ultrasound imaging probe (ATL P4-2) produced the 10ms BH pulses to create a volumetric lesion in tissue. As tissue liquefaction progressed, the velocity was observed to be directed towards the HIFU transducer, monotonically increased, and then saturated at about 1 m/s. Further, when the lesion was fully liquefied and its axial size was comparable to that of the HIFU focal spot, the motion changed direction to the opposite, with velocity independent of tissue type, consistent with streaming. [Work supported by NIH R01EB7643, R01GM122859, R01EB025187, and RSF20-12-00145.]

Partial Respiratory Motion Compensation for Abdominal Extracorporeal Boiling Histotripsy Treatments With a Robotic Arm.

Extracorporeal boiling histotripsy (BH), a noninvasive method for mechanical tissue disintegration, is getting closer to clinical applications. However, the motion of the targeted organs, mostly resulting from the respiratory motion, reduces the efficiency of the treatment. Here, a practical and affordable unidirectional respiratory motion compensation method for BH is proposed and evaluated in ex vivo tissues. The BH transducer is fixed on a robotic arm following the motion of the skin, which is tracked using an inline ultrasound imaging probe. In order to compensate for system lags and obtain a more accurate compensation, an autoregressive motion prediction model is implemented. BH pulse gating is also implemented to ensure targeting accuracy. The system is then evaluated with ex vivo BH treatments of tissue samples undergoing motion simulating breathing with the movement of amplitudes between 5 and 10 mm, the frequency between 16 and 18 breaths/min, and a maximum speed of 14.2 mm/s. Results show a reduction of at least 89% of the value of the targeting error during treatment while only increasing the treatment time by no more than 1%. The lesions obtained by treating with the motion compensation were close in size and affected area to the no-motion case, whereas lesions obtained without the compensation were often incomplete and had larger affected areas. This approach to motion compensation could benefit extracorporeal BH and other histotripsy methods in clinical translation.

A Prototype Therapy System for Boiling Histotripsy in Abdominal Targets Based on a 256-Element Spiral Array.

Boiling histotripsy (BH) uses millisecond-long ultrasound (US) pulses with high-amplitude shocks to mechanically fractionate tissue with potential for real-time lesion monitoring by US imaging. For BH treatments of abdominal organs, a high-power multielement phased array system capable of electronic focus steering and aberration correction for body wall inhomogeneities is needed. In this work, a preclinical BH system was built comprising a custom 256-element 1.5-MHz phased array (Imasonic, Besançon, France) with a central opening for mounting an imaging probe. The array was electronically matched to a Verasonics research US system with a 1.2-kW external power source. Driving electronics and software of the system were modified to provide a pulse average acoustic power of 2.2 kW sustained for 10 ms with a 1-2-Hz repetition rate for delivering BH exposures. System performance was characterized by hydrophone measurements in water combined with nonlinear wave simulations based on the Westervelt equation. Fully developed shocks of 100-MPa amplitude are formed at the focus at 275-W acoustic power. Electronic steering capabilities of the array were evaluated for shock-producing conditions to determine power compensation strategies that equalize BH exposures at multiple focal locations across the planned treatment volume. The system was used to produce continuous volumetric BH lesions in ex vivo bovine liver with 1-mm focus spacing, 10-ms pulselength, five pulses/focus, and 1% duty cycle.

Pilot in vivo studies on transcutaneous boiling histotripsy in porcine liver and kidney

Boiling histotripsy (BH) is a High Intensity Focused Ultrasound (HIFU) method for precise mechanical disintegration of target tissue using millisecond-long pulses containing shocks. BH treatments with real-time ultrasound (US) guidance allowed by BH-generated bubbles were previously demonstrated ex vivo and in vivo in exposed porcine liver and small animals. Here, the feasibility of US-guided transabdominal and partially transcostal BH ablation of kidney and liver in an acute in vivo swine model was evaluated for 6 animals. BH parameters were: 1.5 MHz frequency, 5–30 pulses of 1–10 ms duration per focus, 1% duty cycle, peak acoustic powers 0.9–3.8 kW, sonication foci spaced 1–1.5 mm apart in a rectangular grid with 5–15 mm linear dimensions. In kidneys, well-demarcated volumetric BH lesions were generated without respiratory gating and renal medulla and collecting system were more resistant to BH than cortex. The treatment was accelerated 10-fold by using shorter BH pulses of larger peak power without affecting the quality of tissue fractionation. In liver, respiratory motion and aberrations from subcutaneous fat affected the treatment but increasing the peak power provided successful lesion generation. These data indicate BH is a promising technology for transabdominal and transcostal mechanical ablation of tumors in kidney and liver.

Real-time imaging and control of boiling histotripsy lesion formation using Doppler ultrasound

In boiling histotripsy (BH), millisecond-long HIFU bursts with shocks are used to mechanically liquefy tissue. Most BH studies have been performed under real-time B-mode ultrasound imaging of bubbles generated at the treatment site. However, such approach does not allow quantitative assessment of the degree of tissue liquefaction. In addition, aberration and clutter often limit the echogenicity introduced by bubbles. In this work, plane wave Doppler followed by B-mode imaging were implemented immediately after each BH pulse to visualize residual bubble motion during volumetric BH treatments in ex vivo bovine liver. The changes in the Doppler power and speed were used as metrics to assess the degree of tissue liquefaction. An ATL P6-3 imaging probe connected to a Verasonics Ultrasound Engine (VUE) was mounted in the central opening of a 256-element 1.5 MHz HIFU array driven by another VUE with 1.2 kW external power source enhanced by a capacitor bank. The proposed imaging sequence improved BH targeting and bubble visualization. As tissue liquefaction progressed from partial to full (single versus five BH pulses per focal spot), both Doppler power and residual bubble velocity directed away from the transducer increased (7-fold and 10 cm/s to 20 cm/s, correspondingly) and then saturated. [Work supported by NIH R01EB7643.]

Boiling Histotripsy Ablation of Renal Cell Carcinoma in the Eker Rat Promotes a Systemic Inflammatory Response

Boiling histotripsy (BH) is an experimental focused ultrasound technique that produces non-thermal mechanical ablation. We evaluated the feasibility, short-term histologic effects and the resulting acute inflammatory response to BH ablation of renal cell carcinoma (RCC) in the Eker rat. Genotyped Eker rats were monitored for de novo RCCs with serial ultrasound (US) imaging. When tumors were ≥8 mm, rats underwent ultrasound-guided extracorporeal ablation of the tumor with BH, a pulsed focused US technique that produces non-thermal mechanical ablation of targeted tissues, or a sham US procedure. Treatments targeted approximately 50% of the largest RCC with a margin of normal kidney. BH treated rats were euthanized at 1 (n = 4) or 48 (n = 4) h, and sham patients (n = 4) at 48 h. Circulating plasma cytokine levels were assessed with multiplex assays before and at 0.25, 1, 4, 24 and 48 h following treatment. Kidneys were collected and processed for histologic assessment, immunohistochemistry and intrarenal cytokine concentration measurements. For statistical analysis Student's t-test was used. US-guided BH treatment was successful in all animals, producing hypoechoic regions within the targeted volume consistent with BH treatment effect. Grossly, regions of homogenized tissue were apparent with evidence of focal intra-parenchymal hemorrhage. Histologically, BH produced a sharply demarcated region of homogenized tumor and non-tumor tissue containing acellular debris. BH treatment was associated with significantly increased relative concentration of plasma TNF versus sham treatment (p < 0.05) and transient elevations in high-mobility group box 1 (HMGB1), IL-10 and IL-6 consistent with acute inflammatory response to trauma. Intrarenal cytokine concentrations followed the same trend. At 48 h, enhanced infiltration of CD8+ T cells was observed by immunohistochemistry in both the treated and un-treated contralateral RCC/kidneys in BH-treated animals versus sham treatment. BH treatment was well tolerated with transient gross hematuria and a perinephric hematoma developing in one subject each. The study demonstrates the feasibility of BH ablation of de novo RCC and suggests activation of the acute inflammatory cascade following treatment that appears to stimulate CD8+ T cell infiltration of both treated and untreated tumors. Longer duration chronic studies are ongoing to characterize the longevity and robustness of this response.

Impact of MR-guided boiling histotripsy in distinct murine tumor models

Interest in mechanical high intensity focused ultrasound (HIFU) ablation is rapidly growing. Boiling histotripsy (BH) is applied for mechanical fragmentation of soft tissue into submicron fragments with limited temperature increase using the shock wave and cavitation effects of HIFU. Research on BH has been largely limited to ex vivo experiments. As a consequence, the in vivo pathology after BH treatment and the relation to preexistent tissue characteristics are not well understood. This study reports on in vivo MR guided BH treatment, either with 100 or 200 pulses per focal spot, in three different subcutaneous mouse tumor models: a soft-tissue melanoma (B16OVA), a compact growing thymoma (EL4), and a highly vascularized neuroblastoma (9464D). Extensive treatment evaluation was performed using MR imaging followed by histopathology 2h after treatment.T2 weighted MRI allowed direct in vivo visualization of the BH lesions in all tumor models. The 100-pulse treated area in the B16OVA tumors was larger than the predicted treatment volume (500±10%). For the more compact growing EL4 and 9464D tumors this was 95±13% and 55±33%, respectively. Histopathology after the 100-pulse treatment revealed completely disintegrated lesions in the treated area with sharp borders in the compact EL4 and 9464D tumors, while for B16OVA tumors the lesion contained a mixture of discohesive (partly viable) clusters of cells, micro-vessel remainings, and tumor cell debris. The treatment of B16OVA with 200 pulses increased the fragmentation of tumor tissue. In all tumor types only micro-hemorrhages were detected after ablation (slightly higher after 200-pulse treatment for the highly vascularized 9464D tumors). Collagen staining revealed that the collagen fibers were to a greater or lesser extent still intact and partly clotted together near the lesion border in all tumor models.In conclusion, this study reveals effective mechanical fragmentation of different tumor types using BH without major hemorrhages. However, treatment settings may need to be adjusted to the tissue characteristics for optimal tissue fragmentation.

Pathology and immune effects of magnetic resonance imaging-guided Boiling Histotripsy in murine tumor models

In situ tumor ablation techniques are successfully applied for the destruction of local, often inoperable tumor masses. Following ablation tumor antigens become instantly available for immune cells, but systemic abscopal effects have only occasionally been reported after ablation monotherapy. Which ablation technique combines optimal local destruction with effective antigen release for induction of anti-tumor immunity is largely unknown. We study non-invasive MRI-guided high intensity focused ultrasound-ablation (MRgHiFU) in murine tumor models for local destruction by heating or mechanical disruption using Boiling histotripsy (BH). BH mechanically fragmentizes soft tissue into submicron fragments that are absorbed as part of a physiological healing response. BH treatment was performed using a MR compatible animal HIFU system (Image Guided Therapy, Pessac, France) with a 3 MHz transducer (focal spot size 0.5 x 0.5 x 2.0 mm). A 7T animal MR scanner was used for treatment guidance and evaluation. Here, we w...

Pilot in vivo studies on transcutaneous boiling histotripsy in porcine liver and kidney

Boiling histotripsy (BH) uses millisecond-long pulses of focused ultrasound (FUS) waves with shocks to mechanically homogenize tissue. Here we report a pilot in vivo acute study on transcutaneous volumetric BH ablation of porcine liver and kidney. BH treatment was administered using a 1.5 MHz FUS transducer operating at peak electric power ranging from 0.6 to 4 kW with B-mode ultrasound guidance using an imaging probe mounted coaxially. Sonication protocols delivered 5-30 pulses of 1-10 ms duration and 1% duty factor to focal points spaced 1-1.5 mm apart in a rectangular grid with 5-15 mm linear dimensions. Well-demarcated volumetric BH lesions were successfully generated in both liver and kidney without respiratory gating. The treatment was accelerated 6-fold without affecting the quality of tissue homogenization by using shorter duration BH pulses of larger peak power. These data indicate that transcutaneous volumetric renal and hepatic ablations are feasible in the in vivo porcine model. Studies are ongoing to optimize treatment parameters with the goal of clinical BH ablation of renal and hepatic masses. [This work was supported by NIH R01 EB7643, K01 EB015745, NSBRI through NASA NCC 9-58, RFBR 16-02-00653, and Urology Care Foundation.]