New medicine application of local injection acoustic response stent to enhancement of ultrasound histotripsy

Abstract

The invention discloses a new medicine application of a local injection acoustic response stent in enhancing ultrasound histotripsy. The acoustic response scaffold (ARS) comprises a scaffold material, fluorocarbon gas or liquid fluorocarbon particles with liquid-vapor phase change capacity, and microbubble cloud in the scaffold generates a strong cavitation ablation treatment effect in a target area under the action of low-frequency and low-sound-pressure ultrasound. Fluorocarbon gas or liquid fluorocarbon particles contained in the ARS form microbubble cloud and ideal cavitation nuclei under low-frequency low-sound-pressure ultrasonic excitation, inertia cavitation resonance is achieved under the ultrasonic action, and then cavitation mechanical damage with local tissue controllable strength is generated. The low-intensity ultrasonic histotripsy has the characteristics of low sound pressure, low sound intensity, high targeting property, small tissue damage outside a target region, simplicity in instrument manufacturing and the like, and the targeting property and the safety of treatment are improved. The method does not need very accurate ultrasonic focusing, reduces the operation difficulty, and has application prospects in multiple medical fields such as prostatic hyperplasia, tumor ablation and tumor immunotherapy.

Description

Technical Field

[0001] The present invention relates to the technical field of acoustically-responsive scaffolds (ARS) mechanically ablating diseased tissues through ultrasonic cavitation under pulsed, low-frequency, low-sound-pressure ultrasonic excitation. The ARS can be applied to the treatment of diseased tissues such as malignant solid tumors and prostate hyperplasia, and belongs to the technical field of non-invasive ultrasonic treatment. Background Art

[0002] Ultrasonic tissue ablation (Histotripsy) is a non-invasive, non-ionizing radiation and non-thermal ultrasonic mechanical ablation technology. This method is based on the generation of a bubble cloud in the tissue under the action of a continuous short pulse of 4-5 MPa, and then the extremely high sound pressure pulse of 15-30 MPa is emitted at intervals to stimulate the microbubble cloud to undergo strong inertial cavitation, and the mechanical effects released by cavitation (such as shock waves, microjets, etc.) are used to destroy and decompose tissue. Compared with HIFU (High Intensity Focused Ultrasound), Histotripsy is emitted in a pulsed manner, and the working duty cycle is generally much lower than 1%, so there is no thermal effect and thermal effect damage, and tissue liquefaction and necrosis are more easily metabolized and absorbed by the human body. Studies have shown that Histotripsy has great potential in the treatment of tumors, thrombolysis, benign prostatic hyperplasia (BPH), and softening aortic valve calcification. Especially for the ablation treatment of tumors, a clinical trial of Histotripsy treatment for patients with multifocal primary and metastatic liver tumors showed that Histotripsy can safely target and ablate liver tumor tissue. However, the main disadvantage of Histotripsy is that it requires a large array transducer as a focusing surface and extremely high peak negative pressure (Peak Negative Pressure, or PNP, about 15-30MPa). In fact, it is difficult to focus such a high sound pressure in the body through the narrow human acoustic window. Bone gas obstruction may lead to insufficient sound pressure, and respiratory movement may cause focus deviation. These unfavorable factors may lead to incomplete ablation or mechanical damage to important surrounding structures, causing complications.

[0003] The necessary conditions for Histotripsy mechanical ablation of tissues are to stimulate the human body to produce microbubble clouds and destroy them by cavitation through continuous alternation of low to medium sound pressure and extremely high peak negative pressure pulses. In order to increase the safety, stability and complexity of instrument manufacturing, it is necessary to reduce the treatment sound pressure and artificially create a microbubble cloud with a long duration. Therefore, in our early stage, we injected liquid fluorocarbon compounds through local puncture and ultrasonically stimulated the formation of microbubble clouds, which can greatly reduce the peak negative pressure required for tissue damage. Our previous studies have found that by injecting perfluoropentane phase change microparticle emulsion through local puncture, under the action of low frequency (f) and low sound pressure focused ultrasound (f = 596KHz, PNP ≤ 2MPa), controllable cavitation mechanical ablation of liver tissue can be produced. This technology combines with exogenous microbubble clouds for cavitation ablation, without the need for precise focusing. The cavitation threshold inside the microbubble cloud is extremely low and the cavitation effect is strong. In the non-target area outside the microbubble cloud, the cavitation effect is weak, and there is almost no tissue destruction and ablation effect. Therefore, the targeting and safety of ultrasound treatment are greatly improved. Although this technology requires a sound pressure as low as 2MPa, we found during the experiment that the droplet microemulsion has high fluidity, which will affect the distribution and range of the microbubble cloud. If the microbubble cloud exceeds the target area, it may cause damage to tissues or important structures outside the target area.

[0004] Our previous research found that low-frequency, low-sound-pressure focused ultrasound stimulated the vaporization of perfluoropentane phase-change microparticles and sustained cavitation resonance, which could cause mechanical damage to canine prostate and liver tissues. However, there are also defects such as the high fluidity of perfluoropentane phase-change droplets, which cannot form a high-concentration microbubble cloud in a confined target area.

[0005] The patent application with the publication number CN116036274A discloses a therapeutic use of fluorocarbon phase change microparticles for local ultrasonically stimulated vaporization and cavitation to produce tissue damage, wherein a liquid fluorocarbon compound is wrapped with a membrane material to form a suspension, and then dispersed into an emulsified liquid state after mechanical or ultrasonic oscillation; the membrane material includes lipids, human albumin and sugars, and a fluorocarbon phase change microparticle is prepared; the fluorocarbon phase change microparticles located in the diseased tissue undergo gas-liquid phase change / vaporization under the stimulation of pulsed, low-frequency, low-sound pressure focused ultrasound, and the resulting microbubble cloud / group undergoes microbubble cavitation resonance under the action of continuous low-frequency, low-sound pressure focused ultrasound, resulting in cavitation mechanical damage to the local tissue, thereby producing an ablation therapeutic effect on the diseased tissue. However, the simple fluorocarbon phase change microparticle emulsion in this technology has strong fluidity and cannot be well confined to the tissue target area. Under the action of ultrasound, a diffuse microbubble cloud is formed, which diffuses outside the target area, causing focus deviation to produce incomplete ablation or damage to important structures such as surrounding large blood vessels.

[0006] Therefore, in order to further improve the targeting and safety of ultrasonic tissue ablation, it is critical to reduce the fluidity of fluorocarbon phase change emulsion so that the microbubble cloud can be better confined to the target tissue while ensuring its effective vaporization and cavitation resonance effect. Summary of the invention

[0007] In order to solve the above problems in the prior art, the present invention provides an acoustically-responsive scaffold (ARS) that can confine fluorocarbon particles to a target area and has a good therapeutic effect. The ARS can vaporize and cavitate to produce tissue damage under low-frequency, low-sound-pressure, pulsed ultrasonic excitation.

[0008] In view of this, the technical solution adopted by the present invention is that ARS includes a support material and fluorocarbon particles, the support material wraps the fluorocarbon particles to form an acoustic response support containing fluorocarbon particles, and the volume ratio of the support material solution and the fluorocarbon particle emulsion is 5 to 10:1. ARS is an acoustic support containing air, or fluorocarbon gas, or sulfur hexafluoride gas or liquid fluorocarbon compound; the support material is used as a liquid support material, and the interior contains air, fluorocarbon gas microbubbles, sulfur hexafluoride gas microbubbles, liquid fluorocarbon compound or a mixture of the above gases or compounds, forming an acoustic response support containing the above gas or liquid fluorocarbon compound particles.

[0009] In some embodiments of the present invention, the scaffold material is selected from one or more of sodium alginate, fibrinogen, F127 (polyethylene glycol-polypropylene glycol-polyethylene glycol triblock copolymer), poly N-isopropylacrylamide-polyethylene glycol block copolymer, gelatin, collagen, cellulose, silk protein, keratin, heparin, whey protein, zein, PCLA-PEG-PCLA [poly(ε-caprolactone-co-lactide)-b-poly(ethylene glycol)-bpoly(ε-caprolactone-co-lactide)], polycaprolactone-polylactic acid copolymer, chitin, chitosan, polyvinyl alcohol, polyacrylamide, sodium polyacrylate, hydroxyapatite, methacryloyl hyaluronic acid, methacryloyl chitosan and β-amino ester.

[0010] In some embodiments of the present invention, the air, fluorocarbon gas, sulfur hexafluoride gas or liquid fluorocarbon compound contained in the acoustic response bracket includes: Octafluoropropane (C3F8), Sulfurfluoride (SF6), Perfluorobutane (C4F10), Perfluoropentane (C5F12), Nonafluoro (trifluoromethyl) cyclopentane (C6F12), Perfluorohexane (C6F14), Perfluoroheptane (C7F16), Perfluorooctane (C8F18), etc.; the gas or liquid fluorocarbon compound contained in the acoustic response bracket can be one of the above gases or liquid fluorocarbon compounds, or a mixture of multiple gases or liquid fluorocarbon compounds.

[0011] The present invention also provides a new use of the ARS in preparing a novel low-intensity ultrasonic tissue destruction medicine. The stent enters the human body through local puncture injection and is distributed in the target area.

[0012] In the above-mentioned use of the present invention, ARS needs to be combined with low-frequency, low-sound pressure focused ultrasound to achieve ablative effect. The ARS forms a microbubble cloud or undergoes phase change vaporization to form a microbubble cloud (the microbubble particle size in the microbubble cloud is 1-100 microns) under low-frequency, low-sound pressure ultrasonic excitation, and then inertial cavitation resonance occurs under continuous low-frequency ultrasonic irradiation, and then cavitation mechanical damage with controllable damage degree of local tissue is produced, thereby producing ablative effect on diseased tissue.

[0013] In some embodiments of the present invention, the ultrasonic emission mode of the low-frequency, low-sound pressure ultrasound can be planar ultrasound or focused ultrasound; the ultrasonic emission frequency is in the range of 30kHz-800kHz; the ultrasonic peak negative pressure is in the range of 0.5MPa-8.0MPa; the ultrasonic single pulse width is in the range of 5-200 cycles, and the working duty cycle is less than 5%.

[0014] Furthermore, the low-frequency ultrasound has an emission frequency in the range of 30kHz-800kHz, and can be single-frequency ultrasound, or dual-frequency or multi-frequency combined ultrasonic energy emitted simultaneously.

[0015] In some embodiments of the present invention, the ARS can produce ablation treatment effects such as mechanical damage to human tissue under low-frequency, low-sound pressure ultrasonic excitation, including: mechanical damage and ablation of various pathological tissues such as prostate hyperplasia tissue and malignant solid tumor tissue.

[0016] The present invention also provides a method for preparing ARS, comprising:

[0017] (1) preparing a fluorocarbon microparticle emulsion;

[0018] (2) preparing a scaffold material solution, using physiological saline as a solvent to prepare a sodium alginate solution with a concentration of 20-25%;

[0019] (3) Mix the prepared sodium alginate solution and fluorocarbon microparticle emulsion in a volume ratio of 10:1 and shake thoroughly.

[0020] The present invention uses a stent to wrap the perfluoropentane droplet particles to control their fluidity, so that they are better confined to the target tissue and vaporize under ultrasound to form a high-concentration microbubble cloud and continuous cavitation resonance, further improving their targeting, safety and cavitation intensity.

[0021] The short-term test of the present invention found that when the sound pressure was 4.1MPa, the two-dimensional ultrasound showed that the concentration of microbubble clouds formed by ARS in the target area was higher, and the cavitation resonance might be more intense. The results of taking out the prostate for gross and HE examination 72 hours after treatment also further confirmed that when the sound pressure was 4.1MPa, the mechanical damage to the tissue caused by ultrasonic bubble ablation was the most serious, and a large number of cavity structures of varying sizes appeared in the prostate gland accompanied by bleeding and inflammatory cell infiltration. The damaged area and the damaged area ratio were more than 5 times and 10 times that of the PRO1.5 and PRO2.1 groups, respectively. Although this study shows that the mechanical damage effect of the canine prostate is best at 4.1MPa, it does not mean that all tissues are best at this sound pressure parameter. Because the tissue composition and density of different organs are different (for example, the liver is mainly composed of epithelial tissue, etc., rich in blood vessels, and loose in texture), the tolerance to cavitation resonance is different. Therefore, for different body tissues, the choice of which acoustic parameters should be determined according to the actual situation.

[0022] Long-term test results show that 2D ultrasound data showed that the volume of the prostate was reduced compared to before treatment after ultrasound bubble ablation for one month. Gross observations showed that the damage to the prostate in the short-term experiment was slowly repaired one month after treatment, but the gland still had dark brown stripes or mass scars. HE results showed that the gland structure was severely damaged, showing gland contracture bands, which may be the reason for the reduction in prostate volume.

[0023] The fluorocarbon particles in the present invention become ideal exogenous cavitation nuclei, which can significantly reduce the cavitation threshold and induce enhanced inertial cavitation effect. The application of ARS-mediated ultrasonic tissue destruction prepared in combination with the present invention has the characteristics of short irradiation time, low required ultrasonic sound pressure and sound intensity, minimal tissue damage outside the target area, and convenient operation, thereby improving the targeting and safety of ultrasonic destruction. Although the present invention requires image guidance, it does not require very precise focusing, which reduces the difficulty of operation and has application prospects in multiple clinical treatment fields such as prostate hyperplasia, tumor ablation, and tumor immunotherapy. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 The focused ultrasound device used in the present invention, a is the PRO-KH1 focused ultrasound (FUS) device, b is the appearance of the transducer, and c is the operating interface;

[0025] Figure 2 This is a schematic diagram of ultrasound treatment plan;

[0026] Figure 3 It is the flow performance test of ARS with different concentrations, where a, b, c, d, e, f, g, h, i, j, k, l represent the upright, tilted, and inverted morphological images of ARS with different concentrations, and m, n, o, and p represent the microscopic images of ARS droplets and particles with different concentrations;

[0027] Figure 4 This is a comparison chart of the range of microbubble clouds;

[0028] Figure 5 Two-dimensional ultrasound images of different sound pressure groups before treatment, immediately after treatment, and 72 hours after treatment;

[0029] Figure 6 HE and gross images of prostate tissue 72 hours after short-term experimental treatment;

[0030] Figure 7 These are two-dimensional ultrasound images of the prostate one month after long-term experimental treatment. In the figure, a, b, and c are the microbubble cloud conditions before treatment, immediately after treatment, and one month after treatment. The white dotted line shows the range of the microbubble cloud. d and e are the anterior-posterior diameter, left-right meridian, and upper-lower meridian images of the prostate under two-dimensional ultrasound before treatment. f and g are the anterior-posterior diameter, left-right meridian, and upper-lower meridian images of the prostate under two-dimensional ultrasound one month after treatment.

[0031] Figure 8 HE and gross images of prostate tissues one month after long-term treatment. DETAILED DESCRIPTION

[0032] Example: Acoustically-responsive scaffolds (ARS) based on perfluoropentane phase change particles and sodium alginate and their application in prostate ablation

[0033] Experimental animals

[0034] The research conducted in this invention was approved by the Experimental Animal Welfare and Ethics Committee of the Second Affiliated Hospital of the Army Medical University of the Chinese People's Liberation Army. The male beagles used in the study were purchased from Chongqing Tengxin Huafu Experimental Animal Sales Co., Ltd. All beagles were 38-40 months old and weighed 15±2kg. They were randomly divided into 4 groups: PRO1.5 (PRO-KH1 focused ultrasound + ARS, PNP = 1.5MPa) group, PRO2.1 (PRO-KH1 focused ultrasound + ARS, PNP = 2.1MPa) group, PRO4.1 (PRO-KH1 focused ultrasound + ARS, PNP = 4.1MPa) group, and PRO4.1-LT (PRO-KH1 focused ultrasound + ARS, PNP = 4.1MPa, Long-term) group, 3 in each group. After anesthesia was induced by intramuscular injection of 2.5 ml of xylazine hydrochloride (Shengda Animal Pharmaceutical Co., Ltd. Jilin, China), 7.5 ml of sodium pentobarbital (Sartorius, Germany) was injected intraperitoneally. After successful anesthesia, endotracheal intubation was performed to monitor breathing. The limbs were fixed and the lower abdomen was skinned.

[0035] Preparation of ARS

[0036] Preparation of perfluoropentane phase change microparticle emulsion: Perfluoropentane phase change microparticle emulsion was prepared according to the method in the literature [Li H, He H, Tang J, Luo T, Yang G, Huang L, Dong X, Liu Z: A new sonoablation using acoustic droplet vaporization and focused ultrasound: A feasibility study. Med Phys 2023, 50: 6663-6672.].

[0037] Scaffold preparation: Using physiological saline as a solvent, a sodium alginate solution with a concentration of 20% (20 mg sodium alginate dissolved in 1 ml physiological saline) was prepared.

[0038] After preparing the required reagents, prepare ARS according to the following ratio: 1 ml 20% sodium alginate solution, 100 ul perfluoropentane phase change microparticle emulsion, shake well to mix, place on ice for 5 minutes after mixing and set aside.

[0039] The focused ultrasound device uses a small FUS transducer PRO-KH1 (PRO-KH1, Shenzhen Pro Technology Co., Ltd., China). Figure 1 As shown in a, b, and c, the transducer chip has a diameter of 35 mm, an operating frequency of 797 kHz, and a focal length of 6 cm. The ultrasonic pulse parameters can be adjusted through the treatment software, wherein the peak negative sound pressure is adjustable from -1 to -5 MPa, the pulse width is adjustable from 8 to 100 cycles, and the duty cycle is adjustable from 0.1 to 2%. The sound pressures selected by the instrument of the present invention are 1.55 MPa, 2.10 MPa, and 4.10 MPa, respectively. A cycle of running for 4 seconds and stopping for 1 second is adopted, and the duty cycle is 0.4%.

[0040] Ultrasound therapy options such as Figure 2 Short-term study: Before and immediately after treatment and 72 hours later, all beagle dogs were imaged using a VINNO 70 diagnostic ultrasound system (VINNO Technology Co., Ltd., China) and an X4-12L linear array transducer (VINNO Technology Co., Ltd., China) for two-dimensional ultrasound imaging of the prostate. The same parameters were used for each image acquisition. ARS was injected into the middle of the right and left lobes of the prostate through the lower abdomen under ultrasound guidance. 0.2 ml ( Figure 2 ). An ultrasound coupling pad with a thickness of 3 cm was placed, and two-dimensional ultrasound was used to determine that the ARS in the prostate was located at the focus of the transducer. FUS irradiation was then performed immediately, 3 times, 10 minutes each time, with an interval of 1 minute. The transducer position was adjusted as needed during the treatment interval to ensure that the microbubble cloud produced by the ARS was within the optimal irradiation range. At the end of the treatment, 2 million units of sodium penicillin were injected intramuscularly. The animals were kept for 72 hours and then killed by an overdose of anesthetic. The prostate was immediately removed after sacrifice, and the prostate tissue was cut into multiple layers along the horizontal plane, with each layer about 2 mm thick, for gross examination and photography. It was then fixed with paraformaldehyde (Servicebio Cat) for 24 hours before pathological examination.

[0041] Long-term study: The experimental treatment was the same as that described in the short-term study. Ultrasound two-dimensional imaging of the beagle dog prostate was performed before and immediately after treatment and 1 month later. The animals were killed by overdose of anesthetic after 1 month of feeding and the prostate was immediately removed. The treatment method and examination content of the prostate were the same as those in the short-term study.

[0042] HE staining

[0043] The prostate tissue fixed with paraformaldehyde was dehydrated in graded ethanol solutions, paraffin-embedded sections were rewarmed and fixed, and stained with hematoxylin and eosin. Three sections were randomly selected from each slide and observed under an Olympus upright microscope (10x and 20x).

[0044] Tissue damage assessment

[0045] The damage to prostate tissue caused by the cavitation mechanical effect of ultrasonic ablation is divided into 4 levels based on gross and pathological results:

[0046] Grade 0: No abnormality is found in the prostate;

[0047] Grade 1: The gland cavity is deformed, and the interstitial edema and widening;

[0048] Grade 2: tissue damage, red blood cell infiltration, and inflammatory cell infiltration;

[0049] Grade 3: Tissue liquefaction cavity is formed after 72 hours, and the three-dimensional volume of the prostate ultrasound is reduced after 1 month.

[0050] The pathological images were processed using CaseViewer (CaseViewer 2.4, 3DHISTECH Ltd., Budapest, Hungary) software to delineate the areas of different injury types and calculate the damaged area and damaged area ratio (Table 1).

[0051] Table 1 The damaged area and damaged area ratio of the prostate after treatment (x±SD)

[0052]

[0053] *The difference was statistically significant compared with the PRO1.5-ARS group. # There was a statistically significant difference compared with the PRO2.1-ARS group.

[0054] Statistics

[0055] All data were expressed as mean ± standard deviation (SD). SPSS (SPSS26.0, IBM Corporation, USA) software was used for statistical analysis, and GraphPad Prism (version 8.0; GraphPad Software, LCC, USA) was used for drawing. Paired t-test was used to compare data before and after treatment, one-way analysis of variance was used to compare data between multiple groups, and the least significant difference method was used for further comparison between two groups. MannWhitney U test was used to evaluate the differences between groups for non-normally distributed data. P<0.05 was considered statistically significant.

[0056] result

[0057] All animals had stable vital signs and no signs of awakening during anesthesia, acoustically-responsive scaffold (ARS) injection, and FUS irradiation. No animals died 72 hours and 1 month after treatment. All animals were in good mental state and had no symptoms of bleeding or infection.

[0058] Optimization of ARS

[0059] Previous studies have found that pure perfluoropentane phase change microparticle emulsion has high fluidity and cannot be well confined to the target area and diffuse into the surrounding tissues, which may lead to a decrease in the concentration of phase change microparticles in the target area and damage to the surrounding tissues. In order to overcome this shortcoming, this embodiment designs a new ARS, namely perfluoropentane phase change microparticle emulsion + sodium alginate scaffold. This embodiment sets up 4 concentration gradient sodium alginate scaffolds: 25mg / ml, 20mg / ml, 15mg / ml, 10mg / ml. Subsequently, the fluidity of the container containing ARS is observed by tilting it and the droplet microbubbles are observed under a microscope ( Figure 3 It was found that sodium alginate has the best comprehensive effect at a concentration of 20 mg / ml, which can restrain the strong fluidity of perfluoropentane phase change particles to a certain extent ( Figure 3 g, h, i), and can be well emulsified to form droplet particle emulsions. The droplet particle stability and particle size uniformity are good, and the average particle size is 35±15 microns ( Figure 3 o).

[0060] It was observed that when perfluoropentane phase change microparticle emulsion was simply injected into the liver target area, PFP droplets quickly diffused through blood vessels and tissue gaps, and under the action of FUS, a microbubble cloud was formed that extended far beyond the target area ( Figure 4 a). In the ARS injection group, PFP droplets were well confined to the target area, forming an elliptical microbubble cloud ( Figure 4 b).

[0061] Low-frequency and low-pressure focused ultrasound with different sound pressure parameters stimulates ARS to form microbubble clouds

[0062] Continuous cavitation resonance is a necessary condition for mechanical damage. The cavitation resonance intensity of the microbubble cloud determines the degree of mechanical damage to the tissue. How to make the microbubble cloud cavitate and resonate for a long time, appropriate acoustic parameters are very critical. Acoustic parameters that affect the cavitation effect include sound pressure, pulse width, duty cycle, etc. Among them, sound pressure is the most core indicator.

[0063] In order to investigate the formation of microbubble clouds in the target area by low-frequency, low-sound-pressure focused ultrasound stimulating ARS to produce ADV under different acoustic parameter conditions, the target area ARS was stimulated using acoustic parameters with a center frequency of 797KHz and sound pressures of PNP = 1.5MPa, PNP = 2.1MPa, and PNP = 4.1MPa. It was observed that before the injection of ARS, the prostate gland had a relatively uniform echo, and the stroma and ducts ( Figure 5a, d, g). Because the presence of the sodium alginate scaffold limits the fluidity of the fluorocarbon phase-change droplets, a strong echo cluster is formed in the prostate after ARS injection, which is confined to the needle tip. After injection and excitation by FUS, the brightness of the echo area in the three groups increased, and a strong echo with acoustic shadow was formed. This is because ARS forms a microbubble cloud under the action of FUS, and the echo is enhanced and an acoustic shadow appears. Compared with PRO1.5 and PRO2.1, the echo area of ​​PRO4.1 group is brighter and has a larger range ( Figure 5 b, e, h). 72 hours after the treatment, the microbubble clouds in the PRO4.1 group were still obvious, while the microbubble clouds in the other groups were significantly reduced or even disappeared ( Figure 5 c, f, i). The above results show that ARS undergoes ADV to form microbubble clouds under the action of low-frequency and low-sound-pressure focused ultrasound. The higher the sound pressure, the more intense the cavitation resonance, which provides data support for the selection of acoustic parameters for subsequent long-term experiments.

[0064] Results of mechanical cavitation ablation of prostate tissue

[0065] 72 hours after the end of FUS treatment, the prostate was completely harvested for gross observation and pathological examination. The results of gross examination showed that 72 hours after combined treatment, the prostate target tissues of the three groups showed varying degrees of damage ( Figure 6 g, h, i). The main manifestation of the PRO1.5 group was large-scale tissue bleeding, the PRO2.1 group had a smaller cavity with severe bleeding, and the PRO4.1 group had a significant large-scale cavity-like damage to the prostate tissue with severe bleeding ( Figure 6 i). HE staining showed that the three groups of prostate tissues had white cavity structures of varying sizes, surrounded by inflammatory cell infiltration or glandular cavity deformation and expansion accompanied by red blood cell filling ( Figure 6 d, e, f). The PRO1.5 group was mainly infiltrated by inflammatory cells, with deformed and widened glandular cavities accompanied by red blood cell filling, with small or no cavity structure, and the prostate injury area was 0.98±1.03mm 2 , the damaged area ratio was 0.35±0.41% (Table 1); the PRO2.1 group had a larger cavity structure and glandular cavity deformation accompanied by inflammatory cell infiltration, and the prostate tissue damaged area was 2.97±2.58mm 2 (Table 1), the damage area ratio was 0.60±0.48%. In the prostate tissue of the PRO4.1 group, a large number of cavities of varying sizes appeared, with a damage area of ​​11.68±9.43mm 2(Table 1), the damaged area ratio was 3.89±2.84%. There was no significant difference in the damaged area and damaged area ratio between the PRO1.5 group and the PRO2.1 group. The difference was that the PRO1.5 group had more red blood cell infiltration, and the PRO4.1 group had significantly increased damaged area and damaged area ratio compared with the PRO2.1 and PRO1.5 groups. The above results show that low-frequency and low-sound pressure focused ultrasound excitation of ARS for cavitation resonance can cause damage to prostate tissue, especially when PNP=4.1MPa, it can significantly damage prostate tissue.

[0066] Ultrasound monitoring of prostate volume changes 1 month after ultrasound ablation

[0067] The short-term test results showed that the PRO4.1 group had the best mechanical ablation effect on prostate tissue, so this example selected the acoustic parameters of PRO4.1 to conduct a long-term ablation test on the canine prostate. The results of two-dimensional ultrasound imaging showed that the overall echo of the prostate was uniform before treatment, and some areas had high echo spots, which may be the calcification of the prostate itself. Immediately after treatment, we can clearly see that localized irregular high echo areas appeared in the prostate target area. This is the result of the microbubble cloud formed by ADV under the action of low-frequency and low-sound pressure focused ultrasound of ARS, which produces strong cavitation resonance under continuous irradiation of FUS ( Figure 7 b). Ultrasound two-dimensional imaging was performed again one month after treatment, and it was found that the microbubble cloud had basically disappeared ( Figure 7 c). Subsequently, ultrasound was used to measure the anterior-posterior diameter, left-right diameter, and superior-inferior diameter of the prostate before and one month after treatment to estimate its volume change ( Figure 7 d, e, f, g). Ultrasound measurement results showed that the prostate volume increased from 6.57±0.92cm before treatment to 1 month after ultrasound bubble ablation. 3 Significantly reduced to 4.84±0.15cm 3 Specifically, the anterior-posterior diameter, left-right diameter and superior-inferior diameter of the prostate were reduced from 1.78±0.37 cm, 2.61±0.39 cm and 2.77±0.38 cm before treatment to 1.41±0.35 cm, 2.61±0.39 cm and 2.77±0.38 cm after treatment.

[0068] Prostate macroscopic and pathological findings 1 month after ultrasound ablation

[0069] One month after ultrasound ablation, the prostate was completely harvested for gross observation and pathological examination. The gross results showed that one month after treatment, dark brown stripes or clusters of scars (indicated by the yellow arrow) were visible in the target tissue of the prostate ( Figure 8d, e, f). HE staining results showed that compared with the prostate tissue around the target area, the prostate tissue in the target area was atrophic, the glandular cavity was significantly reduced, the glandular cells were reduced, and the periphery was replaced by a homogeneous light-stained scar structure accompanied by inflammatory cell infiltration ( Figure 8 a, b, c). The atrophy of the local prostate gland may lead to a reduction in prostate volume, which is consistent with the result of ultrasound measurement of prostate volume reduction. In addition, the boundary of the glandular atrophy area is clear, and normal prostate tissue can be seen around it. The above results show that low-frequency, low-sound pressure focused ultrasound excites ARS for cavitation resonance, which can safely and targetedly damage prostate tissue and reduce prostate volume.

[0070] Summarize

[0071] In this experiment, low-frequency, low-sound-pressure, weakly focused ultrasound combined with ARS was used to treat the canine prostate. Ultrasound two-dimensional imaging was performed before and after treatment, and the prostate was taken for gross and pathological examination 72 hours and 1 month after treatment.

[0072] Two-dimensional ultrasound showed that the ARS prepared by the present invention can be well accumulated in the target area. All groups produced microbubble clouds after treatment, but the microbubble clouds produced by the PRO4.1 group were more echogenic and had a larger range than those of the other two groups, and they still existed after 72 hours. Gross observations showed that the prostates of both the PRO4.1 and PRO2.1 groups had many cavities of varying sizes accompanied by severe bleeding, but the cavities in the PRO4.1 group were larger in area and more numerous. No cavities were found in the prostate of the PRO1.5 group, and only a certain degree of bleeding existed. HE detection showed that the total area and damage ratio of the prostate in the PRO4.1 group were significantly higher than those in the other two groups. Long-term damage observations were performed on the canine prostate under the acoustic pressure condition of PNP = 4.1 MPa. Ultrasonic measurements of the anterior-posterior diameter, left-right diameter, and upper-lower diameter of the prostate before and one month after treatment showed that the volume of the prostate was significantly reduced one month after ultrasonic bubble ablation, from 6.57±0.92cm before treatment. 3 Significantly reduced to 4.84±0.15cm 3 Gross observation showed dark brown stripe or mass scars in the target area. HE examination showed atrophy of prostate tissue, shrinkage of glandular cavity and reduction of glandular cells in the target area.

[0073] In summary, when PNP = 4.1 MPa, ultrasound bubble ablation can safely and efficiently mechanically damage prostate tissue and reduce prostate volume. This technology is expected to become a new means of treating BPH in the future. In addition to the perfluoropentane used in Example 1, acoustic response stents can be prepared. On this basis, those skilled in the art can also replace perfluoropentane with common fluorocarbon materials such as Octafluoropropane (C3F8), Sulfur fluoride (SF6), Perfluorobutane (C4F10), Nonafluoro (trifluoromethyl) cyclopentane (C6F12), Perfluorohexane (C6F14), Perfluoroheptane (C7F16), and Perfluorooctane (C8F18). Microbubble clouds can also be formed under low-frequency and low-sound pressure focused ultrasound excitation. The microbubble clouds can resonate by inertial cavitation under continuous ultrasound irradiation, thereby producing cavitation mechanical damage of controllable intensity to local tissues.

Claims

1. A novel acoustic response bracket, characterized in that: It is an acoustic support containing air, fluorocarbon gas, sulfur hexafluoride gas or liquid fluorocarbon compound; the support material is used as the liquid support material, and the interior contains air, fluorocarbon gas microbubbles, sulfur hexafluoride gas microbubbles, liquid fluorocarbon compound or a mixture of the above gases or compounds, forming an acoustic response support containing the above gas or liquid fluorocarbon compound particles.

2. A novel acoustic response bracket according to claim 1, characterized in that: The scaffold material includes sodium alginate, fibrinogen, F127 (polyethylene glycol-polypropylene glycol-polyethylene glycol triblock copolymer), poly N-isopropylacrylamide-polyethylene glycol block copolymer, gelatin, collagen, cellulose, silk protein, keratin, heparin, whey protein, zein, PCLA-PEG-PCLA [poly (ε-caprolactone-co-lactide)-b-poly (ethylene glycol)-bpoly(ε-caprolactone-co-lactide)], polycaprolactone-polylactic acid copolymer, chitin, chitosan, polyvinyl alcohol, polyacrylamide, sodium polyacrylate, hydroxyapatite, methacryloyl hyaluronic acid, methacryloyl chitosan and β-amino ester, etc.

3. A novel acoustic response bracket according to claim 1, characterized in that: The air, fluorocarbon gas, sulfur hexafluoride gas or liquid fluorocarbon compound contained in the acoustic response bracket includes: perfluoropropane / Octafluoropropane (C3F8), sulfur hexafluoride / Sulfur fluoride(SF6), Perfluorobutane / Perfluorobutane(C4F10), Perfluoropentane / Perfluoropentane(C5F12), Nonafluoro(trifluoromethyl)cyclopentane(C6F12), Perfluorohexane / Perfluorohexane(C6F14), Perfluoroheptane / Perfluoroheptane (C7F16), perfluorooctane / Perfluorooctane (C8F18), etc.; the gas or liquid fluorocarbon compound contained in the acoustic response bracket can be one of the above gases or liquid fluorocarbon compounds, or a mixture of multiple gases or liquid fluorocarbon compounds.

4. New use of the ARS according to claims 1-3 in the preparation of a new type of low-intensity ultrasound tissue destruction drug.

5. The use according to claim 4, characterized in that: The ARS enters the human body through local puncture injection and is distributed in the target area.

6. The use according to claim 5, characterized in that: The ARS forms a microbubble cloud or undergoes liquid-gas phase change vaporization to form a microbubble cloud under pulsed, low-frequency, low-sound-pressure ultrasonic excitation. The microbubble particle size in the microbubble cloud is not limited to the range of 1-100 microns. Then, inertial cavitation resonance occurs under low-frequency ultrasonic irradiation, and then cavitation mechanical damage with a controllable degree of damage to local tissue is produced.

7. The use according to claim 6, characterized in that: The ultrasonic emission mode of the pulsed, low-frequency, low-sound-pressure ultrasound can be planar ultrasound or focused ultrasound; the ultrasonic emission frequency is in the range of 30kHz-800kHz; the ultrasonic peak negative pressure is in the range of 0.5MPa-8.0MPa; the ultrasonic single pulse width is in the range of 5-200 cycles, and the working duty cycle is less than 5%.

8. The use according to claim 7, characterized in that: The low-frequency ultrasound has an ultrasound emission frequency in the range of 30kHz-800kHz, and can be single-frequency ultrasound, or dual-frequency or multi-frequency combined ultrasound energy emitted simultaneously or at intervals.

9. The use according to any one of claims 4 to 8, characterized in that: The ARS can produce ablation treatment effects such as mechanical damage to human tissue under low-frequency, low-sound-pressure ultrasonic excitation, including: Mechanical destruction and ablation of various pathological tissues such as prostate hyperplasia tissue and malignant solid tumor tissue.

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