Quality assurance for focused ultrasound-induced blood-brain barrier opening procedure using passive acoustic detection.

BACKGROUND: Focused ultrasound (FUS) combined with microbubbles is a promising technique for noninvasive, reversible, and spatially targeted blood-brain barrier opening, with clinical trials currently ongoing. Despite the fast development of this technology, there is a lack of established quality assurance (QA) strategies to ensure procedure consistency and safety. To address this challenge, this study presents the development and clinical evaluation of a passive acoustic detection-based QA protocol for FUS-induced blood-brain barrier opening (FUS-BBBO) procedure. METHODS: Ten glioma patients were recruited to a clinical trial for evaluating a neuronavigation-guided FUS device. An acoustic sensor was incorporated at the center of the FUS device to passively capture acoustic signals for accomplishing three QA functions: FUS device QA to ensure the device functions consistently, acoustic coupling QA to detect air bubbles trapped in the acoustic coupling gel and water bladder of the transducer, and FUS procedure QA to evaluate the consistency of the treatment procedure. FINDINGS: The FUS device passed the device QA in 9/10 patient studies. 4/9 cases failed acoustic coupling QA on the first try. The acoustic coupling procedure was repeatedly performed until it passed QA in 3/4 cases. One case failed acoustic coupling QA due to time constraints. Realtime passive cavitation monitoring was performed for FUS procedure QA, which captured variations in FUS-induced microbubble cavitation dynamics among patients. INTERPRETATION: This study demonstrated that the proposed passive acoustic detection could be integrated with a clinical FUS system for the QA of the FUS-BBBO procedure. FUNDING: National Institutes of Health R01CA276174, R01MH116981, UG3MH126861, R01EB027223, R01EB030102, and R01NS128461.

Focused Ultrasound-Mediated Delivery of Anti-Programmed Cell Death-Ligand 1 Antibody to the Brain of a Porcine Model.

Immune checkpoint inhibitor (ICI) therapy has revolutionized cancer treatment by leveraging the body's immune system to combat cancer cells. However, its effectiveness in brain cancer is hindered by the blood-brain barrier (BBB), impeding the delivery of ICIs to brain tumor cells. This study aimed to assess the safety and feasibility of using focused ultrasound combined with microbubble-mediated BBB opening (FUS-BBBO) to facilitate trans-BBB delivery of an ICI, anti-programmed cell death-ligand 1 antibody (aPD-L1) to the brain of a large animal model. In a porcine model, FUS sonication of targeted brain regions was performed after intravenous microbubble injection, which was followed by intravenous administration of aPD-L1 labeled with a near-infrared fluorescent dye. The permeability of the BBB was evaluated using contrast-enhanced MRI in vivo, while fluorescence imaging and histological analysis were conducted on ex vivo pig brains. Results showed a significant 4.8-fold increase in MRI contrast-enhancement volume in FUS-targeted regions compared to nontargeted regions. FUS sonication enhanced aPD-L1 delivery by an average of 2.1-fold, according to fluorescence imaging. In vivo MRI and ex vivo staining revealed that the procedure did not cause significant acute tissue damage. These findings demonstrate that FUS-BBBO offers a noninvasive, localized, and safe delivery approach for ICI delivery in a large animal model, showcasing its potential for clinical translation.

First-in-human prospective trial of sonobiopsy in high-grade glioma patients using neuronavigation-guided focused ultrasound.

Sonobiopsy is an emerging technology that combines focused ultrasound (FUS) with microbubbles to enrich circulating brain disease-specific biomarkers for noninvasive molecular diagnosis of brain diseases. Here, we report the first-in-human prospective trial of sonobiopsy in high-grade glioma patients to evaluate its feasibility and safety in enriching plasma circulating tumor biomarkers. A nimble FUS device integrated with a clinical neuronavigation system was used to perform sonobiopsy following an established clinical workflow for neuronavigation. Analysis of blood samples collected before and after FUS sonication showed that sonobiopsy enriched plasma circulating tumor DNA (ctDNA), including a maximum increase of 1.6-fold for the mononucleosome cell-free DNA (cfDNA) fragments (120-280 bp), 1.9-fold for the patient-specific tumor variant ctDNA level, and 5.6-fold for the TERT mutation ctDNA level. Histological analysis of surgically resected tumors confirmed the safety of the procedure. Transcriptome analysis of sonicated and nonsonicated tumor tissues found that FUS sonication modulated cell physical structure-related genes. Only 2 out of 17,982 total detected genes related to the immune pathways were upregulated. These feasibility and safety data support the continued investigation of sonobiopsy for noninvasive molecular diagnosis of brain diseases.

First-in-human prospective trial of sonobiopsy in glioblastoma patients using neuronavigation-guided focused ultrasound.

Sonobiopsy is an emerging technology that combines focused ultrasound (FUS) with microbubbles to enrich circulating brain disease-specific biomarkers for noninvasive molecular diagnosis of brain diseases. Here, we report the first-in-human prospective trial of sonobiopsy in glioblastoma patients to evaluate its feasibility and safety in enriching circulating tumor biomarkers. A nimble FUS device integrated with a clinical neuronavigation system was used to perform sonobiopsy following an established clinical workflow for neuronavigation. Analysis of blood samples collected before and after FUS sonication showed enhanced plasma circulating tumor biomarker levels. Histological analysis of surgically resected tumors confirmed the safety of the procedure. Transcriptome analysis of sonicated and unsonicated tumor tissues found that FUS sonication modulated cell physical structure-related genes but evoked minimal inflammatory response. These feasibility and safety data support the continued investigation of sonobiopsy for noninvasive molecular diagnosis of brain diseases.

Incisionless targeted adeno-associated viral vector delivery to the brain by focused ultrasound-mediated intranasal administration.

BACKGROUND: Adeno-associated viral (AAV) vectors are currently the leading platform for gene therapy with the potential to treat a variety of central nervous system (CNS) diseases. There are numerous methods for delivering AAVs to the CNS, such as direct intracranial injection (DI), intranasal delivery (IN), and intravenous injection with focused ultrasound-induced blood-brain barrier disruption (FUS-BBBD). However, non-invasive and efficient delivery of AAVs to the brain with minimal systemic toxicity remain the major challenge. This study aims to investigate the potential of focused ultrasound-mediated intranasal delivery (FUSIN) in AAV delivery to brain. METHODS: Mice were intranasally administered with AAV5 encoding enhanced green fluorescence protein (AAV5-EGFP) followed by FUS sonication in the presence of systemically injected microbubbles. Mouse brains and other major organs were harvested for immunohistological staining, PCR quantification, and in situ hybridization. The AAV delivery outcomes were compared with those of DI, FUS-BBBD, and IN delivery. FINDINGS: FUSIN achieved safe and efficient delivery of AAV5-EGFP to spatially targeted brain locations, including a superficial brain site (cortex) and a deep brain region (brainstem). FUSIN achieved comparable delivery outcomes as the established DI, and displayed 414.9-fold and 2073.7-fold higher delivery efficiency than FUS-BBBD and IN. FUSIN was associated with minimal biodistribution in peripheral organs, which was comparable to that of DI. INTERPRETATION: Our results suggest that FUSIN is a promising technique for non-invasive, efficient, safe, and spatially targeted AAV delivery to the brain. FUNDING: National Institutes of Health (NIH) grants R01EB027223, R01EB030102, R01MH116981, and UG3MH126861.

Induction Therapy of Retinoic Acid with a Temozolomide-Loaded Gold Nanoparticle-Associated Ultrasound Effect on Glioblastoma Cancer Stem-Like Colonies.

Glioblastoma multiforme (GBM) is the most aggressive glioma. The treatment response is always low, and the condition is typically rapidly fatal. The undifferentiated and self-renewal characteristics of cancer stem cells (CSCs) have been reported, and their potential contribution may cause tumor initiation, recurrence, metastasis, and therapeutic resistance. In particular, glioblastoma stem-like cells exhibit highly invasive properties and drug resistance, serving as a model for the development of novel therapeutic strategies. Induction therapy provides an alternative therapeutic strategy to eliminate the stem cell properties of CSCs and enhance therapeutic sensitivity. The differentiated cells may lose their self-renewal ability, downregulate stem cell-related genes and drug resistance genes, and enhance anticancer drug sensitivity. Therefore, the purpose of this study is to establish a niche for glioblastoma stem-like cell selection as a platform and facilitate the assessment of differentiation therapy on GBM cancer stem-like colonies by retinoic acid (RA) with temozolomide (TMZ)-loaded gold nanoparticles (GNPs) associated with low-intensity ultrasound (LIUS). Herein, a hyaluronic acid-based material system was used to isolate GBM cancer stem-like colonies. Colony formation, size determination, stem cell-related marker expression, and GBM cancer stem-like cell marker expression with the culture period were identified. The effect of TMZ on GBM stem-like colonies on HA-based material systems was also determined, and the results revealed that drug resistance was highly enhanced in GBM colonies compared with that in the control cell population. In addition, GBM colonies also exhibited a significant increase in breast cancer resistance protein expression, which is consistent with the drug resistance effect. Furthermore, several factors, including LIUS, RA, and GNPs, were used to determine the possibility of induction therapy. RA with TMZ-loaded GNP-associated LIUS stimulation exhibited a significant and synergistic effect on the differentiation effect and drug sensitivity enhancement. The GBM cancer stem-like colony system presents an opportunity for the development of new therapeutic strategies, and this study provides an alternative differentiation therapy for malignant tumors.

In vitro study of SDF-1α-loaded injectable and thermally responsive hydrogels for adipose stem cell therapy by SDF-1/CXCR4 axis.

Stem cell-based approaches have become a promising therapeutic strategy for treating ischemic diseases. The aim of this study was to develop injectable hydrogel systems for the local release of stromal cell-derived factor-1α (SDF-1α) to recruit adipose stem cells (ASCs) that express CXCR4 to achieve stem cell therapy and therapeutic angiogenesis. Thermoresponsive and injectable chitosan (CS)/ß-glycerophosphate disodium salt pentahydrate (ßGP) hydrogels with different concentrations of hyaluronic acid (HA) were designed and fabricated to achieve local and sustained release of SDF-1α for ASC recruitment. Herein, the material structures, physical properties, gelation temperature, and gelation time of hydrogels with different compositions were determined. The incorporation of 0.9% HA in CS-based hydrogels not only enhanced the gelation time but also increased the strength of the hydrogels. In addition, the results revealed that the thermoresponsive and injectable CS/ßGP/HA hydrogels showed good biocompatibility. In addition, the in vitro release profiles showed that the hydrogels achieved sustained release of SDF-1α over 7 days and enhanced ASC migration. The results revealed that the hydrogels with HA enhanced the sustained release effect compared with the hydrogel without HA, indicating that the HA content regulated the physical and release properties of the injectable hydrogels. Therefore, thermoresponsive and injectable CS/ßGP/HA hydrogels may provide an alternative for treating ischemic diseases via SDF-1/CXCR4 axis for ASC recruitment and retention.

Strategy of differentiation therapy: effect of dual-frequency ultrasound on the induction of liver cancer stem-like cells on a HA-based multilayer film system.

Cancer stem cells (CSCs) and normal stem cells share the ability to self-renew and drive tumor formation, recurrence, and distant metastasis and are resistant to chemotherapeutic drugs. One potential therapeutic approach for targeting CSCs is to induce CSCs to differentiate into normal cancer cells to eliminate self-renewal and enhance drug sensitivity. We developed a hyaluronic acid (HA)-based multilayer film system for selecting CSC-like hepatocellular carcinoma (HCC) cell colonies. Herein, we assess the differentiation therapy of HCC CSCs using dual-frequency low-intensity ultrasound (LIUS). HA-based multilayer films of poly (allylamine hydrochloride), (PAH/HA)6, were used to isolate CSC colonies. Colony formation, maintenance, and CSC marker expression were identified. The colony-formation rate was investigated, and putative CSC markers for CD44/CD133 expression after 7 days of culture were upregulated on (PAH/HA)6 multilayer films. Dual-frequency LIUS was used to induce CSC colony differentiation, and the phenotype variation, CSC marker expression, gene expression, drug-resistance ability, and invasion ability of CSC colonies with/without LIUS stimulation were compared. The numbers of colonies and CD44/CD133 double-positive cells and the expression levels of stem cell-related genes and proteins associated with stemness all decreased due to differentiation after LIUS exposure. Furthermore, a significant reduction in CSC drug resistance and invasion ability was observed. These results indicate that dual-frequency LIUS induces CSC differentiation and reduces drug resistance and invasion ability. Differentiation of CSCs provides an alternative therapeutic strategy to reverse CSC stemness and force their loss of self-renewal ability. CSC-targeted therapy holds great promise as an effective therapeutic approach for the treatment of human tumors.