Rationale: Glioblastoma multiforme (GBM) is the most aggressive primary malignant brain tumor in adults, characterized by high invasiveness and poor prognosis. Glioma stem cells (GSCs) drive GBM treatment resistance and recurrence, however, the molecular mechanisms activating intracranial GSCs remain unclear. Extracellular vesicles (EVs) are crucial signaling mediators in regulating cell metabolism and can cross the blood-brain barrier (BBB). This study aimed to elucidate how EV cargo contributes to the intracranial GSC state and validate a non-invasive diagnostic strategy for GBM relapse. Methods: We isolated plasma extracellular vesicles (pl-EVs) from three groups: recurrent GBM patients post-resection, non-recurrent GBM patients post-resection, and healthy individuals. Newly diagnosed GBM patients served as an additional control. EVs were characterized and co-cultured with primary GBM cell lines to assess their effect on tumor stemness. EV cargo was analyzed using proteomics to investigate specific EV subpopulations contributing to GBM relapse. Based on these findings, we generated engineered LDHA-enriched EVs (LDHA-EVs) and co-cultured them with patient-derived organoids (PDOs). Metabolomics was performed to elucidate the underlying signal transduction pathways. Results: Our study demonstrated that pl-EVs from recurrent GBM patients enhanced aerobic glycolysis and stemness in GBM cells. Proteomic analysis revealed that plasma EVs from recurrent GBMs encapsulated considerable amounts of the enzyme lactate dehydrogenase A (LDHA). Mechanistically, LDHA-loaded EVs promoted glycolysis, induced cAMP/ATP cycling, and accelerated lactate production, thereby maintained the GSC phenotype. Concurrently, post-surgical therapy-induced stress-modulated hypoxia in residual tumors, promoted LDHA-enriched EV release. Clinically, high levels of circulating LDHA-positive EVs correlated with increased glycolysis, poor therapeutic response, and shorter survival in recurrent GBM patients. Conclusion: Our study highlights LDHA-loaded EVs as key mediators promoting GSC properties and metabolic reprogramming in GBM. These findings provide insights into recurrence mechanisms and suggest potential liquid biopsy approaches for monitoring and preventing GBM relapse.

The success of pancreaticojejunostomy (PJ) critically depends on achieving optimal pancreatic juice drainage. Clinical evidence demonstrates that internal stent placement is highly safe and effective in reducing postoperative complications associated with PJ. However, recent clinical observations have raised concerns regarding potential complications following pancreatic duct stent placement, including intestinal wall perforation caused by the stent. This study aims to describe the design and step-by-step use of a bulb-tipped internal pancreatic duct stent during PJ and to report short-term postoperative outcomes in a consecutive single center series (January 2021-January 2023). The clinical data of 33 patients who underwent PJ with the improved tube from January 2021 to January 2023 were reviewed. General information (gender, age, underlying disease), operation-related information (intraoperative blood loss, postoperative complications, postoperative hospital stay, prognosis) and postoperative diagnosis were retrospectively analyzed. All patients, with a median age of 60 (range, 46-75) years, 18 males and 15 females, underwent PJ successfully, and the improved duct stent was successfully placed during the operation. The median intraoperative blood loss was 100 (range, 50-700) mL. Postoperative complications were observed in 16 patients (48.5%). Among them, a total of 5 patients (15.2%) experienced Grade B/C postoperative pancreatic fistula (POPF). The specific types of complications were as follows: Grade A POPF (n=11, accounting for 33.3%), Grade B POPF [n=4, accounting for 12.1%; among which there were 3 cases of delayed gastric emptying (DGE) and 1 case of surgical site infection], and Grade C POPF (n=1, accounting for 3.0%). Notably, no stent-related complications occurred (0%; 95% confidence interval: 0.0-10.6%). The median postoperative hospitalization was 15 (range, 12-38) days. Among the 31 patients (93.9%) who completed the 3-month follow-up, two cases (6.1%) were lost to follow-up. Preliminary findings suggest that the optimized pancreatic duct stent exhibits favorable early-stage safety in current clinical applications. No stent-related complications were detected within this cohort. As further research progresses, this improved medical device holds promise for demonstrating broader clinical utility and substantial potential in enhancing patient treatment efficacy.

Phenformin has antitumor effects against pancreatic cancer. We report a phenformin complex with the technetium-99m tricarbonyl [99mTc(CO)3], termed 99mTc-Phen, that permitted noninvasive visualization of tumor uptake in models of pancreatic cancer by single-photon emission computed tomography. 99mTc-Phen's nonradioactive, structurally matched counterpart [Re(CO)3]-phenformin, termed Re-Phen, showed antitumor activity against an orthotopic murine pancreatic ductal adenocarcinoma tumor with KrasG12D mutation in immunocompetent mice but not against orthotopically inoculated human MiaPaca-2 PDAC xenografts in nude mice. In vitro, Re-Phen was less potent than phenformin in inhibiting cell proliferation and colony formation and inducing cell death. However, Re-Phen but not phenformin potently reduced mitochondrial membrane potential. Unlike phenformin, Re-Phen caused no significant changes in mitochondrial ATP production or ATP generation by glycolysis at 10-100 μM, suggesting that Re-Phen and phenformin may act through distinct mechanisms. These findings underscore the potential utility of structurally matched Re/99mTc(CO)3-biguanide complexes as a new class of theranostic pairs.

Aim: The aim of this study was to assess the utility of weighted amide proton transfer (APTw) MRI in three different rodent models of hepatocellular carcinoma (HCC).Methods: APTw MRI was evaluated in models of diethylnitrosamine (DEN) induced HCC, N1S1 syngeneic orthotopic xenograft and human HepG2 ectopic xenograft.Results: All models of HCC showed a higher APTw signal over the surrounding normal tissues. In the DEN model, the APTw signal could differentiate HCC lesions from benign nodules. Intra-arterial administration of low-density lipoprotein docosahexaenoic acid (LDL-DHA) nanoparticles to N1S1 xenografts rapidly lowered the tumor APTw signal within 72 h. Direct injections of LDL-DHA nanoparticles into HepG2 xenografts also showed similar therapeutic responses.Conclusion: We have demonstrated the utility of APTw imaging in the diagnostic/therapeutic management of HCC.

Background: Microvascular visualization is crucial in understanding the mechanisms of several pathologies. For instance, visualization of the tumor microenvironment is important in understanding angiogenesis and role in cancer progression. Visualization would provide insights to cancer diagnosis, predicting metastatic growth, and evaluating therapeutic protocols. Similarly, understanding the microvascular network could be beneficial for study of degenerative diseases and tissue repair. The use of microscale computed tomography (micro-CT) and vascular casting agents provides high-resolution images of tissue vasculature in volumetric space. The purpose of this research was to compare a selection of commercially available contrast agents to determine the optimal solution for vascular visualization. Methods: A population of 16 female nude athymic mice (Charles Rivers Laboratories) were implanted with MDA-MB-231 breast cancer cells (ATCC) orthotopically in the lower left mammary fat pad to investigate the tumor microenvironment. Once tumors reach sufficient size, animals were equally divided into four groups based on the micro-CT agent to be administered, namely, control (no contrast agent), barium sulfate (BaSO4), Vascupaint, or Microfil. Animals were anesthetized prior to transcarotid micro-cannulation to infuse 2 mL of the specific contrast agent for intravascular distribution throughout the animal. The jugular vein on the other side of the carotid artery was opened to drain blood flow. Following successful perfusion, animals and extracted organs underwent high-resolution micro-CT scanning (OI/CT, MILabs). Images were reconstructed and analyzed using analysis software to extract mean intensity signals. Results: Preliminary post-mortem micro-CT results reveal Vascupaint and BaSO4 are useful for microvascular visualization. Both Vascupaint and BaSO4 produced significant contrast-enhanced micro-CT image enhancement in the brain (3.39 +/- 0.93 and 6.27 +/- 3.78, respectively) and kidney (12.85 +/- 1.98 and 32.87 +/- 10.03, respectively) as compared to Microfil (0.22 +/- 0.07 and 0.91 +/- 0.63, respectively; P<0.05). For the various contrast agents, there were no differences in image enhancement from the liver, spleen, or tumor tissue (P>0.21). Moreover, use of Vascupaint and BaSO4 allowed for visualization of smaller microvascular structures with average diameters of 20.54 +/- 4.15 and 25.82 +/- 3.75 mu m, which were smaller compared to the 91.66 +/- 24.91 mu m measurements from Microfil-enhanced micro-CT images (P<0.004). Conclusions: Our study suggests that the use of Vascupaint and BaSO4 is more than sufficient for ex vivo visualization of microvascular structures with contrast-enhanced micro-CT imaging as these contrast agents more effectively perfused smaller blood vessels.

BACKGROUND:The prevalence of liver diseases, especially steatosis, requires a more convenient and noninvasive tool for liver diagnosis, which can be a surrogate for the gold standard biopsy. Magnetic resonance (MR) measurement offers potential, however ultrasound (US) has better accessibility than MR. PURPOSE:This study aims to suggest a multiparametric US approach which demonstrates better quantification and imaging performance than MR imaging-based proton density fat fraction (MRI-PDFF) for hepatic steatosis assessment. METHODS:We investigated early-stage steatosis to evaluate our approach. An in vivo (within the living) animal study was performed. Fat inclusions were accumulated in the animal livers by feeding a methionine and choline deficient (MCD) diet for 2 weeks. The animals (n = 19) underwent US and MR imaging, and then their livers were excised for histological staining. From the US, MR, and histology images, fat accumulation levels were measured and compared: multiple US parameters; MRI-PDFF; histology fat percentages. Seven individual US parameters were extracted using B-mode measurement, Burr distribution estimation, attenuation estimation, H-scan analysis, and shear wave elastography. Feature selection was performed, and the selected US features were combined, providing quantification of fat accumulation. The combined parameter was used for visualizing the localized probability of fat accumulation level in the liver; This procedure is known as disease-specific imaging (DSI). RESULTS:The combined US parameter can sensitively assess fat accumulation levels, which is highly correlated with histology fat percentage (R = 0.93, p-value < 0.05) and outperforms the correlation between MRI-PDFF and histology (R = 0.89, p-value < 0.05). Although the seven individual US parameters showed lower correlation with histology compared to MRI-PDFF, the multiparametric analysis enabled US to outperform MR. Furthermore, this approach allowed DSI to detect and display gradual increases in fat accumulation. From the imaging output, we measured the color-highlighted area representing fatty tissues, and the fat fraction obtained from DSI and histology showed strong agreement (R = 0.93, p-value < 0.05). CONCLUSIONS:We demonstrated that fat quantification utilizing a combination of multiple US parameters achieved higher performance than MRI-PDFF; therefore, our multiparametric analysis successfully combined selected features for hepatic steatosis characterization. We anticipate clinical use of our proposed multiparametric US analysis, which could be beneficial in assessing steatosis in humans.

Tumors become inoperable due to their size or location, making neoadjuvant chemotherapy the primary treatment. However, target tissue accumulation of anticancer agents is limited by the physical barriers of the tumor microenvironment. Low-intensity focused ultrasound (FUS) in combination with microbubble (MB) contrast agents can increase microvascular permeability and improve drug delivery to the target tissue after systemic administration. The goal of this research was to investigate image-guided FUS-mediated molecular delivery in volume space. Three-dimensional (3-D) FUS therapy functionality was implemented on a programmable ultrasound scanner (Vantage 256, Verasonics Inc.) equipped with a linear array for image guidance and a 128-element therapy transducer (HIFUPlex-06, Sonic Concepts). FUS treatment was performed on breast cancer-bearing female mice (N= 25). Animals were randomly divided into three groups, namely, 3-D FUS therapy, two-dimensional (2-D) FUS therapy, or sham (control) therapy. Immediately prior to the application of FUS therapy, animals received a slow bolus injection of MBs (Definity, Lantheus Medical Imaging Inc.) and near-infrared dye (IR-780, surrogate drug) for optical reporting and quantification of molecular delivery. Dye accumulation was monitored viain vivooptical imaging at 0, 1, 24, and 48 h (Pearl Trilogy, LI-COR). Following the 48 h time point, animals were humanely euthanized and tumors excised forex vivoanalyzes. Optical imaging results revealed that 3-D FUS therapy improved delivery of the IR-780 dye by 66.4% and 168.1% at 48 h compared to 2-D FUS (p= 0.18) and sham (p= 0.047) therapeutic strategies, respectively.Ex vivoanalysis revealed similar trends. Overall, 3-D FUS therapy can improve accumulation of a surrogate drug throughout the entire target tumor burden after systemic administration.

The treatment of glioblastoma has limited clinical progress over the past decade, partly due to the lack of effective drug delivery strategies across the blood-brain-tumor barrier. Moreover, discrepancies between preclinical and clinical outcomes demand a reliable translational platform that can precisely recapitulate the characteristics of human glioblastoma. Here we analyze the intratumoral blood-brain-tumor barrier heterogeneity in human glioblastoma and characterize two genetically engineered models in female mice that recapitulate two important glioma phenotypes, including the diffusely infiltrative tumor margin and angiogenic core. We show that pulsed laser excitation of vascular-targeted gold nanoparticles non-invasively and reversibly modulates the blood-brain-tumor barrier permeability (optoBBTB) and enhances the delivery of paclitaxel in these two models. The treatment reduces the tumor volume by 6 and 2.4-fold and prolongs the survival by 50% and 33%, respectively. Since paclitaxel does not penetrate the blood-brain-tumor barrier and is abandoned for glioblastoma treatment following its failure in early-phase clinical trials, our results raise the possibility of reevaluating a number of potent anticancer drugs by combining them with strategies to increase blood-brain-tumor barrier permeability. Our study reveals that optoBBTB significantly improves therapeutic delivery and has the potential to facilitate future drug evaluation for cancers in the central nervous system.

Bone microstructure refers to the arrangement and quality of bone tissue at the microscopic level. Understanding the bone microstructure of the skeleton is crucial for gaining insight into the pathophysiology of osteoporosis and improving its treatment. However, handling bone samples can be complex due to their hard and dense properties. Secondly, specialized software makes image processing and analysis difficult. In this protocol, we present a cost-effective and easy-to-use solution for trabecular bone microstructure analysis. Detailed steps and precautions are provided. Micro-CT is a non-destructive three-dimensional (3D) imaging technique that provides high-resolution images of trabecular bone structure. It allows for the objective and quantitative evaluation of bone quality, which is why it is widely regarded as the gold standard method for bone quality assessment. However, histomorphometry remains indispensable as it offers crucial cellular-level parameters, bridging the gap between two-dimensional (2D) and 3D assessments of bone specimens. As for the histologic techniques, we chose to decalcify the bone tissue and then perform traditional paraffin embedding. In summary, combining these two methods can provide more comprehensive and accurate information on bone microstructure.

Introduction: Hepatocellular carcinoma (HCC) remains a leading cause of cancer-related deaths worldwide. Transarterial chemoembolization (TACE) treatment of HCC restricts the blood supply to the tumor. Conventional assessment of tumor response is performed by dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) or computed tomography (DCE-CT) at 4 to 6 weeks after the TACE procedure. An earlier indication of treatment effectiveness would improve patient management and prognosis. Super-resolution ultrasound (SR-US) imaging allows a ten-fold improvement in spatial resolution compared to traditional ultrasound (US) and allows visualization of microvascular networks. The goal of this preclinical research study was to use SR-US imaging to assess the effectiveness of TACE treatment in a relatively brief time frame. Methods: Eight male Sprague Dawley rats weighing around 300 g were injected with 5 million N1S1 cells in the upper left liver lobe and the tumors were allowed to grow for 14 days. The TACE procedure consisted of transhepatic arterial delivery of a mixture of 25 µL Lipiodol and 25 µL Doxorubicin (10 mg/kg) via a polyethylene microcatheter. After intravenous injection of a microbubble (MB) contrast agent, contrast-enhanced US images were acquired (N = 3000) with a preclinical system (Vevo 3100, FUJIFILM VisualSonics Inc) equipped with an MX201 linear array transducer. US images were processed to remove high motion frames, followed by tissue suppression filtering, and then MB localization and accumulation. Morphological filtering was used to enhance the vessel structures. In vivo US, MRI, and CT imaging were performed at baseline before TACE and again at 1 and 2 wk. After euthanasia, tumor tissue was removed for ex vivo analysis. Results: A rat model of HCC for assessing TACE treatment was introduced. Based on tumor size changes and residual perfusion after the TACE procedure, treatments were determined to be a complete responder, partial responder, or non-responder. Initial results demonstrate that SR-US imaging can sensitively detect any reduction in HCC perfusion as measured by a microvascular density (MVD) metric within 2 wk after a single TACE procedure. MVD measurements from the SR-US images were consistence with tumor volume data from MRI and CT imaging. Conclusions: In vivo SR-US images of tumor microvascular networks provided insight into treatment efficacy. Citation Format: Junjie Li, Katherine Brown, Megan Yociss, John Eisenbrey, Kenneth Hoyt. Assessing the effectiveness of transarterial chemoembolization using super-resolution ultrasound imaging and a rat model of hepatocellular carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2460.

Introduction: The first option for cancer treatment is often chemotherapy. However, studies have shown that only 1% of the injected dose reaches the target cancer as the tumor microenvironment presents a physical barrier for optimal drug delivery. Focused ultrasound (FUS) in combination with microbubble (MB) contrast agents is an emerging therapy to improve drug delivery by temporarily increasing microvascular permeability. This research details the development and testing of a novel ultrasound (US) image-guided FUS system and method for enhancing drug delivery to tumor tissue and volume space with comparison to a 2-dimensional (2-D) US therapeutic technology. Methods: Real-time US therapy was implemented on a programmable US system (Vantage 256, Verasonics Inc) equipped with a dual US transducer configuration for interleaved anatomical imaging and volumetric treatment delivery (HIFU-Plex, Sonic Concepts Inc). Both US imaging and therapeutic transducers are co-registered 128 element arrays with center frequencies of 3.5 and 2.0 MHz, respectively. The latter is a concentric array that enables beam steering in 3-dimensional (3-D) space. US treatment was performed at a peak negative pressure of 0.7 MPa (mechanical index, MI of 0.45), pulse repetition frequency of 10 Hz, and duty cycle of 10%. BALB/c mice (N = 22, Charles River Laboratory) were implanted with 2.0 × 105 breast cancer cells (4T1, ATCC). Once tumors reached 0.6 cm in size, mice were randomly divided into a 3-D or 2-D US therapy group or sham control. US therapy was performed following an intravascular injection of microbubbles (Definity, Lantheus Medical Imaging) and IR-780 dye. Note microbubbles function as a therapeutic mediator whereas the fluorescent dye represents a surrogate small molecule drug. Live animal fluorescent imaging was performed at baseline before US therapy (0 h) and again at 1, 24, and 48 h. Following the 48 h timepoint, animals were euthanized and tumors surgically excised for ex vivo analysis. Results: 3-D US-mediated therapy improved molecular delivery to tumor tissue by 150 and 180% at 24 and 48 h, respectively, when compared to our previously established 2-D US therapeutic approach (p = 0.22) or sham therapy (p = 0.07). A similar trend was observed during ex vivo imaging of excised tumor samples treated with 3-D US therapy as compared to the 2-D US therapeutic approach (p = 0.41) or sham therapy (p = 0.04). Dye extraction further confirmed these observations (p > 0.12). Conclusions: 3-D US therapy improved molecular delivery to the tumor volume compared to our previously established 2-D US-based method as confirmed by a series of optical imaging studies. Citation Format: Ryan Margolis, Junjie Li, Lokesh Basavarajappa, Kenneth Hoyt. Image-guided focused ultrasound-mediated drug delivery for improved cancer treatment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2469.

Head and neck squamous cell carcinoma (HNSCC) is a common form of cancer with more than 54,000 cases expected in the United States in 2022. It is known that advanced disease correlates with an overexpression of epidermal growth factor receptor (EGFR) and leads to a poor prognosis and less favorable outcomes. Cetuximab is a monoclonal antibody that targets the extracellular domain of EGFR. However, the therapeutic effect of cetuximab is in part limited by the physical barriers of the tumor microenvironment. One promising technique that overcomes these challenges is focused ultrasound (FUS) therapy, which safely and temporarily increases microvascular permeability and drug extravasation. This study investigated the use of a novel three-dimensional (3-D) FUS approach to improve cetuximab delivery and treatment of HNSCC. HNSCC-bearing mice were divided into three groups: control and drug $\pm$ FUS. Prior to therapy, each mouse received a bolus injection of saline and microbubbles (50 $\mu \mathbf{L}$ , Definity, Lantheus Medical Imaging) via a tail vein inj ection. FUS therapy was performed using a customizable ultrasound image-guided scanner (HIFUPlex-06, Verasonics Inc). For the drug dosed mice, IRDye750 (Li-COR Biosciences) was conjugated to cetuximab and administered as a single dose of 3.30 mg. $\text{kg}^{-1}$ . In vivo optical imaging (Pearl Trilogy, LI-COR Biosciences) was used to measure drug accumulation at $0 \ \mathbf{d}$ (baseline) and again at 1, 3, and 7 $\mathbf{d}$ . Optical imaging found mice treated with drug + FUS exhibited signal intensity increases of 143.9 and 83.0 % over mice treated with drug alone at 1 and 3 $\mathbf{d}$ , respectively $(p < 0.01)$ . This combined treatment of drug + FUS improved survival by 40 % compared to mice dosed with cetuximab alone $(p=0.04)$ .

Objective: Hepatocellular carcinoma (HCC) is a highly prevalent form of liver cancer diagnosed annually in 600,000 people worldwide. A common treatment is transarterial chemoembolization (TACE), which interrupts the blood supply of oxygen and nutrients to the tumor mass. The need for repeat TACE treatments may be assessed in the weeks after therapy with contrast-enhanced ultrasound (CEUS) imaging. Although the spatial resolution of traditional CEUS has been restricted by the diffraction limit of ultrasound (US), this physical barrier has been overcome by a recent innovation known as super-resolution US (SRUS) imaging. In short, SRUS enhances the visi-ble details of smaller microvascular structures on the 10 to 100 mu m scale, which unlocks a host of new clinical opportunities for US. Methods: In this study, a rat model of orthotopic HCC is introduced and TACE treatment response (to a doxorubi-cin-lipiodol emulsion) is assessed using longitudinal SRUS and magnetic resonance imaging (MRI) performed at 0, 7 and 14 d. Animals were euthanized at 14 d for histological analysis of excised tumor tissue and determination of TACE response, that is, control, partial response or complete response. CEUS imaging was performed using a pre-clinical US system (Vevo 3100, FUJIFILM VisualSonics Inc.) equipped with an MX201 linear array transducer. After administration of a microbubble contrast agent (Definity, Lantheus Medical Imaging), a series of CEUS images were collected at each tissue cross-section as the transducer was mechanically stepped at 100 mu m incre-ments. SRUS images were formed at each spatial position, and a microvascular density metric was calculated. Microscale computed tomography (microCT, OI/CT, MILabs) was used to confirm TACE procedure success, and tumor size was monitored using a small animal MRI system (BioSpec 3T, Bruker Corp.). Results: Although there were no differences at baseline (p > 0.15), both microvascular density levels and tumor size measures from the complete responder cases at 14 d were considerably lower and smaller, respectively, than those in the partial responder or control group animals. Histological analysis revealed tumor-to-necrosis levels of 8.4%, 51.1% and 100%, for the control, partial responder and complete responder groups, respectively (p < 0.005). Conclusion: SRUS imaging is a promising modality for assessing early changes in microvascular networks in response to tissue perfusion-altering interventions such as TACE treatment of HCC.

Introduction The acute exacerbation of chronic obstructive pulmonary disease (AECOPD) has a seriously negative impact on patients’ healths condition and disease progression. Bacterial infection is closely related to AECOPD, and antibiotics are frequently used in clinical practice. The lack of specific biomarkers for rational antibiotics use always leads to antibiotics abuse in chronic obstructive pulmonary disease (COPD) flare-ups. Eosinopenia has been considered to be related to increased bacterial load of potentially pathogenic organisms at the onset of COPD exacerbations. Therefore, this study aims to investigate whether eosinopenia could be used as a reference for the use of antibiotics in AECOPD. Methods and analysis In this study, a hospital-based retrospective cohort design will be adopted to analyse the clinical data of inpatients who are primarily diagnosed with AECOPD in West China Hospital of Sichuan University from 1 January 2010 to 31 December 2020. Relevant data will be extracted from the Clinical Big Data Platform for Scientific Research in West China Hospital, including demographic characteristics, blood eosinophil count, procalcitonin, C reactive protein, microbial cultivation, antibiotics use, length of hospital stay, non-invasive ventilation use, intensive care unit transfer and mortality, etc. The collected data will be described and inferred by corresponding statistical methods according to the data type and their distributions. Multiple binary logistic regression models will be used to analyse the relationship between blood eosinophil count and bacterial infection. The antibiotics use, and patient morbidity and mortality will be compared between patients with or without eosinopenia. Ethics and dissemination This study has been approved by the Biomedical Ethics Review Board of West China Hospital of Sichuan University (Approval No. 2020-1056). And the research results will be published in a peer-reviewed journal. Trial registration number ChiCTR2000039379.

Glioblastoma multiforme (GBM) is the most prevalent malignant tumor in the central nervous system. It has diverse phenotypes, including diffuse single-cell infiltration in which the tumor cells co-opt the normal microvasculature, and the neovascularization of an expanding tumor mass. The blood-brain-tumor barrier (BBTB) is a significant obstacle to GBM treatment and restricts entry of most FDA-approved effective oncology drugs. Herein, we report that picosecond laser excitation of vascular-targeted plasmonic gold nanoparticles (AuNPs) can non-invasively and reversibly modulate the BBTB permeability (optoBBTB). OptoBBTB enhances the delivery of paclitaxel (Taxol) in two genetically engineered glioma models (GEMM) that span the spectrum of GBM phenotypes. OptoBBTB followed by Taxol delivery effectively suppresses tumor growth and prolongs the survival time of both GEMM. Moreover, our results raise the possibility that paclitaxel, which is amongst the most widely used oncology drugs because of its proven efficacy but has been abandoned for GBM following its failure to efficacy in early phase clinical trials due to poor blood-brain barrier (BBB) penetration, could now be reconsidered in combination with strategies to increase BBB permeability. In summary, optoBBTB is a novel and effective approach to increase the delivery of therapeutics with limited BBB permeability to treat neoplastic and non-neoplastic brain diseases.

H‐scan ultrasound (US) imaging is a novel tissue characterization technique to detect apoptosis‐induced changes in cancer cells after the initiation of effective drug treatment. The objective of the proposed research was to assess the sensitivity of 3‐dimensional (3D) H‐scan US technique for monitoring the response of breast cancer‐bearing animals to neoadjuvant chemotherapy and correlate results to diffusion‐weighted magnetic resonance imaging (DW‐MRI) measurements of programmed cancer cell death.

Objectives Three-dimensional (3D) H-scan is a new ultrasound (US) technique that images the relative size of acoustic scatterers. The goal of this research was to evaluate use of 3D H-scan US imaging for monitoring early breast cancer response to neoadjuvant therapy using a preclinical murine model of breast cancer. Materials and Methods Preclinical studies were conducted using luciferase-positive breast cancer–bearing mice (n = 40). Anesthetized animals underwent US imaging at baseline before administration with an apoptosis-inducing drug or a saline control. Image data were acquired using a US scanner equipped with a volumetric transducer following either a shorter- or longer-term protocol. The later included bioluminescent imaging to quantify tumor cell viability. At termination, tumors were excised for ex vivo analysis. Results In vivo results showed that 3D H-scan US imaging is considerably more sensitive to tumor changes after apoptosis-inducing drug therapy as compared with traditional B-scan US. Although there was no difference at baseline ( P > 0.99), H-scan US results from treated tumors exhibited progressive decreases in image intensity (up to 62.2% by day 3) that had a significant linear correlation with cancer cell nuclear size ( R 2 > 0.51, P < 0.001). Results were validated by histological data and a secondary longitudinal study with survival as the primary end point. Discussion Experimental results demonstrate that noninvasive 3D H-scan US imaging can detect an early breast tumor response to apoptosis-inducing drug therapy. Local in vivo H-scan US image intensity correlated with cancer cell nuclear size, which is one of the first observable changes of a cancer cell undergoing apoptosis and confirmed using histological techniques. Early imaging results seem to provide prognostic insight on longer-term tumor response. Overall, 3D H-scan US imaging is a promising technique that visualizes the entire tumor and detects breast cancer response at an early stage of therapy.

The use of ultrasound (US) for tumor tissue characterization remains an exciting prospect. The purpose of this research project was to introduce a 3-dimensional (3-D) H-scan US imaging system to monitor response of breast cancer to neoadjuvant chemotherapy. Studies were conducted using female mice ( $N= 20$ ) implanted with 1 million breast cancer cells (MDA-MB-231). Once tumors were formed, animals were treated with a sham drug or low or high doses of cisplatin. In vivo US imaging was performed using Vevo 3100 system (FUJIFILM VisualSonics Inc) equipped with an MX201 transducer and motorized positioner. To generate the H-scan US images, Gaussian-weighted Hermite polynomial filters were convolved with the radiofrequency (RF) data to measure the relative strength of the backscattered US signals. Animals also underwent diffusion-weighted magnetic resonance imaging (DW-MRI)) using a preclinical scanner (BioSpec 3T, Bruker Corp) for generation of apparent diffusion coefficient (ADC) maps as a measure of intratumoral water diffusion. In vivo results demonstrated that 3-D H-scan US imaging was more sensitive to tumor changes after neoadjuvant chemotherapy as compared to B-scan US. While there was no difference at baseline (p > 0.60), H-scan US images from the cisplatin treated tumors exhibited increased intensity at 7 d (36.4 ± 6.9% and 43.8 ± 8.1% for low and high dosed groups, respectively) indicating a decrease in aggregate US scatterer size. Collectively, these observations were confirmed by histological analysis. Overall, 3-D H-scan US imaging is a promising new tool for monitoring cancer response to neoadjuvant therapy. In vivo results matched those found using DW-MRI, which is an established modality for assessing anticancer treatment using apoptosis-inducing drugs.

Hepatocellular carcinoma (HCC) represents a significant disease burden and mortality risk worldwide. Transarterial chemoembolization (TACE) is a minimally invasive treatment procedure that restricts tumor blood flow after local injection of an embolization agent and anticancer drug mixture. Assessment of tumor response to TACE is currently determined by using magnetic resonance imaging (MRI) or computed tomography (CT) at 4 to 6 $\mathbf{wk}$ . Recently, it was shown that contrast-enhanced ultrasound (CEUS) imaging at 1 to 2 wk may result in a similarly if not better effective assessment while giving an earlier indication of tumor response and improving treatment management. However, measurement of tumor perfusion levels is limited by conventional processing of CEUS images and could be improved by adapting methods used for super-resolution ultrasound (SRUS) imaging. The overall goal of this research was to develop an improved CEUS-based assessment of liver cancer response to TACE therapy. Using a rat model of HCC, tumors were treated with TACE or a shame procedure ( $(N=3$ per group). Immediately following tail vein injection of a microbubble (MB) contrast agent, CEUS imaging was performed using a preclinical ultrasound system (Vevo 3100, FUJIFILM VisualSonics Inc) equipped with a 15 MHz linear transducer. Imaging was performed in anesthetized animals at baseline before TACE and again 7 and 14 d thereafter. Improved CEUS images were generated by applying spatiotemporal filtering followed by MB detection and localization. Microvessel density (MVD) was calculated from CEUS images as a percentage of tumor area with microvascular structures. On average, MVD decreased by 56.0 % in the TACE treated animals after 1 wk, versus 11.9 % in the control animals. Tumor diameter decreased by 20.3 % in the treated group animals after 1 wk, while tumors grew 12.1 % in the control group. The level of microvascular detail in the CEUS images provided insight into treatment efficacy within only 2 wk of a single TACE procedure.