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Supplementary Figure S3. BRG1-BRD inhibits γ-H2AX formation and DSB repair in irradiated MDA-MB-231 cells and this activity is enhanced by dimerization.

Improved therapeutic strategies are required to minimize side effects associated with radioiodine gene therapy to avoid unnecessary damage to normal cells and radiation-induced secondary malignancies. We previously reported that codon-optimized sodium iodide symporter (oNIS) enhances absorption of I-131 and that the brahma-associated gene 1 bromodomain (BRG1-BRD) causes inefficient DNA damage repair after high-energy X-ray therapy. To increase the therapeutic effect without applying excessive radiation, we considered the combination of oNIS and BRG1-BRD as gene therapy for the most effective radioiodine treatment. The antitumor effect of I-131 with oNIS or oNIS+BRD expression was examined by tumor xenograft models along with functional assays at the cellular level. The synergistic effect of both BRG1-BRD and oNIS gene overexpression resulted in more DNA double-strand breaks and led to reduced cell proliferation/survival rates after I-131 treatment, which was mediated by the p53/p21 pathway. We found increased p53, p21, and nucleophosmin 1 (NPM1) in oNIS- and BRD-expressing cells following I-131 treatment, even though the remaining levels of citrulline and protein arginine deiminase 4 (PAD4) were unchanged at the protein level.

Purpose Translocator protein 18-kDa (TSPO) positron emission tomography (PET) is a valuable tool to detect neuroinflammed areas in a broad spectrum of neurodegenerative diseases. However, the clinical application of second-generation TSPO ligands as biomarkers is limited because of the presence of human rs6971 polymorphism that affects their binding. Here, we describe the ability of a new TSPO ligand, [18F]BS224, to identify abnormal TSPO expression in neuroinflammation independent of the rs6971 polymorphism. Methods An in vitro competitive inhibition assay of BS224 was conducted with [3H]PK 11195 using membrane proteins isolated from 293FT cells expressing TSPO-wild type (WT) or TSPO-mutant A147T (Mut), corresponding to a high-affinity binder (HAB) and low-affinity binder (LAB), respectively. Molecular docking was performed to investigate the interaction of BS224 with the binding sites of rat TSPO-WT and TSPO-Mut. We synthesized a new 18F-labeled imidazopyridine acetamide ([18F]BS224) using boronic acid pinacol ester 6 or iodotoluene tosylate precursor 7, respectively, via aromatic 18F-fluorination. Dynamic PET scanning was performed up to 90 min after the injection of [18F]BS224 to healthy mice, and PET imaging data were obtained to estimate its absorbed doses in organs. To evaluate in vivo TSPO-specific uptake of [18F]BS224, lipopolysaccharide (LPS)-induced inflammatory and ischemic stroke rat models were used. Results BS224 exhibited a high affinity (Ki = 0.51 nM) and selectivity for TSPO. The ratio of IC50 values of BS224 for LAB to that for HAB indicated that the TSPO binding affinity of BS224 has low binding sensitivity to the rs6971 polymorphism and it was comparable to that of PK 11195, which is not sensitive to the polymorphism. Docking simulations showed that the binding mode of BS224 is not affected by the A147T mutation and consequently supported the observed in vitro selectivity of [18F]BS224 regardless of polymorphisms. With optimal radiochemical yield (39 ± 6.8%, decay-corrected) and purity (> 99%), [18F]BS224 provided a clear visible image of the inflammatory lesion with a high signal-to-background ratio in both animal models (BPND = 1.43 ± 0.17 and 1.57 ± 0.37 in the LPS-induced inflammatory and ischemic stroke rat models, respectively) without skull uptake. Conclusion Our results suggest that [18F]BS224 may be a promising TSPO ligand to gauge neuroinflammatory disease-related areas in a broad range of patients irrespective of the common rs6971 polymorphism.

Glioblastoma is the most common and fatal primary glioma and has a severe prognosis. It is a challenge for neurosurgeons to remove brain tumor tissues completely by resection. Meanwhile, fluorescence-guided surgery (FGS) is a technique used in glioma surgery to enhance the visualization of tumor edges to clarify the extent of tumor resection. Indocyanine green (ICG) is the only FDA-approved NIR fluorescent agent. It non-covalently binds to human serum albumin (HSA). Secreted protein acidic and rich in cysteine (SPARC) is an extracellular glycoprotein expressed in gliomas and binds to albumin, suggesting that it plays an important role in tumor uptake of the ICG-HSA complex. Here we demonstrate the binding properties of HSA or SPARC to ICG using surface plasmon resonance and saturation binding assay. According to in vitro and in vivo studies, the results showed that the uptake of ICG-HSA complex was higher in SPARC-expressing glioblastoma cell line and tumor region compared with the uptake of free ICG. Here, we visualized the SPARC-dependent uptake of ICG and ICG-HSA complex in U87MG. Our results demonstrated that the ICG-HSA complex is likely to be used as an efficient imaging agent targeting SPARC-expressing tumors, especially glioblastoma.

Although FDG-PET is widely used in cancer, its role in gastric cancer (GC) is still controversial due to variable [18F]fluorodeoxyglucose ([18F]FDG) uptake. Here, we sought to develop a genetic signature to predict high FDG-avid GC to plan individualized PET and investigate the molecular landscape of GC and its association with glucose metabolic profiles noninvasively evaluated by [18F]FDG-PET. Based on a genetic signature, PETscore, representing [18F]FDG avidity, was developed by imaging data acquired from thirty patient-derived xenografts (PDX). The PETscore was validated by [18F]FDG-PET data and gene expression data of human GC. The PETscore was associated with genomic and transcriptomic profiles of GC using The Cancer Genome Atlas. Five genes, PLS1, PYY, HBQ1, SLC6A5, and NAT16, were identified for the predictive model for [18F]FDG uptake of GC. The PETscore was validated in independent PET data of human GC with qRT-PCR and RNA-sequencing. By applying PETscore on TCGA, a significant association between glucose uptake and tumor mutational burden as well as genomic alterations were identified. Our findings suggest that molecular characteristics are underlying the diverse metabolic profiles of GC. Diverse glucose metabolic profiles may apply to precise diagnostic and therapeutic approaches for GC.

Recent advances in immune modulation have made impressive progress in cancer immunotherapy. Because dynamic nature of the immune response often makes it difficult to evaluate therapeutic outcomes, innovative imaging technologies have been developed to enable non-invasive visualization of immune cells and tumors in their microenvironment. This review summarizes the current tumor immunology and describes new innovative imaging methods with great potential to obtain non-invasive real-time insights into the complex functions of the immune system and into the management of cancer immunotherapy.

PurposeThe service recovery paradox (SRP) refers to a particular effect whereby an excellent recovery can turn angry and frustrated customers into loyal ones. Researchers who have studied the SRP have reported mixed findings, with some providing evidence in its support and others not finding any such evidence. To address this discrepancy, this study aims to investigate the SRP.Design/methodology/approachThis study re-examined the phenomenon of the SRP with a field study and provided further evidence in a subsequent experimental study in which the failure and recovery conditions were carefully manipulated.FindingsThe results of this study suggest that the SRP was observed in neither the field study nor the scenario experiment.Practical implicationsThis study can influence the current service management of restaurants with regard to service failures in several ways.Originality/valueThis research is a pioneering effort to examine the SRP by conducting both a field study and a scenario experiment.

Immune checkpoint inhibitors (ICIs) are widely used in cancer immunotherapy, requiring effective methods for response monitoring. This study evaluated changes in 18F-2-fluoro-2-deoxy-D-glucose (FDG) and 18F-fluorothymidine (FLT) uptake by tumors following ICI treatment as potential imaging biomarkers in mice. Tumor uptakes of 18F-FDG and 18F-FLT were measured and compared between the ICI treatment and control groups. A combined imaging index of glucose–thymidine uptake ratio (GTR) was defined and compared between groups. In the ICI treatment group, tumor growth was effectively inhibited, and higher proportions of immune cells were observed. In the early phase, 18F-FDG uptake was higher in the treatment group, whereas 18F-FLT uptake was not different. There was no difference in 18F-FDG uptake between the two groups in the late phase. However, 18F-FLT uptake of the control group was markedly increased compared with the ICI treatment group. GTR was consistently higher in the ICI treatment group in the early and late phases. After ICI treatment, changes in tumor cell proliferation were observed with 18F-FLT, whereas 18F-FDG showed altered metabolism in both tumor and immune cells. A combination of 18F-FLT and 18F-FDG PET, such as GTR, is expected to serve as a potentially effective imaging biomarker for monitoring ICI treatment.

The 18 kDa translocator protein (TSPO) has been proposed as a biomarker for the detection of neuroinflammation. Although various PET probes targeting TSPO have been developed, a highly selective probe for detecting TSPO is still needed because single nucleotide polymorphisms in the human TSPO gene greatly affect the binding affinity of TSPO ligands. Here, we describe the visualization of neuroinflammation with a multimodality imaging system using our recently developed TSPO-targeting radionuclide PET probe [18F]CB251, which is less affected by TSPO polymorphisms. Methods: To test the selectivity of [18F]CB251 for TSPO polymorphisms, 293FT cells expressing polymorphic TSPO were generated by introducing the coding sequences of wild-type (WT) and mutant (Alanine → Threonine at 147th Amino Acid; A147T) forms. Competitive inhibition assay was conducted with [3H]PK11195 and various TSPO ligands using membrane proteins isolated from 293FT cells expressing TSPO WT or mutant-A147T, representing high-affinity binder (HAB) or low-affinity binder (LAB), respectively. IC50 values of each ligand to [3H]PK11195 in HAB or LAB were measured and the ratio of IC50 values of each ligand to [3H]PK11195 in HAB to LAB was calculated, indicating the sensitivity of TSPO polymorphism. Cellular uptake of [18F]CB251 was measured with different TSPO polymorphisms, and phantom studies of [18F]CB251-PET using 293FT cells were performed. To test TSPO-specific cellular uptake of [18F]CB251, TSPO expression was regulated with pCMV-TSPO (or shTSPO)/eGFP vector. Intracranial lipopolysaccharide (LPS) treatment was used to induce regional inflammation in the mouse brain. Gadolinium (Gd)-DOTA MRI was used to monitor the disruption of the blood-brain barrier (BBB) and infiltration by immune cells. Infiltration of peripheral immune cells across the BBB, which exacerbates neuroinflammation to produce higher levels of neurotoxicity, was also monitored with bioluminescence imaging (BLI). Peripheral immune cells isolated from luciferase-expressing transgenic mice were transferred to syngeneic inflamed mice. Neuroinflammation was monitored with [18F]CB251-PET/MR and BLI. To evaluate the effects of anti-inflammatory agents on intracranial inflammation, an inflammatory cytokine inhibitor, 2-cyano-3, 12-dioxooleana-1, 9-dien-28-oic acid methyl ester (CDDO-Me) was administered in intracranial LPS challenged mice. Results: The ratio of IC50 values of [18F]CB251 in HAB to LAB indicated similar binding affinity to WT and mutant TSPO and was less affected by TSPO polymorphisms. [18F]CB251 was specific for TSPO, and its cellular uptake reflected the amount of TSPO. Higher [18F]CB251 uptake was also observed in activated immune cells. Simultaneous [18F]CB251-PET/MRI showed that [18F]CB251 radioactivity was co-registered with the MR signals in the same region of the brain of LPS-injected mice. Luciferase-expressing peripheral immune cells were located at the site of LPS-injected right striatum. Quantitative evaluation of the anti-inflammatory effect of CDDO-Me on neuroinflammation was successfully monitored with TSPO-targeting [18F]CB251-PET/MR and BLI. Conclusion: Our results indicate that [18F]CB251-PET has great potential for detecting neuroinflammation with higher TSPO selectivity regardless of polymorphisms. Our multimodal imaging system, [18F]CB251-PET/MRI, tested for evaluating the efficacy of anti-inflammatory agents in preclinical studies, might be an effective method to assess the severity and therapeutic response of neuroinflammation.

The corporate social responsibility (CSR) literature documents inconsistent results regarding the relationship between CSR and employees’ organizational citizenship behavior (OCB). Additionally, few empirical studies examine the mediating factors explaining how employees’ CSR perception affects their attitudes and behaviors. Thus, we propose an extended M-R model linking hotels’ CSR activities, employees’ perceived meaningfulness of work and organizational pride, and organizational citizenship behavior. We recruited 363 Chinese hotel employees in Guangzhou for our sample and employed partial least squares structural equation modeling for our analysis. The results showed that CSR practices benefiting both internal (i.e., employees) and external stakeholders (i.e., community and customers) positively influenced employees’ organizational pride. However, only CSR activities benefiting customers significantly affected employees’ perceived meaningfulness of work. CSR influenced employees’ OCB through meaningfulness of work and organizational pride. The findings provide hotel management insights regarding the development of CSR initiatives to enhance employees’ organizational attitudes and work performance. The study limitations are that the survey methodology has weaknesses and that we do not consider the influence of corporate culture on OCB.

Claudin-3 (CLDN3), a tight junction protein, regulates cell-to-cell interactions in epithelial or endothelial cell sheets. During tumorigenesis, epithelial cells are transformed, and tumor cells proliferate through out-of-plane division, resulting in external exposure of CLDN3. Since alterations of CLDN3 expression are associated with cancer progression and higher CLDN3 expression is observed in most ovarian cancers, we tested the feasibility of using a CLDN3-specific antibody as a novel imaging tracer.After reducing the CLDN3-specific antibodies to expose the -SH groups, click chemistry was used to conjugate the radioactive isotopeThe labeling efficiency of NOTA-We have successfully conjugated a radioisotope and a fluorescent protein with CLDN3-specific antibodies and verified the specific binding of labeled antibodies to OVCAR-3 tumors in a mouse model. Our data suggested that CLDN3-specific human monoclonal antibodies could be used as a useful theranostic tracer.

Hypoxic conditions, which large or infiltrative hypovascular tumors may encounter, also produce acidic environments. Carbonic anhydrase-IX (CA-IX), an enzyme involved in lowering pH, is overexpressed in hepatocellular carcinoma (HCC). In the present study, whether inhibition of CA-IX enhances the efficacy of a hexokinase II inhibitor in an in vivo murine model was examined and its prognostic implication in HCC patients was investigated. CA-IX expression was evaluated using quantitative real-time PCR and western blot analysis using human HCC cell lines. 3-bromopyruvate (3-BP), a hexokinase II inhibitor, and acetazolamide, a carbonic anhydrase inhibitor, were used to target hexokinase II and CA-IX in vitro and in vivo, respectively. A human HCC cell line (Huh-7) was tested as a subcutaneous tumor model in BALB/c nu/nu mice. The prognostic role of CA-IX was evaluated in the TCGA database. Quantitative real-time PCR and western blot analysis revealed that CA-IX expression was activated in the presence of 3-BP. Further analysis showed that introducing an additional stress by treating the orally active CA-IX inhibitor (acetazolamide) can synergistically increase the efficacy of 3-BP in vivo, which was confirmed using a mouse model. We also found that HCC patients with high CA-IX expression show poor overall survival in TCGA database. These results indicate CA-IX is a promising therapeutic target for enhancing the efficacy of 3-BP and can be a prognostic factor for HCC.

2-Deoxy-2-[18F]fluoro-d-glucose ([18F]FDG) accumulation in inflammatory lesions can confound the diagnosis of cancer. In this study, we investigated [18F]FDG accumulation and efflux in relation to the genes and proteins involved in glucose metabolism in murine inflammation and cancer models. [18F]FDG accumulation and [18F]FDG efflux were measured in cancer cells (breast cancer, glioma, thyroid cancer, and hepatoma cells) and RAW 264.7 cells (macrophages) activated with lipopolysaccharide (LPS). The levels of mRNA expression were measured by real-time quantitative PCR (qPCR). The expression of glucose metabolism–related proteins was detected by western blotting. Dynamic [18F]FDG positron emission tomography-computed tomography (PET/CT) images were acquired for 2 h in tumor-bearing BALB/c nude mice and inflammatory mice induced by turpentine oil. [18F]FDG accumulation in MDA-MB-231 (breast cancer) increased with time, but that of HepG2 (hepatoma) reached a constant level after 120 min. [18F]FDG efflux in HepG2 was faster than that in MDA-MB-231. HepG2 strongly expressed glucose-6-phosphatase (G6Pase) compared with MDA-MB-231. [18F]FDG accumulation increased with time, and [18F]FDG efflux accelerated after the activation of RAW 264.7 cells. The expression levels of G6Pase, glucose transporter1 and glucose transporter3 (GLUT1 and GLUT3), and hexokinase II (HK II) increased after the activation of RAW 264.7 cells. [18F]FDG efflux in activated macrophages was faster than that in MDA-MB-231 cancer cells. MDA-MB-231 strongly expressed HK II protein compared with the activated RAW 264.7. In murine models, [18F]FDG accumulation in MDA-MB-231 cancer and inflammatory lesions increased with time, but that in HepG2 tumor increased until 20–30 min (SUVmeans ± SD (tumor/muscle), 3.0 ± 1.3) and then decreased (2.1 ± 0.9 at 110–120 min). There was no difference in the pattern of [18F]FDG accumulation with time in MDA-MB-231 tumors and inflammatory lesions. We found that [18F]FDG efflux accelerated in activated macrophages reflecting increased G6Pase expression after activation and lower expression of HK II protein than that in MDA-MB-231 cancer cells.

Prostate-specific membrane antigen (PSMA) is overexpressed in most prostate adenocarcinoma (AdPC) cells and acts as a target for molecular imaging. However, some case reports indicate that PSMA-targeted imaging could be ineffectual for delineation of neuroendocrine (NE) prostate cancer (NEPC) lesions due to the suppression of the PSMA gene (FOLH1). These same reports suggest that targeting somatostatin receptor type 2 (SSTR2) could be an alternative diagnostic target for NEPC patients. This study evaluates the correlation between expression ofFOLH1, NEPC marker genes andSSTR2. We evaluated the transcript abundance forFOLH1andSSTR2genes as well as NE markers across 909 tumors. A significant suppression ofFOLH1in NEPC patient samples and AdPC samples with high expression of NE marker genes was observed. We also investigated protein alterations of PSMA and SSTR2 in an NE-induced cell line derived by hormone depletion and lineage plasticity by loss of p53. PSMA is suppressed following NE induction and cellular plasticity in p53-deficient NEPC model. The PSMA-suppressed cells have more colony formation ability and resistance to enzalutamide treatment. Conversely, SSTR2 was only elevated following hormone depletion. In 18 NEPC patient-derived xenograft (PDX) models we find a significant suppression ofFOLH1and amplification ofSSTR2expression. Due to the observedFOLH1-supressed signature of NEPC, this study cautions on the reliability of using PMSA as a target for molecular imaging of NEPC. The observed elevation ofSSTR2in NEPC supports the possible ability of SSTR2-targeted imaging for follow-up imaging of low PSMA patients and monitoring for NEPC development.

Human serum albumin (HSA) is the most abundant plasma protein. The main reason for using HSA as a versatile tool for drug delivery is based on its ability to accumulate in tumors. However, the mechanism of albumin accumulation in tumors is not yet clear. Many researchers using HSA as a drug-carrier have focused on the passive tumor targeting by enhanced permeability and retention (EPR) effect, while other investigators proposed that albumin binding proteins mediate albumin accumulation in tumors. We investigated whether HSA accumulation in tumors is mediated by the EPR effect or by secreted protein acidic and rich in cysteine (SPARC), which is known to be an albumin-binding protein. Methods: To investigate the role of SPARC on HSA accumulation in tumors, we compared HSA uptake in U87MG glioblastoma cells with different SPARC expression. U87MG cells generally express high levels of SPARC and were, therefore, used as SPARC-rich cells. SPARC-less U87MG (U87MG-shSPARC) cells were established by viral-shSPARC transduction. We detected cellular uptake of fluorescence-labeled HSA by confocal microscopy in U87MG and U87MG-shSPARC cells. To demonstrate the mechanism of HSA accumulation in tumors, we injected FNR648-labeled HSA and FITC-labeled dextran in U87MG and U87MG-shSPARC tumor-bearing mice and observed their micro-distribution in tumor tissues. Results: HSA was internalized in cells by binding with SPARC in vitro. HSA accumulation in U87MG glioma was associated with SPARC expression in vivo. FITC-dextran was distributed in U87MG tumors in the vicinity of blood vessels. The distribution of HSA, on the other hand, was observed in the regions remote from blood vessels of U87MG tumor tissues but not in U87MG-shSPARC tumor tissues. Conclusion: Our results demonstrate that the tumor-distribution of HSA is affected not only by the EPR-effect but also by SPARC expression. SPARC enhances HSA accumulation in U87MG glioma and mediates active targeting of HSA in tumors.

Although FDG-PET is widely used in cancer, its role in gastric cancer (GC) is still controversial due to variable [Based on a genetic signature, PETscore, representing [Five genes, PLS1, PYY, HBQ1, SLC6A5, and NAT16, were identified for the predictive model for [Our findings suggest that molecular characteristics are underlying the diverse metabolic profiles of GC. Diverse glucose metabolic profiles may apply to precise diagnostic and therapeutic approaches for GC.

Purpose Translocator protein 18-kDa (TSPO) positron emission tomography (PET) is a valuable tool to detect neuroinflammed areas in a broad spectrum of neurodegenerative diseases. However, the clinical application of second-generation TSPO ligands as biomarkers is limited because of the presence of human rs6971 polymorphism that affects their binding. Here, we describe the ability of a new TSPO ligand, [18F]BS224, to identify abnormal TSPO expression in neuroinflammation independent of the rs6971 polymorphism. Methods An in vitro competitive inhibition assay of BS224 was conducted with [3H]PK 11195 using membrane proteins isolated from 293FT cells expressing TSPO-wild type (WT) or TSPO-mutant A147T (Mut), corresponding to a high-affinity binder (HAB) and low-affinity binder (LAB), respectively. Molecular docking was performed to investigate the interaction of BS224 with the binding sites of rat TSPO-WT and TSPO-Mut. We synthesized a new 18F-labeled imidazopyridine acetamide ([18F]BS224) using boronic acid pinacol ester 6 or iodotoluene tosylate precursor 7, respectively, via aromatic 18F-fluorination. Dynamic PET scanning was performed up to 90 min after the injection of [18F]BS224 to healthy mice, and PET imaging data were obtained to estimate its absorbed doses in organs. To evaluate in vivo TSPO-specific uptake of [18F]BS224, lipopolysaccharide (LPS)-induced inflammatory and ischemic stroke rat models were used. Results BS224 exhibited a high affinity (Ki = 0.51 nM) and selectivity for TSPO. The ratio of IC50 values of BS224 for LAB to that for HAB indicated that the TSPO binding affinity of BS224 has low binding sensitivity to the rs6971 polymorphism and it was comparable to that of PK 11195, which is not sensitive to the polymorphism. Docking simulations showed that the binding mode of BS224 is not affected by the A147T mutation and consequently supported the observed in vitro selectivity of [18F]BS224 regardless of polymorphisms. With optimal radiochemical yield (39 ± 6.8%, decay-corrected) and purity (> 99%), [18F]BS224 provided a clear visible image of the inflammatory lesion with a high signal-to-background ratio in both animal models (BPND = 1.43 ± 0.17 and 1.57 ± 0.37 in the LPS-induced inflammatory and ischemic stroke rat models, respectively) without skull uptake. Conclusion Our results suggest that [18F]BS224 may be a promising TSPO ligand to gauge neuroinflammatory disease-related areas in a broad range of patients irrespective of the common rs6971 polymorphism. Supplementary Information The online version contains supplementary material available at 10.1007/s00259-021-05617-4.

Purpose Translocator protein 18-kDa (TSPO) positron emission tomography (PET) is a valuable tool to detect neuroinfammed areas in a broad spectrum of neurodegenerative diseases. However, the clinical application of second-generation TSPO ligands as biomarkers is limited because of the presence of human rs6971 polymorphism that afects their binding. Here, we describe the ability of a new TSPO ligand, [ 18F]BS224, to identify abnormal TSPO expression in neuroinfammation independent of the rs6971 polymorphism. Methods An in vitro competitive inhibition assay of BS224 was conducted with [3H]PK 11195 using membrane proteins isolated from 293FT cells expressing TSPO-wild type (WT) or TSPO-mutant A147T (Mut), corresponding to a high-afnity binder (HAB) and low-afnity binder (LAB), respectively. Molecular docking was performed to investigate the interaction of BS224 with the binding sites of rat TSPO-WT and TSPO-Mut. We synthesized a new 18F-labeled imidazopyridine acetamide ([18F]BS224) using boronic acid pinacol ester 6 or iodotoluene tosylate precursor 7, respectively, via aromatic 18F-fuorination. Dynamic PET scanning was performed up to 90 min after the injection of [18F]BS224 to healthy mice, and PET imaging data were obtained to estimate its absorbed doses in organs. To evaluate in vivo TSPO-specifc uptake of [18F] BS224, lipopolysaccharide (LPS)-induced infammatory and ischemic stroke rat models were used. Results BS224 exhibited a high afnity (Ki=0.51 nM) and selectivity for TSPO. The ratio of IC50 values of BS224 for LAB to that for HAB indicated that the TSPO binding afnity of BS224 has low binding sensitivity to the rs6971 polymorphism and it was comparable to that of PK 11195, which is not sensitive to the polymorphism. Docking simulations showed that the binding mode of BS224 is not afected by the A147T mutation and consequently supported the observed in vitro selectivity of [ 18F]BS224 regardless of polymorphisms. With optimal radiochemical yield (39±6.8%, decay-corrected) and purity (>99%), [18F]BS224 provided a clear visible image of the infammatory lesion with a high signal-to-background ratio in both animal models (BPND =1.43±0.17 and 1.57±0.37 in the LPS-induced infammatory and ischemic stroke rat models, respectively) without skull uptake. Conclusion Our results suggest that [ 18F]BS224 may be a promising TSPO ligand to gauge neuroinfammatory disease-related areas in a broad range of patients irrespective of the common rs6971 polymorphism.

There has been considerable interest in the clinical use of exosomes as delivery vehicles for treatments as well as for promising diagnostic biomarkers, but the physiological distribution of exosomes must be further elucidated to validate their efficacy and safety. Here, we aimed to develop novel methods to monitor exosome biodistribution in vivo using positron emission tomography (PET) and optical imaging. Exosomes were isolated from cultured mouse breast cancer cells and labeled for PET and optical imaging. In mice, radiolabeled and fluorescently labeled exosomes were injected both via lymphatic and hematogenous metastatic routes. PET and fluorescence images were obtained and quantified. Radioactivity and fluorescence intensity of ex vivo organs were measured. PET signals from exosomes in the lymphatic metastatic route were observed in the draining sentinel lymph nodes. Immunohistochemistry revealed greater exosome uptake in brachial and axillary versus inguinal lymph nodes. Following administration through the hematogenous metastasis pathway, accumulation of exosomes was clearly observed in the lungs, liver, and spleen. Exosomes from tumor cells were successfully labeled with 64Cu (or 68Ga) and fluorescence and were visualized via PET and optical imaging, suggesting that this simultaneous and rapid labeling method could provide valuable information for further exosome translational research and clinical applications.

Despite improved therapeutic efficacy of the locked nucleic acid (LNA)- and peptide nucleic acid (PNA)-modified antisense microRNAs (anti-miRs), their wider application in clinical practice is still not thoroughly investigated. This study aimed to investigate the stability and therapeutic efficacy of the modified LNA- and PNA-type anti-miRs in a murine prostate cancer model under various treatment conditions. After verifying the anti-cancer potential of anti-miR21 by targeting tumor suppressor PTEN, the potential of the modified LNA- and PNA-type anti-miR21s was compared in vitro and in vivo. We found that PNA-type anti-miR21 showed better stability and therapeutic efficacy in the xenografted mouse tumor model than the LNA-type anti-miR21. Furthermore, PNA-type anti-miR21 treatment showed reduced tumor metastasis. This study may serve as a ground for exploring diverse choices in therapeutic oligonucleotide modification techniques to improve cancer treatment.

Clinically, there has been significant interest in the use of exosomes for diagnostic applications as promising biomarkers and therapeutic applications as therapeutic vehicles. However, knowledge ofin vivophysiological biodistribution of exosomes was difficult to assess until now. Physiological distribution of exosomes in the body must be elucidated for clinical application. In this study, we aimed to develop reliable and novel methods to monitor biodistribution of exosomes usingin vivoPET and optical imaging.MethodsExosomes were isolated from cultured medium of 4T1, mouse breast cancer cells. Exosomes were labeled with Cy7 and64Cu (or68Ga). In mice, radio/fluorescent dye-labeled exosomes were injected through the lymphatic routes (footpad injection) and hematogenous metastatic routes (tail vein injection). Fluorescence and PET images were obtained and quantified. Radio-activity ofex vivoorgans was measured by gamma counter.ResultsPET signals from exosomes in the lymphatic metastatic route were observed in the draining lymph nodes, which are not distinguishable with optical imaging. Immunohistochemistry revealed greater uptake of exosomes in brachial and axillary lymph nodes than inguinal lymph node. After administration through the hematogenous metastasis pathway, accumulation of exosomes was clearly observed in PET images in the lungs, liver, and spleen, showing results similar toex vivogamma counter data.ConclusionExosomes from tumor cells were successfully labeled with64Cu (or68Ga) and visualized by PET imaging. These results suggest that this cell type-independent, quick, and easy exosome labeling method using PET isotopes could provide valuable information for further application of exosomes in the clinic.

338 Background: HER2 is highly overexpressed in many kinds of cancers with a poor prognosis. Recently, near-infrared (NIR) fluorescence-based imaging is a growing field for both pre-clinical and clinical application. In this study, we aimed to synthesize Human Epidermal Receptor2 (HER2)-specific near-infrared (NIR) fluorescence probes and evaluate their applicability in cancer-specific image-guided surgeries using an animal model. Methods: An NIR dye emitting light of 800 nm (IRDye800CW, Li-COR, USA) was conjugated to trastuzumab and HER2-specific affibody using click mechanism. HER2 affinity was assessed by the surface plasmon resonance technique. HER2 positive/negative gastric cancer cell lines (NCI-N87 and SNU-601) were subcutaneously implanted into female BALB/c-nu (6 to 8 weeks old) mice. The biodistribution and fluorescence signal intensity were measured by Lumina II (Perkin Elmer, MA, USA) and a laparoscopic NIR camera (InTheSmart, Seoul, Korea) after injecting the probes intravenously. Results: Trastuzumab-IRDye800CW showed higher affinity to HER2 (K(D) = 2.093(3)pM) than unconjugated trastuzumab(K(D) = 25.75pM). The significant signal of fluorescence was targeted to the HER2-positive tumors at 24hr after injection, while no or low signal retention was observed in negative group. The peak appears at 24hr after injection. On the other hand, small difference of affinity was shown between HER2-target affibody-IRDye800CW (K(D) = 4.71nM) and unlabeled pure affibody(K(D) = 1.42nM). The renal clearance of HER2-target affibody conjugated with IRDye800CW was so fast that we could not detect the signal. Conclusions: Our results suggest that trastuzumab conjugated with IRDye800CW can be a feasible tool to monitor HER2 status in pre-clinical cancer imaging. Moreover, this probe can provide complementary means for assessment of HER2 expression in gastric cancer patients and/or be used to further detection of HER2-positive lesions during image-guided surgery.

Rheumatoid arthritis (RA) is a chronic disease with systemic inflammation resulting in destruction of multiple articular cartilages and bones. Activated macrophage plays a pivotal role during the disease course and has been one of main targets to inhibit inflammatory reaction of RA by using biological disease-modifying anti-rheumatic drugs (bDMARDs). 18F-FEDAC is one of PET imaging agents targeting TSPO, which is overexpressed in activated macrophages. The aim of this study was to evaluate the roles of 18F-FEDAC PET as an in vivo imaging of activated macrophages on etanercept (ETN), a TNF-antagonist as one of bDMARDs in collagen induced arthritis mice. In RAW 264.7 cells, the expressions of TSPO as well as iNOS and infiltrated nucleus of NF-κB were induced by activation with lipopolysaccharide and interferon-gamma. TSPO expression was slightly attenuated by ETN treatment, not by methotrexate (MTX) as a cytotoxic agent. However, cell uptake of 18F-FEDAC did not show significant changes according to both of the treatments. Similarly in CIA mice, 18F-FEDAC uptake in inflamed paws on PET imaging did not show significant changes during both of the treatments, contrary to the uptake decrease of 18F-FDG, a glucose analog to reflect metabolic or active inflammatory activity. Interestingly, when we divided joints according to the degree of 18F-FEDAC uptake before ETN treatment, the joints of high 18F-FEDAC uptake showed better response to ETN than the joints with low 18F-FEDAC uptakes. In case of 18F-FDG, there was no such kinds of patterns. We can speculate that 18F-FEDAC PET imaging may identify activated macrophage-induced arthritis because that 18F-FEDAC can reflect activated macrophages, which is the therapeutic target of ETN by TNF antagonistic effect. Thus, in vivo imaging using 18F-FEDAC may be used as a predictor of therapeutic effects among those kinds of bDMARDs having anti-inflammatory actions to inhibit activated macrophage.

The current study examines CSR issues in the casino context from employees' perspectives, especially due to the essential role of employees for business success in service industries. This study first investigates the main effect of employees' perceived CSR on organizational commitment, mediated by job satisfaction. This study further explores this mediated relationship by incorporating employees' perceptions of the casino industry as a moderator, based on social identity theory which appears particularly well-suited to support the proposed moderating effect in the casino context due to the industry's controversial nature. The results support the proposed hypotheses, making theoretical and managerial implications, especially with the finding of employees' perceptions of the casino industry as a new and significant moderator. Such finding also makes a managerial contribution by suggesting the importance of creating and/or improving employees' general perceptions about the casino industry which will eventually help to enhance the employees' commitment to their organization.

Although glucose transporter 1 (GLUT1) and hexokinase 2 (HK2) are known as major proteins involved in the molecular mechanisms for accumulating 2-deoxy-2-[18F]fluoro-D-glucose ([18F]FDG) in cancer cells, sometimes, [18F] FDG accumulation cannot be explained by the expression of these two proteins. We investigated the involvement of adenine nucleotide translocase 2 (ANT2), which catalyzes ADP/ATP exchange at the mitochondrial inner membrane, in [18F] FDG accumulation. ANT2 expression was evaluated in various cancer cell lines and human cancer tissues (microarrays) using western blot and immunohistochemical (IHC) staining, respectively. The expression levels of ANT2 were compared to [18F] FDG accumulation and pathologic findings, including differentiation grade. Additionally, we modulated ANT2 expression levels using ANT2 siRNA and an ANT2 expression vector in cancer cells and murine xenografted tumors. [18F] FDG accumulation correlated with ANT2 expression in various cancer cell lines; this was not explained by GLUT1 and/or HK2 expression. At both the cell and tissue levels, ANT2 expression was high in less-differentiated or more malignant type of cancers. [18F] FDG accumulation changed according to the modulation of the ANT2 expression level. In various cancer cells and tissues, the expression levels of ANT2 explained [18F] FDG accumulation better than those of GLUT1 and HK2. ANT2 can be used as a marker of dedifferentiated pathology and aggressiveness of cancer.

Purpose Study on the pathogen and the pathogen-related disease require the information at both cellular and organism level. However, lack of appropriate high-quality antibodies and the difference between the experimental animal models make it difficult to analyze in vivo mechanism of pathogen-related diseases. For more reliable research on the infection and immune-response of pathogen-related diseases, accurate analysis is essential to provide spatiotemporal information of pathogens and immune activity to avoid false-positive or mis-interpretations. In this regards, we have developed a method for tracking Francisella tularensis in the animal model without using the specific antibodies for the F. tularensis. Materials and methods A dual reporter plasmid using GFP-Lux with putative bacterioferritin promoter (pBfr) was constructed and transformed to F. tularensis live vaccine strain to generate F. tularensis LVS (FtLVS)-GFP-Lux for both fluorescence and bioluminescence imaging. For vaccination to F. tularensis infection, FtLVS and lipopolysaccharide (LPS) from FtLVS were used. Results We visualized the bacterial replication of F. tularensis in the cells using fluorescence and bioluminescence imaging, and traced the spatio-temporal process of F. tularensis pathogenesis in mice. Vaccination with LPS purified from FtLVS greatly reduced the bacterial replication of FtLVS in animal model, and the effect of vaccination was also successfully monitored with in vivo imaging. Conclusion We successfully established dual reporter labeled F. tularensis for cellular and whole body imaging. Our simple and integrated imaging analysis system would provide useful information for in vivo analysis of F. tularensis infection as well as in vitro experiments, which have not been fully explained yet with various technical problems.

Activated macrophages have been known to play pivotal roles in the pathogenesis of rheumatoid arthritis (RA). 18F-FEDAC (N-benzyl-N-methyl-2-[7,8-dihydro-7-(2-18F-fluoroethyl)-8-oxo-2-phenyl-9H-purin-9-yl]acetamide) is a radiolabeled ligand for the 18-kDa translocator protein (TSPO), which is abundant in activated macrophages. We evaluated the feasibility of using 18F-FEDAC in a murine RA model. Methods: RAW 264.7 mouse macrophages were activated by lipopolysaccharide. TSPO expression levels in activated and inactivated macrophages were measured by quantitative polymerase chain reaction and Western blotting. The cellular uptake and specific binding of 18F-FEDAC were measured using a γ-counter. For the in vivo study, collagen-induced arthritis (CIA) was developed in DBA/1 mice, and the clinical score for arthritis was measured regularly. 18F-FEDAC and 18F-FDG PET images were acquired on days 23 and 37 after the first immunization. Histologic examinations were performed to evaluate macrophages and TSPO expression. Results: We found increased TSPO messenger RNA and protein expression in activated macrophages. Uptake of 18F-FEDAC in activated macrophages was higher than that in nonactivated cells and was successfully blocked by the competitor, PK11195. In CIA mice, joint swelling was apparent on day 26 after the first immunization, and the condition worsened by day 37. 18F-FEDAC uptake by arthritic joints increased early on (day 23), whereas 18F-FDG uptake did not. However, 18F-FDG uptake by arthritic joints markedly increased at later stages (day 37) to a higher level than 18F-FEDAC uptake. The 18F-FEDAC uptake correlated weakly with summed severity score (P = 0.019, r = 0.313), whereas the 18F-FDG uptake correlated strongly with summed severity score (P < 0.001, r = 0.897). Histologic sections of arthritic joints demonstrated an influx of macrophages compared with that in normal joints. Conclusion:18F-FEDAC enabled the visualization of active inflammation sites in arthritic joints in a CIA model by targeting TSPO expression in activated macrophages. The results suggest the potential usefulness of 18F-FEDAC imaging in the early phase of RA.

Unfamiliar aspects of ethnic foods are found to enhance both perceived authenticity and perceived risk. Whereas the former positively influences customers’ evaluations of foods and induces consumption behavior, the latter negatively affects customer behavior. Due to these conflicting effects, ethnic restaurateurs face a dilemma of whether to emphasize unfamiliar aspects to increase customer perception of authenticity or to eliminate them to reduce perceived risk. Thus, this study investigated (1) the effects of food information on perceived authenticity and perceived risk and (2) the comparative influences of perceived authenticity and perceived risk on customer satisfaction and purchase intention. The results revealed that unfamiliar ingredients influenced both perceived authenticity and perceived risk, whereas unfamiliar dish names only affected perceived authenticity. Additionally, the results support the notion that ethnic restaurant managers should endeavor to convey the uniqueness of their ethnic foods to maximize the effect of perceived authenticity over that of perceived risk.

The extant literature reports inconsistent results regarding the effect of employee gender on customers' attitudes and behaviors. Furthermore, the previous research on congruency bias has mainly focused on female-dominant jobs. To address these issues, this study examined how customers' gender stereotypes affect their service evaluations and revisit intentions. Grounded in role congruity theory, congruency bias, and justification-suppression models, the results showed that employee gender and gender congruency with the norm for the job have differential impacts on customer evaluations. Furthermore, this study found asymmetric impacts of congruency bias on customer experiences. For example, customers showed lower satisfaction and revisit intention when female employees failed in a male-dominant job than when their male counterparts did. However, they showed equal levels of satisfaction and revisit intention when male and female employees failed in a female-dominant job. The results provide important insights into how to reduce customers' gender bias toward different gender-dominated jobs.

The extant literature documents the significance of an auspicious naming strategy using lucky names and/or numbers to promote consumer behavior. However, this practice has rarely been examined in the hospitality literature. Therefore, by integrating value-belief-norm (VBN) theory and the theory of reasoned action (TRA), this study developed a consumer behavior model to examine the influence of customer perceptions of auspiciously named foods on their attitudes and purchase intention. In total, 475 surveys were collected using a panel data service in China. This study examined the structural relationships among personal values, superstitious beliefs, attitudes, social norms, and purchase intentions. The results revealed the salient role of social norms in determining auspicious consumption. Additionally, age was found to moderate the effects of attitude and social norms on purchase intention. These results provide important insight into how to develop effective marketing strategies to increase the sales of auspiciously named food products.

Introduction: Claudin-3 (CLDN3), a tight junction protein, regulates cell-to-cell interactions in epithelial or endothelial cell sheets. During tumorigenesis, epithelial cells are transformed and tumor cells proliferate through out-of-plane division, resulting in external exposure of CLDN3. This alteration of CLDN3 expression is associated with cancer progression, correlating with malignancy in various carcinomas. Since CLDN3 is particularly overexpressed in most ovarian cancers and used as an effective diagnostic marker, we tested the possibility of using a CLDN3-specific antibody as a novel imaging probe. Materials and Methods: After reducing the CLDN3 specific antibody to expose the -SH group, click chemistry was used to conjugate radioactive isotope 111In or fluorescent protein FNR648. Human ovarian cancer OVCAR-3 cells and human glioblastoma (U87MG) cells were used as CLDN3 positive and negative cells, respectively. Flow cytometry was used to measure the binding of CLDN3 IgG1 monoclonal antibody to theses cell lines. To establish xenograft model, OVCAR-3 cells were injected subcutaneously into the mice. 111In-labeled CLDN3 antibody (370 kBq/50 μl) was administered intravenously to mice with. After 24 hours, organs including tumor were excised and measured with a γ-counter. Images were acquired with IVIS and SPECT/CT. Results: The labeling efficiency of NOTA-111In or antibody-NOTA-111In was 98.52% or 100%, respectively. FNR648 labeled CLDN3 antibody was bound to the cell surface of OVCAR-3 at 83.4% and to U87MG at 5.7%, respectively. In OVCAR-3 tumor xenografted mice, mice injected with CLDN3 antibody showed 2.5-fold higher tumor uptake (20.4 ± 7.4% ID/g) than mice injected with human IgG (8.8 ± 2.6% ID/g) at 24 hour p.i. The fluorescence signal of CLDN3 antibody peaked at 24 hour p.i. Conclusion: We successfully conjugated radioisotope and fluorescent protein using the CLDN3 specific antibody. Since the specific binding of CLDN3 antibodies to OVCAR-3 tumors has been validated in a mouse model and diagnostic radionuclides can be replaced with therapeutic radionuclides, this human monoclonal antibody could be used as a useful theranostic probe. Citation Format: Sera Oh, Hobin Yang, Ho Rim Oh, Chul-Hee Lee, Young-Hwa Kim, Gi Jeong Cheon, Keon Wook Kang, Young Kee Shin, Hyewon Youn. A human Claudin-3 monoclonal antibody as a potential multimodal theranostic probe in ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2820.