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The development of molecular imaging probes to identify key cellular changes within lung metastases may lead to noninvasive detection of metastatic lesions in the lung. In this study, we constructed a macrophage-targeted clickable albumin nanoplatform (CAN) decorated with mannose as the targeting ligand using a click reaction to maintain the intrinsic properties of albumin

Background Immunotherapy with clodronate-encapsulated liposomes, which induce macrophage depletion, has been studied extensively. However, previously reported liposomal formulation-based drugs (Clodrosome® and m-Clodrosome®) are limited by their inconsistent size and therapeutic efficacy. Thus, we aimed to achieve consistent therapeutic effects by effectively depleting macrophages with uniform-sized liposomes.Results We developed four types of click chemistry-based liposome nanoplatforms that were uniformly sized and encapsulated with clodronate, for effective macrophage depletion, followed by conjugation with Man-N3 and radiolabeling. Functionalization with Man-N3 improves the specific targeting of M2 macrophages, and radioisotope labeling enables in vivo imaging of the liposome nanoplatforms. The functionalized liposome nanoplatforms are stable under physiological conditions. The difference in the biodistribution of the four liposome nanoplatforms in vivo were recorded using positron emission tomography imaging. Among the four platforms, the clodronate-encapsulated mannosylated liposome effectively depleted M2 macrophages in the normal liver and tumor microenvironment ex vivo compared to that by Clodrosome® and m-Clodrosome®.Conclusion The newly-developed liposome nanoplatform, with finely tuned size control, high in vivo stability, and excellent ex vivo M2 macrophage targeting and depletion effects, is a promising macrophage-depleting agent.

Supplementary Figures 1-6. Supplementary Figure 1. Mouse models of RIPF. Supplementary Figure 2. The ocurrence of EndMT and an EMT in the development of RIPF. Supplementary Figure 3. 2-ME inhibits EMT in the development of RIPF. Supplementary Figure 4. 2-ME inhibits RIPF. Supplementary Figure 5. H&E staining in human RIPF tissues Supplementary Figure 6. Whole-body or thoracic irradiation of mice induces apoptosis and detachment of lung endothelial cells.

Purpose: Radiation-induced pulmonary fibrosis (RIPF) is a late side effect of thoracic radiotherapy. The purpose of our study was to gain further insight into the development of RIPF.Experimental Design/Results: Here, we observed that irradiation of mouse lungs induced collagen deposition, particularly around blood vessels, in the early phase of RIPF. Such deposition subsequently became evident throughout the irradiated tissues. Accompanied by the collagen deposition, vascular EndMT (endothelial-to-mesenchymal transition) began to develop in the early phase of RIPF, before the appearance of EMT (epithelial-to-mesenchymal transition) of alveolar epithelial (AE) II cells in the substantive fibrotic phase. Concomitant with the EndMT, we detected vascular endothelial cell (EC)–specific hypoxic damage in the irradiated lung tissues. In human pulmonary artery endothelial cells (HPAEC), the radiation-induced EndMT via activation of TGFβ-R1/Smad signaling was dependent on HIF1α expression. A novel HIF1α inhibitor, 2-methoxyestradiol (2-ME), inhibited the irradiation-induced EndMT via downregulation of HIF1α-dependent Smad signaling. In vivo, 2-ME inhibited the vascular EndMT, and decreased the collagen deposition associated with RIPF. Furthermore, HIF1α-related EndMT was observed also in human RIPF tissues.Conclusions: We provide the first evidence that an EndMT occurs in RIPF development and that the EndMT may be effectively inhibited by modulating vascular EC-specific hypoxic damage. Clin Cancer Res; 21(16); 3716–26. ©2015 AACR.

A number of researchers in Korea have tried to set-up the production of radionuclides and develop new radiopharmaceuticals for several decades. Thanks to their 60-year endeavor to advance the field of radiopharmaceutical sciences, now we have a lot of research units and facilities in Korea. Still, there are huge number of issues to be solved in radiopharmaceutical sciences; however, our efforts will be continued to develop new radiopharmaceuticals and to apply the new radiopharmaceuticals into nuclear medicine field.

Background Immune checkpoint inhibitors such as anti-programmed cell death protein 1 (PD1) block tumor growth by reinvigorating the immune system; however, determining their efficacy only by the changes in tumor size may prove inaccurate. As the immune cells including macrophages in the tumor microenvironment (TME) are associated with the response to anti-PD1 therapy, tumor-associated macrophages (TAMs) imaging using nanoparticles can noninvasively provide the immune enrichment status of TME. Herein, the mannosylated-serum albumin (MSA) nanoparticle was labeled with radioactive isotope 68Ga to target the mannose receptors on macrophages for noninvasive monitoring of the TME according to anti-PD1 therapy. Results B16F10 tumor-bearing mice were treated with anti-PD1, and the response to anti-PD1 was determined by the tumor volume. According to the flow cytometry, the responders to anti-PD1 showed an increased proportion of TAMs, as well as lymphocytes, and the most enriched immune cell population in the TME was also TAMs. For noninvasive imaging of TAMs as a surrogate of immune cell augmentation in the TME via anti-PD1, we acquired [68Ga]Ga-MSA positron emission tomography. According to the imaging study, an increased number of TAMs in responders at the early phase of anti-PD1 treatment was observed in both B16F10 and MC38 tumor-bearing mice models. Conclusion As representative immune cells in the TME, non-invasive imaging of TAMs using MSA nanoparticles can reflect the immune cell enrichment status in the TME closely associated with the response to anti-PD1. As non-invasive imaging using MSA nanoparticles, this approach shows a potential to monitor and evaluate anti-tumor response to immune checkpoint inhibitors.

Exposure to ionizing radiation leads to severe damages in radiosensitive organs and induces acute radiation syndrome, including effects on the hematopoietic system and gastrointestinal system. In this study, the radioprotective ability of KMRC011, a novel toll-like receptor 5 (TLR5) agonist, was investigated in C57BL6/N mice exposed to lethal total-body gamma-irradiation. In a 30-day survival study, KMRC011-treated mice had a significantly improved survival rate compared with control after 11 Gy total-body irradiation (TBI), and it was found that the radioprotective activity of KMRC011 depended on its dosage and repeated treatment. In a 5-day short-term study, we demonstrated that KMRC011 treatment stimulated cell proliferation and had an anti-apoptotic effect. Furthermore, KMRC011 increased the expressions of genes related to DNA repair, such as Rad21, Gadd45b, Sod2 and Irg1, in the small intestine of lethally irradiated mice. Interestingly, downregulation of NF-κB p65 in the mouse intestine by KMRC011 treatment was observed. This data indicated that KMRC011 exerted a radioprotective activity partially by regulating NF-κB signaling. Finally, peak expression levels of G-CSF, IL-6, IFN-γ, TNF-α and IP-10 induced by KMRC011 treatment were different depending on the route of administration and type of cytokine. These cytokines could be used as candidate biomarkers for the evaluation of KMRC011 clinical efficacy. Our data indicated that KMRC011 has radioprotective activity in lethally irradiated mice and may be developed as a therapeutic agent for radioprotection.

We introduce an efficient cell tracking imaging protocol using positron emission tomography (PET). Since macrophages are known to home and accumulate in tumor tissues and atherosclerotic plaque, we design a PET imaging protocol for macrophage cell tracking using aza-dibenzocyclooctyne-tethered PEGylated mesoporous silica nanoparticles (DBCO-MSNs) with the short half-life F-18-labeled azide-radiotracer via an in vivo strain-promoted alkyne azide cycloaddition (SPAAC) covalent labeling reaction inside macrophage cells in vivo. This PET imaging protocol for in vivo cell tracking successfully visualizes the migration of macrophage cells into the tumor site by the bioorthogonal SPAAC reaction of DBCO-MSNs with [18F]fluoropentaethylene glycolic azide ([18F]2) to form 18F-labeled aza-dibenzocycloocta-triazolic MSNs (18F-DBCOT-MSNs) inside RAW 264.7 cells. The tissue radioactivity distribution results were consistent with PET imaging findings. In addition, PET images of atherosclerosis in ApoE−/- mice fed a western diet for 30 weeks were obtained using the devised macrophage cell-tracking protocol.

O-2-18F-fluoroethyl-l-tyrosine ([18F]FET) has been widely used for glioblastomas (GBM) in clinical practice, although evaluation of its applicability in non-clinical research is still lacking. The objective of this study was to examine the value of [18F]FET for treatment evaluation and prognosis prediction of anti-angiogenic drug in an orthotopic mouse model of GBM. Human U87MG cells were implanted into nude mice and then bevacizumab, a representative anti-angiogenic drug, was administered. We monitored the effect of anti-angiogenic agents using multiple imaging modalities, including bioluminescence imaging (BLI), magnetic resonance imaging (MRI), and positron emission tomography-computed tomography (PET/CT). Among these imaging methods analyzed, only [18F]FET uptake showed a statistically significant decrease in the treatment group compared to the control group (P=0.02 and P=0.03 at 5 and 20 mg/kg, respectively). This indicates that [18F]FET PET is a sensitive method to monitor the response of GBM bearing mice to anti-angiogenic drug. Moreover, [18F]FET uptake was confirmed to be a significant parameter for predicting the prognosis of anti-angiogenic drug (P=0.041 and P=0.007, on Days 7 and 12, respectively, on Pearson’s correlation; P=0.048 and P=0.030, on Days 7 and 12, respectively, on Cox regression analysis). However, results of BLI or MRI were not significantly associated with survival time. In conclusion, this study suggests that [18F]FET PET imaging is a pertinent imaging modality for sensitive monitoring and accurate prediction of treatment response to anti-angiogenic agents in an orthotopic model of GBM.

Introduction Macrophages play a key role in atherosclerotic plaque formation in atherosclerosis, but its detailed understanding has poorly investigated until now. Thus, we sought to demonstrate a noninvasive technique for macrophage tracking to atherosclerotic lesions in apolipoprotein E −/− (ApoE −/− ) mice with an imaging system based on sodium iodide symporter (NIS) gene coupled with 99m Tc-single-photon emission computed tomography (SPECT). Methods and results Macrophage cells (RAW264.7) were stably transduced with retrovirus expressing NIS gene (RAW-NIS). In RAW-NIS cells, uptake of 125 I was higher than the parental cells. [ 18 F]FDG signals in the aorta at 30weeks on an ApoE −/− mice with high cholesterol diet were higher (1.7±0.12% injected dose (ID)) than those in control group (0.84±0.06% ID). Through 99m Tc-SPECT/computed tomography (CT), in the RAW-NIS cell injected group, the 99m Tc-pertechnetate uptake in aorta was higher than control groups. However, according to atorvastatin treatment, RAW-NIS cell recruitment reduced to the aorta. Area of 99m Tc-pertechnetate uptake was positively correlated with immunostaining results against macrophage antigen (CD68). Cholesterol and low-density lipoprotein levels of atorvastatin-treated group showed lower than those of atorvastatin-untreated group, but did not reach statistical difference. Conclusions This novel approach to tracking macrophages to atherosclerotic plaques in vivo can be applied for studies of arterosclerotic vascular disease.

To obtain an additional pharmacological agent for the diagnosis of inflammation, we investigated the medical use of (89)Zr-oxalate as a positron emission tomography (PET) probe for the in vivo imaging of inflammation and compared its efficacy to that of 2-deoxy-2-[(18)F]fluoro-d-glucose ([(18)F]FDG) and sodium [(18)F]fluoride. (89)Zr-oxalate exhibited observable higher uptake in a macrophage cell line than in tumor cells. The inflammatory lesions and tumors were clearly visualized by PET imaging and autoradiography using (89)Zr-oxalate. Compared to [(18)F]FDG and sodium [(18)F]fluoride, (89)Zr-oxalate demonstrated a high selectivity index to the tumor at an early time point after injection and to inflammation at a delayed time point after injection (24 h). Through histological examination, large numbers of macrophages and neutrophils were observed in the tumor lesions with the highest (89)Zr-oxalate uptake. In a rheumatoid arthritis (RA) mouse model, (89)Zr-oxalate demonstrated a high level of accumulation in inflammatory lesions. (89)Zr-oxalate is a new strategic tool for tumor imaging and inflammatory processes.

The construction of biosensors with antibodies as the biological sensing element requires the integration of antibodies into the biosensor such that their activity is reproducibly retained. In this work, a peptide ligand was identified from phage-displayed heptameric peptide libraries for the purpose of antibody immobilization on biosensor surfaces. For biopanning of peptides specific to the Fc region of the antibody, rabbit anti-goat immunoglobulin G (IgG) antibody was immobilized on a culture dish coated with goat IgG, which enabled exposure of the Fc region of rabbit antibody on the surface with the Fab region bound to the surface-adsorbed goat IgG. The library phages were added to the dish and phages displaying antibody-binding peptides were competitively eluted using the rabbit antibody. After four rounds of biopanning, 22 phage plaques were randomly selected and their nucleotide sequences corresponding to the random peptide were determined. Three enriched sequences were selected and analyzed for their rabbit antibody-binding activity. One of these sequences (KHRFNKD), which was rich in positive charge and homologous to the IgG-binding domains of Staphylococcal protein A, exhibited a high and specific affinity to rabbit antibody. This peptide was immobilized on a quartz crystal microbalance (QCM) biosensor, and the resulting QCM was found to be capable of capturing antibody and detecting antigen, with potential applications in antibody purification and enzyme immunoassay.

This study was to evaluate the anti-atherosclerotic effect of Korean red ginseng (KRG) in apolipoprotein E knockout (ApoE KO) mice. Female ApoE KO mice were divided into 2 groups and were treated via intragastric inoculation for 21 weeks. The body weight increased gradually in both the KRG extract inoculation group and the control group inoculated with saline. Plaque lesion areas in the aortic sinus were reduced largely in the KRG inoculation group than in the control group. Triglyceride and inflammatory cytokines such as monocyte chemoattractant protein-1 (MCP-1) and tumor necrosis factor-alpha (TNF-α) were significantly reduced in the KRG inoculation group. Additionally, the KRG inoculation group showed significantly lower cardiovascular inflammation by 18F-fluoro-2-deoxy-d-glucose positron emission tomography/computed tomography compared to that in the control group. Taken together, KRG alleviates atherosclerotic lesions through MCP-1 and TNF-α suppression. These results suggest that KRG could be a valuable therapeutic strategy for the treatment of atherosclerosis.

Purpose: Radiation-induced pulmonary fibrosis (RIPF) is a late side effect of thoracic radiotherapy. The purpose of our study was to gain further insight into the development of RIPF. Experimental Design/Results: Here, we observed that irradiation of mouse lungs induced collagen deposition, particularly around blood vessels, in the early phase of RIPF. Such deposition subsequently became evident throughout the irradiated tissues. Accompanied by the collagen deposition, vascular EndMT (endothelial-to-mesenchymal transition) began to develop in the early phase of RIPF, before the appearance of EMT (epithelial-to-mesenchymal transition) of alveolar epithelial (AE) II cells in the substantive fibrotic phase. Concomitant with the EndMT, we detected vascular endothelial cell (EC)–specific hypoxic damage in the irradiated lung tissues. In human pulmonary artery endothelial cells (HPAEC), the radiation-induced EndMT via activation of TGFβ-R1/Smad signaling was dependent on HIF1α expression. A novel HIF1α inhibitor, 2-methoxyestradiol (2-ME), inhibited the irradiation-induced EndMT via downregulation of HIF1α-dependent Smad signaling. In vivo, 2-ME inhibited the vascular EndMT, and decreased the collagen deposition associated with RIPF. Furthermore, HIF1α-related EndMT was observed also in human RIPF tissues. Conclusions: We provide the first evidence that an EndMT occurs in RIPF development and that the EndMT may be effectively inhibited by modulating vascular EC-specific hypoxic damage. Clin Cancer Res; 21(16); 3716–26. ©2015 AACR.

Three-dimensional (3D) carbon nitride (C3 N4 )-based materials show excellent performance in a wide range of applications because of their suitable band structures. To realize the great promise of two-dimensional (2D) allotropes of various 3D materials, it is highly important to develop routes for the production of 2D C3 N4 materials, which are one-atom thick, in order to understand their intrinsic properties and identify their possible applications. In this work, water-dispersible, atomically thin, and small carbon nitride nanodots were produced using the chemical oxidation of graphitic C3 N4 . Various analyses, including X-ray diffraction, X-ray photoelectron, Fourier-transform infrared spectroscopy, and combustion-based elemental analysis, and thermogravimetric analysis, confirmed the production of 3D oxidized C3 N4 materials. The 2D C3 N4 nanodots were successfully exfoliated as individual single layers; their lateral dimension was several tens of nanometers. They showed strong photoluminescence in the visible region as well as excellent performances as cell-imaging probes in an in vitro study using confocal fluorescence microscopy.

Introduction In vivo tracking of the transplanted stem cells is important in pre-clinical research of stem cell therapy for myocardial infarction. We examined the feasibility of adenovirus-mediated sodium iodide symporter (NIS) gene to cell tracking imaging of transplanted stem cells in a canine infarcted myocardium by clinical single photon emission computed tomography (SPECT). Methods Beagle dogs were injected intramyocardially with NIS-expressing adenovirus-transfected canine stem cells (Ad-hNIS-canine ADSCs) a week after myocardial infarction (MI) development. 99m Tc-methoxyisobutylisonitrile ( 99m Tc-MIBI) and 99m Tc-pertechnetate ( 99m TcO 4 − ) SPECT imaging were performed for assessment of infarcted myocardium and viable stem cell tracking. Transthoracic echocardiography was performed to monitor any functional cardiac changes. Results Left ventricular ejection fraction (LVEF) was decreased after LAD ligation. There was no significant difference in EF between the groups with the stem cell or saline injection. 125 I uptake was higher in Ad-hNIS-canine ADSCs than in non-transfected ADSCs. Cell proliferation and differentiation were not affected by hNIS-carrying adenovirus transfection. 99m Tc-MIBI myocardial SPECT imaging showed decreased radiotracer uptake in the infarcted apex and mid-anterolateral regions. Ad-hNIS-canine ADSCs were identified as a region of focally increased 99m TcO 4 − uptake at the lateral wall and around the apex of the left ventricle, peaked at 2 days and was observed until day 9. Conclusions Combination of adenovirus-mediated NIS gene transfection and clinical nuclear imaging modalities enables to trace the fate of transplanted stem cells in infarcted myocardium for translational in vivo cell tracking study for prolonged duration.

The acute organ motion estimation in multimodal imaging improves quality of diagnosis and therapy of tumor. It is particularly important in the organ related involuntary movement such as lung and liver. The aim of this study was to compare the region specific motion in rodent using radioactive molecular sieve PET image and MRI image. The molecular sieve was contained 0.37 MBq F-18 and coated with hydrogel. For comparison of internal and external motion, molecular sieve was placed inside lung and liver and attached on the surface of each organ region in SD-rat. PET study was performed using a small animal PET scanner (Inveon) after IV injection of FDG 37 MBq/0.2 mL. List-mode data was synchronized with respiratory gating trigger signal from external monitoring system (Biovet). MRI study was performed using 3-T clinical MRI system (Magnetom Tim Trio) with human wrist coil. Coronal MRI images were acquired using T2-weighted turbo spin echo (TSE) sequence with respiratory triggering. The parameter is as follow settings: TR = 1000 ms, TE = 36 ms, FA = 20. We analyzed moving pattern and variation of movement in lung and liver region following respiratory cycle in both of PET and MRI images. Moving patterns in PET image were different in accordance with where molecular sieve was placed. The variation of lung and liver internal motion was 0.93 and 0.52 in PET image, respectively. Estimated organ motion in MRI image revealed moving pattern based on respiratory cycle. The maximum variation of lung and liver region was 1.75 and 1.36 in MRI image, respectively. We recognized organ motion was different depending on the region and the monitoring signal was overestimated compared to real motion in both of region in PET. This study demonstrated that region-specific motion estimation would realize through aid of MRI images without ext