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Integrin αvβ3 is an effective marker of angiogenesis in cancer, and αvβ3-specific imaging can yield important details about this complex physiological process. We utilized the recently reported and highly αvβ3-specific peptide, bicyclic RGD (bcRGD), as the basic structure of an in vivo αvβ3 imaging probe, and synthesized a radioiodinated form of bcRGD, namely [125I]bcRGD, with high radiochemical purity (>99%) and high molar activity (81 GBq/μmol). As expected, [125I]bcRGD exhibited high selectivity for αvβ3 compared with αvβ5 and α5β1 in vitro. [125I]bcRGD showed significantly higher accumulation in U-87MG cells (1.6% dose/mg) with high expression of αvβ3 compared to A549 cells (0.3% dose/mg) with only moderate expression. Furthermore, 30 min after administration to tumor-bearing mice, [125I]bcRGD showed significantly higher accumulation in U-87MG tumors (3.8% ID/g) than in A549 tumors (2.1% ID/g), and the radioactivity accumulation ratios of U-87MG tumor/blood and U-87MG tumor/muscle were 4.0 and 6.0, respectively. These results highlight the promising properties of [123/125I]bcRGD for use as an in vivo αvβ3 imaging probe, as well as the utility of bcRGD as a basic structure of molecular probes for both imaging and therapeutic applications. bcRGD may exhibit broad use in future theranostics applications targeting integrin αvβ3-related diseases.

Introduction Lactate could serve as an energy source and signaling molecule in the brain, although there is insufficient in vivo evidence to support this possibility. Here we aimed to use a one-pot enzymatic synthetic procedure to synthesize l -[3-11C]lactate that can be used to evaluate chemical forms in the blood after intravenous administration, and as a probe for pharmacokinetic analysis of lactate metabolism in in vivo positron emission tomography (PET) scans with normal and fasted rats. Methods Racemic [3-11C]alanine obtained from 11C-methylation of a precursor and deprotection was reacted with an enzyme mixture consisting of alanine racemase, d -amino acid oxidase, catalase, and lactate dehydrogenase to yield l -[3-11C]lactate via [3-11C]pyruvate. The optical purity was measured by HPLC. Radioactive chemical forms in the arterial blood of Sprague Dawley rats with or without insulin pretreatment were evaluated by HPLC 10 min after bolus intravenous injection of l -[3-11C]lactate. PET scans were performed on normal and fasted rats administered with l -[3-11C]lactate. Results l -[3-11C]Lactate was synthesized within 50 min and had decay corrected radiochemical yield, radiochemical purity, and optical purity of 13.4%, >95%, and >99%, respectively. The blood radioactivity peaked immediately after l -[3-11C]lactate injection, rapidly decreased to the minimum value within 90 s, and slowly cleared thereafter. HPLC analysis of blood samples revealed the presence of [11C]glucose (78.9%) and l -[3-11C]lactate (12.1%) 10 min after administration of l -[3-11C]lactate. Insulin pretreatment partly inhibited glyconeogenesis conversion leading to 55.4% as [11C]glucose and 38.9% as l -[3-11C]lactate simultaneously. PET analysis showed a higher SUV in the brain tissue of fasted rats relative to non-fasted rats. Conclusions We successfully synthesized l -[3-11C]lactate in a one-pot enzymatic synthetic procedure and showed rapid metabolic conversion of l -[3-11C]lactate to [11C]glucose in the blood. PET analysis of l -[3-11C]lactate indicated the possible presence of active lactate usage in rat brains in vivo.

Matrix metalloproteinase-12 (MMP-12, macrophage elastase) is a member of the MMP family that is responsible for the degradation of extracellular matrix, and is associated with the inflammatory process of chronic obstructive pulmonary disease (COPD). COPD, characterized by progressive and irreversible airflow obstruction, is recently a major cause of mortality and morbidity worldwide. Herein, to develop radioiodinated probes for the early diagnosis of COPD, we designed and synthesized novel MMP-12-targeted dibenzofuran compounds (1–3) with a variety of linker structures (carbamate, amide, and sulfonamide). In competitive enzyme activity assays, it was revealed that the linker structures significantly affected the inhibitory activity against and selectivity for MMP-12. Compound 1, with carbamate linker, demonstrated potent MMP-12 inhibitory activity (IC50 = 8.5 nM) compared to compound 2, with amide linker, and compound 3, with sulfonamide linker. Using bromo-substituted carbamate 13 as a radioiodination precursor, [125I]1 was successfully prepared to high radiochemical purity (over 98%) and good specific radioactivity (4.1 GBq/μmol). These results suggest that radioiodinated compound 1 is potent as a novel MMP-12-targeted probe.

A high-affinity and low-molecular-weight fluorescent Zn2+ sensor 1 based on a 2,2′-bipyridine scaffold, which functions as both the chelating moiety for Zn2+ and the fluorophore, was developed and evaluated in biological applications. Controlling the occurrence of tautomerism of the chelating moiety for Zn2+ by introducing an amino group at the 6-position of the pyridine ring dramatically increased the binding affinity toward Zn2+. Fluorescent sensor 1 exhibited a nanomolar-range dissociation constant (Kd = 2.2 nM), a large Stokes shift (140 nm), and an 8.6-fold turn-on response to Zn2+ under physiological conditions. Fluorescence images revealed that fluorescent sensor 1 exhibits good properties with respect to aqueous solubility and cell permeability and can quantitatively detect the Zn2+ levels in living cells. Furthermore, a 65Zn2+ radioactive zinc isotope uptake study revealed the real concentration of accumulated Zn2+ at the detection limit. The novel fluorescent sensor 1 is a promising sensor for use in biological applications.

Near-infrared (NIR: 800-1000nm) fluorescent probes, which activate their fluorescence following interaction with functional biomolecules, are desirable for noninvasive and sensitive tumor diagnosis due to minimal tissue interference. Focusing on bioavailability and applicability, we developed a probe with a self-assembling polymer micelle, a lactosome, encapsulating various quantities of NIR dye (IC7-1). We also conjugated anti-HER2 single chain antibodies to the lactosome surface and examined the probe's capacity to detect HER2 in cells and in vivo . Micelles encapsulating 20mol% IC7-1 (hIC7L) showed 30-fold higher fluorescence (λ em : 858nm) after micelle denaturation compared to aqueous buffer. Furthermore, antibody modification allowed specific activation of the probe (HER2-hIC7L) following internalization by HER2-positive cells, with the probe concentrating in lysosomes. HER2-hIC7L intravenously administered to mice clearly and specifically visualized HER2-positive tumors by in vivo optical imaging. These results indicate that HER2-hIC7L is a potential activatable NIR probe for sensitive tumor diagnosis. From the Clinical Editor Near-infrared probes that activate their fluorescence following interaction with specific biomolecules are desirable for noninvasive and sensitive tumor detection due to minimal tissue interference. This team of authors developed a probe termed hIC7L and demonstrate its potential in HER2 tumor diagnosis.

We report a novel series of fluorescent Zn 2+ sensors based on a pyridine–pyridone core structure, which functions as both the chelating part for Zn 2+ and as the fluorophore. Pyridine–pyridone derivatives obtained by replacement of an electron-donating group with an electronic-withdrawing group at the 3-position of the pyridone ring showed almost no background fluorescence because electron transfer to the fluorophore was inhibited. However, these derivatives showed strong fluorescence with Zn 2+ . Coordination with Zn 2+ greatly influenced the electron transfer of the fluorophore, and, as a result, enabled fluorescence ON/OFF switching. In these sensors, 4-[4-(methylsulfanyl)-2-oxo-6-(pyridin-2-yl)-1,2-dihydropyridin-3-yl]benzoic acid ( 5 ) displayed excellent properties: small molecular weight (MW = 338), good water solubility, large Stokes shift (132 nm), and fluorescence ON/OFF switching with Zn 2+ . In addition, fluorescence images of Zn 2+ with 5 in living cells show that the new fluorescent sensor is useful for biological applications.

We examined exendin(9-39), an antagonist of glucagon-like peptide-1 (GLP-1) receptor (GLP-1R), as a potential probe for imaging of pancreatic beta-cells. To evaluate in vitro receptor specificity, binding assay was performed using dispersed mouse islet cells. Binding assay showed competitive inhibition of [(125)I]BH-exendin(9-39) binding by non-radioactive exendin(9-39). To assess in vivo selectivity, the biodistribution was evaluated by intravenous administration of [(125)I]BH-exendin(9-39) to mice. Radioactivity of harvested pancreas reached highest levels at 60 and 120min among organs examined except lung. Pre-administration of excess non-radioactive exendin(9-39) remarkably and specifically blocked the radioactivity of pancreas. After [(125)I]BH-exendin(9-39) injection into transgenic mice with pancreatic beta-cells expressing GFP, fluorescent and radioactive signals of sections of pancreas were evaluated with an image analyzer. Imaging analysis showed that the fluorescent GFP signals and the radioactive signals were correspondingly located. Thus, the GLP-1R antagonist exendin(9-39) may serve as a useful probe for pancreatic beta-cell imaging.

Introduction Despite extensive attempts to develop cyclooxygenase (COX)-2 imaging radiotracers, no suitable positron emission tomography (PET)/single photon emission computed tomography (SPECT) tracers are currently available for in vivo imaging of COX-2 expression. The aims of this study were to synthesize and evaluate a radioiodinated derivative of lumiracoxib, 2-[(2-fluoro-6-iodophenyl)-amino]-5-methylphenylacetic acid (FIMA), which is structurally distinct from other drugs in the class and has weakly acidic properties, as a SPECT tracer for imaging COX-2 expression. Methods The COX inhibitory potency was assessed by measuring COX-catalyzed oxidation with hydrogen peroxide. Cell uptake characteristics of 125 I-FIMA were assessed in control and linterfero/interferon-γ-stimulated macrophages. The biodistribution of 125 I-FIMA was determined by the ex vivo tissue counting method in rats. Results The COX-2 inhibitory potency of FIMA (IC 50 =2.46 μM) was higher than that of indomethacin (IC 50 =20.9 μM) and was comparable to lumiracoxib (IC 50 =0.77 μM) and diclofenac (IC 50 =0.98 μM). The IC 50 ratio (COX-1/COX-2=182) indicated FIMA has a high isoform selectivity for COX-2. 125 I-FIMA showed a significantly higher accumulation in COX-2 induced macrophages than in control macrophages, which decreased with nonradioactive FIMA in a concentration dependent manner. The biodistribution study showed rapid clearance of 125 I-FIMA from the blood and most organs including the liver and kidneys. No significant in vivo deiodination was observed with radioiodinated FIMA. Conclusions FIMA showed high inhibitory potency and selectivity for COX-2. Radioiodinated FIMA showed specific accumulation into COX-2 induced macrophages, no significant in vivo deiodination and rapid blood clearance. Radioiodinated FIMA deserves further investigation as a SPECT radiopharmaceutical for imaging COX-2 expression.

Since a very complicated technique is necessary to measure cerebral blood flow (CBF) with [14C]iodoantipyrine in small animals, a practical and easy method is needed. In this paper, the differential uptake ratio (DUR) at 5 min after injection of N-isopropyl-p-[125I]iodoamphetamine (IMP) was estimated as an index of CBF in normal rats and compared with the quantitative CBF value measured using [15O]H2O and dynamic PET scan. A good correlation between the two values was obtained. The results indicate that DUR of [125I]IMP at 5 min after injection in rats is a useful and practical indicator of CBF since neither arterial blood sampling nor metabolite correction is necessary.

In the basic research on cerebral perfusion disorders, a new method for measurement of oxygen extraction fraction (OEF) and cerebral metabolic rate for oxygen (CMRO2) has been desired. Although the O-15-labeled gas inhalation method is performed in clinical studies, application of the inhalation method to small animals requires too many intensive procedures. Therefore, this study investigated a new method of assessing OEF in small animals using intravenously injectable oxygen (injectable 15O-O2). Additionally, it was compared with the inhalation method, using a monkey for further validation. Using injectable 15O-O2, 72 MBq/ml of radioactivity was obtained after 15O-O2 gas circulation into an artificial lung. OEF with injectable 15O-O2 was calculated using the same equation as that applied to the bolus inhalation method. The OEF value obtained by injectable 15O-O2 was well in accordance with that evaluated by the arterial-venous difference of oxygen concentration. Furthermore, the OEF and CMRO2 images obtained with two methods using a monkey brain showed a similar distribution. These results indicate that this method is useful for the estimation of OEF and CMRO2 in small animals using an animal positron emission tomography system and may accelerate the basic research of cerebral perfusion diseases.

In the basic research about cerebral perfusion disorders, a new method for measurement of oxygen extraction fraction (OEF) and cerebral metabolic rate for oxygen (CMRO 2 ) has been desired. Although O-15-labeled gas inhalation method is performed in clinical studies, application of the inhalation method to small animals requires too many intensive procedures. Therefore, this study investigated a new method of assessing OEF in small animals using intravenously injectable oxygen (injectable 15 O-O 2 ). Moreover, OEF after transient ischemia–reperfusion was estimated to validate the usefulness of the method. Injectable 15 O-O 2 , 72 MBq/ml of radioactivity, was obtained after 15 O-O 2 gas circulation into the artificial lung. OEF after injection of injectable 15 O-O 2 was calculated using the same equation as that applied to the bolus inhalation method. OEF value obtained by injectable 15 O-O 2 was well in accordance with that evaluated by arterial-venous difference of oxygen concentration. Furthermore, misery perfusion was depicted in the lesioned hemisphere after transient ischemia–reperfusion. These results indicated that this method is useful for the estimation of OEF and CMRO 2 in small animals using an animal positron emission tomography (PET) system and may accelerate the basic research of cerebral perfusion diseases.