Development of Low Molecular Weight Ligands for Integrin αvß3.

We designed and synthesized non-peptide organic molecular ligands for integrin αvß3. Candidate ligands featured amidino analog and carboxy groups as binding sites on either side of a spacer, which consisted of benzophenone or an analog, such as diphenyl sulfide, diphenyl sulfoxide, diphenyl sulfone, or diphenyl ether. Competitive binding assays to integrin αvß3 with respect to [125I]echistatin were used to determine inhibitory activity of the synthetic ligands. Ligands bearing 2-aminobenzimidazoyl and glycyl groups separated by a benzophenone spacer demonstrated more potent binding than did a linear Arg-Gly-Asp (RGD) tripeptide that represents the native integrin αvß3 binding motif. Ligands possessing 2-aminobenzimidazoyl and carboxy groups and diphenyl sulfoxide or diphenyl ether spacers inhibited binding of [125I]echistatin with IC50 values similar to that of the linear RGD tripeptide.

Renal dysfunction can occur in advanced-stage Duchenne muscular dystrophy.

INTRODUCTION: With improved treatments, patients with Duchenne muscular dystrophy (DMD) can survive far beyond adolescence. However, advanced-stage DMD patients are at risk of developing renal dysfunction. In this study, long-term renal function outcomes and associated risk factors in advanced stage DMD were analyzed. METHODS: Fifty-one patients were classified into three different age groups (<20, 20-29, and ≥30 years of age), and cystatin C (CysC) levels were compared among groups. RESULTS: Median serum CysC levels were 0.74 mg/L, 0.63 mg/L, and 0.76 mg/L in the age groups of <20, 20-29, and ≥30 years, respectively (P = .003). Five of the nine patients in the ≥30 years age group showed elevated serum CysC and decreased cardiac function compared with the other four in the group (P = .014). DISCUSSION: Our results indicate an association between cardiac and renal dysfunction in patients with advanced-stage DMD.

PET probe detecting non-small cell lung cancer susceptible to epidermal growth factor receptor tyrosine kinase inhibitor therapy.

Tyrosine kinase inhibitors for epidermal growth factor receptor (EGFR-TKIs) are used as molecular targeted therapy for non-small cell lung cancer (NSCLC) patients. The therapy is applied to the patients having EGFR-primary L858R mutation, but drug tolerance caused by EGFR-secondary mutation is occurred within one and half years. For the non-invasive detection of the EGFR-TKIs treatment positive patients by positron emission tomograpy (PET) imagaing, fluorine-18 labeled thienopyrimidine derivative, [18F]FTP2 was newly synthesized. EGFR inhibition assay, cell uptake study, and blocking study indicated [18F]FTP2 binds with high and selective affinity for EGFR with L858R mutation, and not with L858R/T790M dual mutations. On animal PET study using tumor bearing mice, H3255 cells expressing L858R mutated EGFR was more clearly visualized than H1975 cells expressing L858R/T790M dual mutated EGFR. [18F]FTP2 has potential for detecting NSCLC which is susceptible to EGFR-TKI treatment.

Comparison of Radioiodine- or Radiobromine-Labeled RGD Peptides between Direct and Indirect Labeling Methods.

Radiolabeled cyclic peptides containing the (Arg-Gly-Asp) RGD sequence for use in positron emission tomography (PET) imaging, single-photon emission computed tomography (SPECT) imaging, and targeted radionuclide therapy of cancer have been reported. In this study, RGD was used as a model carrier peptide for diagnosis and therapy of cancer. To evaluate the characteristics of radiohalogen-labeled peptides, several kinds of labeled RGD peptides [125I-c(RGDyK), 77Br-c(RGDyK), [125I]SIB-c(RGDfK), [77Br]SBrB-c(RGDfK), [125I]SIB-EG2-c(RGDfK), and [77Br]SBrB-EG2-c(RGDfK)] were designed, prepared, and evaluated. In these initial studies, 77Br (t1/2=57.0 h) and 125I (t1/2=59.4 d) were used because of their longer half-lives. Precursor peptides were synthesized using a standard 9-fluorenylmethyloxycarbonyl (Fmoc)-based solid-phase methodology. Radiolabeled peptides were prepared by chloramine-T method or conjugation of RGD peptides with [125I]N-succinimidyl 3-iodobenzoate ([125I]SIB) or [77Br]N-succinimidyl 3-bromobenzoate ([77Br]SBrB). Measurement of the partition coefficients, integrin binding assay, and biodistribution experiments in tumor-bearing mice were performed. 125I and 77Br labeling were successfully performed using similar methods, and in vitro characteristics and biodistributions were similar between the 125I-labeled and corresponding 77Br-labeled peptides. [125I]SIB- and [77Br]SBrB-conjugated RGD peptides showed higher partition coefficients, lower tumor uptakes, and higher intestinal uptake than 125I-c(RGDyK) and 77Br-c(RGDyK). [125I]SIB-EG2-c(RGDfK) and [77Br]SBrB-EG2-c(RGDfK), which possess an ethylene glycol linker, decreased lipophilicity and uptake in intestine compared with [125I]SIB-c(RGDfK) and [77Br]SBrB-c(RGDfK), which possess no linker. However, the improvement in biodistribution of [125I]SIB-EG2-c(RGDfK) and [77Br]SBrB-EG2-c(RGDfK)] was insufficient. In conclusion, directly radiohalogenated c(RGDyK) peptides are potentially more useful for tumor imaging and therapy than indirectly radiohalogenated ones.

Indocyanine Green-Labeled Polysarcosine for in Vivo Photoacoustic Tumor Imaging.

Photoacoustic (PA) imaging has been considered an attractive imaging modality for sensitive and in-depth imaging of biomolecules with a high resolution in vivo. PA imaging probes utilizing fluorescence dyes, including indocyanine green (ICG), have been proposed to enhance PA signal intensity. On the other hand, nanomicelles modified with polysarcosine (PSar), a biocompatible hydrophilic polymer, on their surface have previously achieved rapid tumor uptake, suggesting active transport of PSar into tumor tissues. Thus, we hypothesized that PSar-based materials might be utilized as diagnostic probes for targeting tumors and therefore evaluated the potential of PSar labeled with an ICG derivative, ICG-PSar, as a PA imaging probe for targeting cancer. In this study, ICG-PSars with differing molecular weights (10, 20, and 30 kDa) were synthesized. In vitro cellular uptake studies using ICG-PSar demonstrated rapid uptake in colon26 tumor cells partially via macropinocytosis-mediated endocytosis. In vivo fluorescence imaging and biodistribution study indicated that ICG-PSar30k exhibited high accumulation in the tumor (8.4% dose/g), with high tumor-to-blood ratios reaching 4.6 at 24 h post injection of the probe. Finally, in vivo PA imaging studies showed that PA signal increased in tumors (251%) but not in blood vessels, achieving high contrast tumor imaging at 24 h after ICG-PSar30k probe injection. These results suggest that ICG-PSar has potential as a tumor-targeting PA imaging probe.

Development of anti-HER2 fragment antibody conjugated to iron oxide nanoparticles for in vivo HER2-targeted photoacoustic tumor imaging.

Photoacoustic (PA) imaging is a promising imaging modality that provides biomedical information with high sensitivity and resolution. Iron oxide nanoparticles (IONPs) have been regarded as remarkable PA contrast agents because of their low toxicity and biodegradable properties. However, IONP delivery is restricted by its modest leakage and retention in tumors. In this study, we designed IONPs (20nm, 50nm, and 100nm) conjugated with anti-HER2 moieties [whole IgG, single-chain fragment variable (scFv), and peptide] for HER2-targeted PA tumor imaging. The binding affinity, cellular uptake, and in vivo biodistribution were examined. We propose 20-nm anti-HER2 scFv-conjugated IONPs (SNP20) as a novel PA contrast agent. SNP20 demonstrated high affinity and specific binding to HER2-expressing cells; it selectively visualized HER2-positive tumors in PA imaging studies. These data indicate that SNP20 is a potential PA contrast agent for imaging of HER2-expressing tumors. FROM THE CLINICAL EDITOR: Iron oxide nanoparticles have been demonstrated to be good contrast agents for tumor imaging. They may also be useful in photoacoustic (PA) imaging, which can provide high sensitivity data and image resolution. The authors here coupled iron oxide nanoparticles with anti-HER2 antibody fragment and showed significant retention of these nanoparticles in tumors. This combination may provide another option for enhanced imaging of tumors.

Evasion from accelerated blood clearance of nanocarrier named as "Lactosome" induced by excessive administration of Lactosome.

BACKGROUND: Nanoparticle of Lactosome, which is composed of poly(l-lactic acid)-base depsipeptide with diameter of 35nm, accumulates in solid tumors by the enhanced permeability and retention (EPR) effect. However, a pharmacokinetic alteration of Lactosome was observed when Lactosome was repeatedly administered. This phenomenon is named as the Lactosome accelerated blood clearance (ABC) phenomenon. In this study, the effect of Lactosome dose on the ABC phenomenon was examined and discussed in terms of immune tolerance. METHODS: To tumor transplanted mice, Lactosome (0-350mg/kg) was administrated. At 7days after the first administration, indocyanine green (ICG)-labeled Lactosome (ICG-Lactosome, 0-350mg/kg) was injected. Near-infrared fluorescence imaging was performed, and biodistribution of ICG-Lactosome was evaluated. Further, the produced amounts of anti-Lactosome IgM were determined by enzyme-linked immunosorbent assay (ELISA). RESULTS: ICG-Lactosome accumulated in the tumor region when the first Lactosome dose exceeded over 150mg/kg. The amounts of anti-Lactosome IgM were inversely correlated with the first Lactosome doses. Even after establishment of the Lactosome ABC phenomenon with the first Lactosome dose as low as 5.0mg/kg, the Lactosome ABC phenomenon can be evaded apparently by dosing ICG-Lactosome over 50mg/kg regardless of anti-Lactosome IgM production. CONCLUSIONS: There are two different mechanisms for evasion from the Lactosome ABC phenomenon before and after its establishment. In either mechanism, however, the Lactosome ABC phenomenon can be evaded by excessive administration of Lactosome. GENERAL SIGNIFICANCE: Lactosome is a potential nanocarrier for drug and/or imaging agent delivery, which can be used for frequent administrations without significant pharmacokinetic alterations.

In vivo imaging of membrane type-1 matrix metalloproteinase with a novel activatable near-infrared fluorescence probe.

Membrane type-1 matrix metalloproteinase (MT1-MMP) is a protease activating MMP-2 that mediates cleavage of extracellular matrix components and plays pivotal roles in tumor migration, invasion and metastasis. Because in vivo noninvasive imaging of MT1-MMP would be useful for tumor diagnosis, we developed a novel near-infrared (NIR) fluorescence probe that can be activated following interaction with MT1-MMP in vivo. MT1-hIC7L is an activatable fluorescence probe comprised of anti-MT1-MMP monoclonal antibodies conjugated to self-assembling polymer micelles that encapsulate NIR dyes (IC7-1, λem : 858 nm) at concentrations sufficient to cause fluorescence self-quenching. In aqueous buffer, MT1-hIC7L fluorescence was suppressed to background levels and increased approximately 35.5-fold in the presence of detergent. Cellular uptake experiments revealed that in MT1-MMP positive C6 glioma cells, MT1-hIC7L showed significantly higher fluorescence that increased with time as compared to hIC7L, a negative control probe lacking the anti-MT1-MMP monoclonal antibody. In MT1-MMP negative MCF-7 breast adenocarcinoma cells, both MT1-hIC7L and hIC7L showed no obvious fluorescence. In addition, the fluorescence intensity of C6 cells treated with MT1-hIC7L was suppressed by pre-treatment with an MT1-MMP endocytosis inhibitor (P < 0.05). In vivo optical imaging using probes intravenously administered to tumor-bearing mice showed that MT1-hIC7L specifically visualized C6 tumors (tumor-to-background ratios: 3.8 ± 0.3 [MT1-hIC7L] vs 3.1 ± 0.2 [hIC7L] 48 h after administration, P < 0.05), while the probes showed similarly low fluorescence in MCF-7 tumors. Together, these results show that MT1-hIC7L would be a potential activatable NIR probe for specifically detecting MT1-MMP-expressing tumors.

The potential of o-bromo-trans-decalinvesamicol as a new PET ligand for vesicular acetylcholine transporter imaging.

We investigated the characteristics of the regional rat brain distribution of radio-brominated o-bromo-decalinvesamicol (OBDV) in vivo to evaluate its potential as a PET ligand for vesicular acetylcholine transporter (VAChT). In in vivo biodistribution study, the specific brain regional accumulation of [(77) Br]OBDV was revealed 30 min after intravenous injection. The specific brain regional accumulation of [(77) Br]OBDV was significantly inhibited by co-injection of (+/-)-vesamicol. In contrast, no significant inhibition of the uptake of [(77) Br]OBDV in all brain regions was observed with co-injection of (+)-pentazocine (selective σ-1 receptor agonist) and (+)-3-(3-hydroxyphenyl)-N-propylpiperidine, [(+)-3-PPP] (σ-1 and σ-2 receptor agonist) with [(77) Br]OBDV. [(77) Br]OBDV accumulation in VAChT-rich brain regions was observed in ex vivo autoradiography. These results showed that [(77) Br]OBDV selectively bound to VAChT with high affinity in rat brain in vivo. Hence, OVBDV radiolabelled with more suitable (76) Br was suggested to be a potent VAChT ligand for PET.

Morphology control between twisted ribbon, helical ribbon, and nanotube self-assemblies with his-containing helical peptides in response to pH change.

pH-Responsive molecular assemblies with a variation in morphology ranging from a twisted ribbon, a helical ribbon, to a nanotube were prepared from a novel A3B-type amphiphilic peptide having three hydrophilic poly(sarcosine) (A block) chains, a hydrophobic helical dodecapeptide (B block), and two histidine (His) residues between the A3 and B blocks. The A3B-type peptide adopted morphologies of the twisted ribbon at pH 3.0, the helical ribbon at pH 5.0, and the nanotube at pH 7.4, depending upon the protonation states of the two His residues. On the other hand, another A3B-type peptide having one His residue between the A3 and B blocks showed a morphology change only between the helical ribbon and the relatively planar sheets with pH variation in this range. The morphology change is thus induced by one- or two-charge generation at the linking site of the hydrophilic and hydrophobic blocks of the component amphiphiles but in different ways.

Size control of core-shell-type polymeric micelle with a nanometer precision.

Amphiphilic polydepsipeptides having a hydrophobic poly(L-lactic acid) block and varying numbers of a hydrophilic poly(sarcosine) block ranging from 1 to 3, AB-, A2B-, and A3B-type, were prepared and studied on their molecular assemblies. The morphologies were found to be polymeric micelles for the AB- and the A3B-type polydepsipeptides, but worm-like micelles for the A2B-type polydepsipeptide. The hydrodynamic diameter of the A3B-type polydepsipeptide (22 nm) became smaller than the AB-type polydepsipeptide (34 nm). The polymeric micelle sizes composed of the AB-type polydepsipeptide were adjustable up to ca. 100 nm with incorporation of poly(L-lactic acid) into the hydrophobic core. On the other hand, with varying mixing ratio of the AB-type and A3B-type polydepsipeptides, the hydrodynamic diameters were tunable to become smaller sizes with a precise control in the range from 22 to 34 nm. The polydispersity indices of the polymeric micelles were less than 0.1, indicating that we can obtain the homogeneous polymeric micelles with diameters in the range from 20 to 100 nm under a precise control.

Suppressive immune response of poly-(sarcosine) chains in peptide-nanosheets in contrast to polymeric micelles.

Nanoparticles are expected to be applicable for the theranostics as a carrier of the diagnostic and therapeutic agents. Lactosome is a polymeric micelle composed of amphiphilic polydepsipeptide, poly(sarcosine)64-block-poly(L-lactic acid)30, which was found to accumulate in solid tumors through the enhanced permeability and retention effect. However, lactosome was captured by liver on the second administration to a mouse. This phenomenon is called as the accelerated blood clearance phenomenon. On the other hand, peptide-nanosheet composed of amphiphilic polypeptide, poly(sarcosine)60-block-(L-Leu-Aib)6, where the poly(L-lactic acid) block in lactosome was replaced with the (L-Leu-Aib)6 block, abolished the accelerated blood clearance phenomenon. The ELISA and in vivo near-infrared fluorescence imaging revealed that peptide-nanosheets did not activate the immune system despite the same hydrophilic block being used. The high surface density of poly(sarcosine) chains on the peptide-nanosheet may be one of the causes of the suppressive immune response.

Micelle-based activatable probe for in vivo near-infrared optical imaging of cancer biomolecules.

Near-infrared (NIR: 800-1000 nm) 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 (λem: 858 nm) 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.

Solid tumor-targeting theranostic polymer nanoparticle in nuclear medicinal fields.

Polymer nanoparticles can be prepared by self-assembling of amphiphilic polymers, and various types of molecular assemblies have been reported. In particular, in medicinal fields, utilization of these polymer nanoparticles as carriers for drug delivery system (DDS) has been actively tried, and some nanoparticulate drugs are currently under preclinical evaluations. A radionuclide is an unstable nucleus and decays with emission of radioactive rays, which can be utilized as a tracer in the diagnostic imaging systems of PET and SPECT and also in therapeutic purposes. Since polymer nanoparticles can encapsulate most of diagnostic and therapeutic agents with a proper design of amphiphilic polymers, they should be effective DDS carriers of radionuclides in the nuclear medicinal field. Indeed, nanoparticles have been recently attracting much attention as common platform carriers for diagnostic and therapeutic drugs and contribute to the development of nanotheranostics. In this paper, recent developments of solid tumor-targeting polymer nanoparticles in nuclear medicinal fields are reviewed.

High efficacy of particle beam therapies against tumors under hypoxia and prediction of the early stage treatment effect using 3'-deoxy-3'-[18F]fluorothymidine positron emission tomography.

OBJECTIVE: Compared with radiation therapy using photon beams, particle therapies, especially those using carbons, show a high relative biological effectiveness and low oxygen enhancement ratio. Using cells cultured under normoxic conditions, our group reported a greater suppressive effect on cell growth by carbon beams than X-rays, and the subsequent therapeutic effect can be predicted by the cell uptake amount of 3'-deoxy-3'-[18F]fluorothymidine (18F-FLT) the day after treatment. On the other hand, a hypoxic environment forms locally around solid tumors, influencing the therapeutic effect of radiotherapy. In this study, the influence of tumor hypoxia on particle therapies and the ability to predict the therapeutic effect using 18F-FLT were evaluated. METHODS: Using a murine colon carcinoma cell line (colon 26) cultured under hypoxic conditions (1.0% O2 and 5.0% CO2), the suppressive effect on cell growth by X-ray, proton, and carbon irradiation was evaluated. In addition, the correlation between decreased 18F-FLT uptake after irradiation and subsequent suppression of cell proliferation was investigated. RESULTS: Tumor cell growth was suppressed most efficiently by carbon-beam irradiation. 18F-FLT uptake temporarily increased the day after irradiation, especially in the low-dose irradiation groups, but then decreased from 50 h after irradiation, which is well correlated with the subsequent suppression on tumor cell growth. CONCLUSIONS: Carbon beam treatment shows a strong therapeutic effect against cells under hypoxia. Unlike normoxic tumors, it is desirable to perform 18F-FLT positron emission tomography 2-3 days after irradiation for early prediction of the treatment effect.

PET/MRI multimodality imaging to evaluate changes in glymphatic system function and biomarkers of Alzheimer's disease.

The glymphatic system is considered to play a pivotal role in the clearance of disease-causing proteins in neurodegenerative diseases. This study employed MR diffusion tensor imaging (DTI) to evaluate glymphatic system function and its correlation with brain amyloid accumulation levels measured using [11C]Pittsburgh compound-B (PiB) PET/MRI. Fifty-six patients with mild cognitive impairment and early Alzheimer's disease (AD: 70 ± 11 y) underwent [11C]PiB PET/MRI to assess amyloid deposition and were compared with 27 age-matched cognitively normal volunteers (CN: 69 ± 10y). All participants were evaluated for cognitive function using the Mini Mental State Examination (MMSE) before [11C]PiB PET/MRI. DTI images were acquired during the PET/MRI scan with several other MR sequences. The DTI analysis along the perivascular space index (DTI-ALPS index) was calculated to estimate the functional activity of the glymphatic system. Centiloid scale was applied to quantify amyloid deposition levels from [11C]PiB PET images. All patients in the AD group showed positive [11C]PiB accumulation, whereas all CN participants were negative. ALPS-index for all subjects linearly correlated with PiB centiloid, MMSE scores, and hippocampal volume. The correlation between the ALPS-index and PiB accumulation was more pronounced than with any other biomarkers. These findings suggest that glymphatic system dysfunction is a significant factor in the early stages of Alzheimer's disease.

Droplets Adhesion to Surgical Masks during Standard Oral Surgery.

The most common routes of transmission of coronavirus disease 2019 are droplet and contact infections. During dental treatment, several instruments and procedures used generate droplets of saliva and blood, such as during the extraction of an impacted third molar (M3). Surgical masks are often used during tooth extraction. However, the surface structures of surgical masks against droplets are not fully understood. Therefore, we analyzed the droplets that adhered to the surgical masks during impacted M3 extraction using electron microscopy. The surgical mask was divided into three layers and observed using electron microscopy. The outer and inner layers had a similar mesh-like structure, whereas the middle layer had a denser three-dimensional structure. Droplets ranging from 20-100 µm in size, generated during the extraction, adhered to the fibers of the outer layer of the mask. Fewer droplets adhered to the middle layer than to the outer layer. Droplets did not reach the inner layer. In conclusion, we suggest that a surgical mask can prevent droplet infection when performing impacted M3 extraction. This study is expected to contribute to the study of infection control strategies during dental treatments in the future.

[18F]FES PET Resolves the Diagnostic Dilemma of COVID-19-Vaccine-Associated Hypermetabolic Lymphadenopathy in ER-Positive Breast Cancer.

Coronavirus disease (COVID-19) vaccination is known to cause a diagnostic dilemma due to false-positive findings on [18F]FDG PET in vaccine-associated hypermetabolic lymphadenopathy. We present two case reports of women with estrogen-receptor (ER)-positive cancer of the breast who were vaccinated for COVID-19 in the deltoid muscle. [18F]FDG positron emission tomography (PET) demonstrated primary breast cancer and multiple axillary lymph nodes with increased [18F]FDG uptake, diagnosed as vaccine-associated [18F]FDG-avid lymph nodes. Subsequent [18F]FES PET revealed single axillary lymph node metastasis in the vaccine-associated [18F]FDG-avid lymph nodes. To the best of our knowledge, this is the first study showing the usefulness of [18F]FES PET in diagnosing axillary lymph node metastasis in COVID-19-vaccinated patients harboring ER-positive breast cancer. Thus, [18F]FES PET has potential applications in the detection of true-positive metastatic lymph nodes in patients with ER-positive breast cancer regardless of the ipsilateral or contralateral side, who have received COVID-19 vaccination.

Self-assemblies of triskelion A2B-type amphiphilic polypeptide showing pH-responsive morphology transformation.

A pH-responsive rolled-sheet morphology was prepared from a triskelion A(2)B-type amphiphilic polypeptide having a histidine residue as a pH-responsive unit. The dimensions of the rolled sheet were 85 nm diameter and 210 nm length with a sheet turn number of 2.0 at pH 7.4. Upon decreasing the pH from 7.4 to 5.0, the layer spacing of the rolled sheets was widened from ca. 9 to ca. 19 nm due to electrostatic repulsion caused by histidine protonation. This morphology change occurred reversibly with a pH change between 7.4 and 5.0. The molecular packing in the rolled sheets was shown to be loosened at pH 5.0 on the basis of electron diffraction measurements. The tightness of the rolled sheets was thus controlled reversibly by a pH change due to a single protonation in the amphiphilic polypeptide.

Radiobrominated probe targeting activated p38α in inflammatory diseases.

OBJECTIVE: p38α, a member of the mitogen-activated protein kinase superfamily, is activated by external stimuli, followed by nuclear translocation for the regulation of inflammatory responses at the transcriptional and translational levels in inflammatory diseases. Thus, activated p38α would be an appropriate target molecule for in vivo noninvasive imaging and targeted radionuclide therapy. For this purpose, we designed a radiobrominated compound, 6-(4-[77Br]bromo-2-fluorophenoxy)-8-methyl-2-(tetrahydro-2H-pyran-4-ylamino)-pyrido[2,3-d]pyrimidin-7(8H)-one ([77Br]4-BR), based on a potent p38α selective inhibitor, R1487, for use with single-photon emission computed tomography. We synthesized [77Br]4-BR and evaluated its effectiveness as an activated p38α imaging probe compared with our previous radioiodinated probe (6-(2-fluoro-4-[125I]iodophenoxy)-8-methyl-2-(tetrahydro-2H-pyran-4-ylamino)-pyrido[2,3-d]pyrimidin-7(8H)-one ([125I]4-IR)) in a mouse inflammatory model. METHODS: We designed [77Br]4-BR by replacing the radioiodine of [125I]4-IR or the fluorine of R1487 with radiobromine at the 4-position of the phenoxy ring. We synthesized 4-BR via a four-step process. The inhibitory potency of 4-BR was measured using an ADP-Glo™ kinase assay system. Radiosynthesis of [77Br]4-BR was performed via an organotin-radiobromine exchange reaction using the corresponding tributyltin precursor. Radioactivity biodistribution was evaluated in normal ddY mice and turpentine oil-induced inflammation model mice for 120 min after intravenous administration of [77Br]4-BR. The temporal changes in radioactivity in blood fractions were compared between [77Br]4-BR and [125I]4-IR. RESULTS: 4-BR was synthesized at a total yield of 9.1% and showed a p38α inhibitory potency similar to that of 4-IR. [77Br]4-BR was successfully obtained from a tributyltin precursor with high radiochemical yield (89.9%), purity (95.9%), and molar activity (2.0 TBq/µmol). [77Br]4-BR showed accumulation of high radioactivity in the inflamed tissue (3.4% ± 0.9% ID/g, peaking at 15 min), rapid delivery throughout the body, and rapid blood clearance with approximately half of the blood radioactivity existing as an intact form at 60 min. Although the maximum radioactivity accumulation in inflamed tissue after [77Br]4-BR administration was approximately half that of [125I]4-IR because of its faster blood clearance and lower free fraction in the input function, the inflamed tissue-to-blood ratio was comparable between [77Br]4-BR and [125I]4-IR. CONCLUSIONS: [77Br]4-BR would be a promising imaging agent for detecting activated p38α in inflammatory diseases.