High Binding Ratios Research Articles

TLR2 agonist prevents the progression of periapical lesions in mice by reducing osteoclast activity and regulating the frequency of Tregs.

To evaluate the role of regulatory T lymphocytes (Tregs) in the presence or absence of the synthetic ligand Pam3Cys during the progression of periapical lesion in wild-type (WT) and toll-like receptor 2 knockout (TLR2KO) mice. A total of 130 C57BL/6 male WT and TLR2KO mice were allocated into control (n = 5) and experimental (periapical lesion induction) (n = 10) groups. In specific groups (WT+Pam3cys and TLR2KO+Pam3cys), the synthetic ligand Pam3cys was administered intraperitoneally every 7 days, according to the experimental period (14, 21 and 42 days). At the end of those periods, the animals were euthanized, and the mandible and the spleen were submitted to histotechnical processing. Mandible histological sections were analysed by haematoxylin and eosin, TRAP histoenzymology and immunohistochemistry (FOXP3, RANK, RANKL and OPG). Spleen sections were analysed by immunohistochemistry (FOXP3). The inflammatory infiltrate and bone resorption were more intense in the TLR2KO group compared to the WT group. The animals that received the Pam3cys had smaller periapical lesions when compared to the animals that did not receive the ligand (p < .05). TLR2KO animals showed a significant increase in the number of osteoclasts when compared to TLR2KO+Pam3cys group (p < .05). At 21 days, the WT+Pam3cys group had a lower number of osteoclasts when compared to the WT animals (p = .02). FOXP3 expression was more intense in the WT+Pam3cys groups when compared to the WT animals in the 42 days (p = .03). In the spleen analysis, the WT+Pam3cys group also had a higher expression of FOXP3 when compared to the WT animals at 14 and 42 days (p = .02). Concerning RANKL, there was a reduction in staining in the KOTLR2+Pam3cys groups at 21 and 42 days (p = .03) and a higher binding ratio between RANK/RANKL in animals that did not receive the ligand. Administration of the Pam3cys increased the proliferation of Tregs, showed by FOXP3 expression and prevented the progression of the periapical lesion in WT mice. On the other hand, in the TLR2KO animals, Treg expression was lower with larger areas of periapical lesions. Finally, systemic administration of the Pam3cys in KO animals was able to limit the deleterious effects of the absence of the TLR2 receptor.

Prevalence and pathogenesis of macro-TSH in neonates: Analysis of umbilical cord blood from 939 neonates and their mothers.

Background: Macro-thyroid stimulating hormone (macro-TSH) is a large molecular weight TSH that causes elevated serum TSH concentrations due to its slow clearance. It is primarily a complex of TSH and anti-TSH autoantibodies. The aims of this study were to examine the prevalence and nature of macro-TSH in neonates and to determine how to cope with macro-TSH in neonates suspected to have congenital hypothyroidism through neonatal mass screening. Methods: The presence of macro-TSH was examined using polyethylene glycol (PEG), gel filtration chromatography (GFC), and 125I-TSH binding studies in 939 umbilical cord blood samples from neonates and their mothers. Results: Among 138 serum samples with a PEG precipitation ratio of TSH > 68.9% (mean + SD), human anti-mouse antibodies were found in nine samples. The presence of macro-TSH was examined in the remaining 129 serum samples using a 125I-TSH binding study and GFC. The 125I-TSH binding study revealed that four babies (0.43%) had significantly high ratios of 125I-TSH binding to their sera. Two of the babies were siblings, and their mother and the other two mothers also showed significantly high binding ratios. The 125I-TSH binding was displaced by a large amount (1 µg) of unlabeled human TSH in a similar way between babies and their mothers in all cases, suggesting the presence of anti-TSH autoantibodies in their sera. Further characterization of the autoantibodies in one baby and its mother showed a low affinity and high specificity to human TSH, and the nature was very similar between them. These findings may indicate that the anti-TSH autoantibodies that developed in the mother were transferred to the baby through the placenta and formed macro-TSH by binding to neonatal TSH. GFC revealed macro-TSH in only one baby and its mother, probably because of the dissociation of TSH from autoantibodies during the analytical procedure. Conclusions: Macro-TSH was found in 0.43% of neonates and their mothers all had macro-TSH as well. We recommend that if a baby's serum TSH concentration is high enough to consider levothyroxine treatment suspecting congenital hypothyroidism but the fT4 level is normal, their mother's macro-TSH should be checked.

An Albumin-Binding PSMA Ligand with Higher Tumor Accumulation for PET Imaging of Prostate Cancer.

Prostate-specific membrane antigen (PSMA) is an ideal target for the diagnosis and treatment of prostate cancer. Due to the short half-life in blood, small molecules/peptides are rapidly cleared by the circulatory system. Prolonging the half-life of PSMA probes has been considered as an effective strategy to improve the tumor detection. Herein, we reported a 64Cu-labeled PSMA tracer conjugating with maleimidopropionic acid (MPA), 64Cu-PSMA-CM, which showed an excellent ability to detect PSMA-overexpressing tumors in delayed time. Cell experiments in PSMA-positive 22Rv1 cells, human serum albumin binding affinity, and micro-PET imaging studies in 22Rv1 model were performed to investigate the albumin binding capacity and PSMA specificity. Comparisons with 64Cu-PSMA-BCH were performed to explore the influence of MPA on the biological properties. 64Cu-PSMA-CM could be quickly prepared within 30 min. The uptake of 64Cu-PSMA-CM in 22Rv1 cells increased over time and it could bind to HSA with a high protein binding ratio (67.8 ± 1.5%). When compared to 64Cu-PSMA-BCH, 64Cu-PSMA-CM demonstrated higher and prolonged accumulation in 22Rv1 tumors, contributing to high tumor-to-organ ratios. These results showed that 64Cu-PSMA-CM was PSMA specific with a higher tumor uptake, which demonstrated that MPA is an optional strategy for improving the radioactivity concentration in PSMA-expressing tumors and for developing the ligands for PSMA radioligand therapy.

Ultrasound-induced red bean protein–lutein interactions and their effects on physicochemical properties, antioxidant activities and digestion behaviors of complexes

The complexation of red bean protein (RBP) and lutein (LU) enhances the functional properties of proteins and increases their applicability as nutraceuticals for LU. The effects of ultrasonic treatment on the interactions between RBP and LU, physicochemical properties, antioxidant activities, and digestion behaviors of RBP–LU complexes were investigated. Ultrasonic treatment promoted the non-covalent binding of RBP with LU through hydrophobic interactions, causing the binding site to be closer to tryptophan than tyrosine. This process resulted in a more flexible secondary structure and less compact tertiary conformation of complexes with higher binding ratios, surface hydrophobicity, surface charge, and smaller particle size. The enhanced RBP–LU interactions and structural modifications of complexes induced by ultrasound possibly enhanced the functional properties. Ultrasonic treatment maximized the emulsifying and antioxidant activities of these complexes, and also enhanced the release of LU in the intestine by altering the digestion behavior of the complex, thereby improving its bioaccessibility. An appropriate ultrasonic treatment could facilitate RBP–LU interactions and modify the structure of complexes to improve their functional properties.

Variability in detection of SARS-CoV-2-specific antibody responses following mild infection: a prospective multicentre cross-sectional study, London, United Kingdom, 17 April to 17 July 2020.

IntroductionImmunoassays targeting different SARS-CoV-2-specific antibodies are employed for seroprevalence studies. The degree of variability between immunoassays targeting anti-nucleocapsid (anti-NP; the majority) vs the potentially neutralising anti-spike antibodies (including anti-receptor-binding domain; anti-RBD), particularly in mild or asymptomatic disease, remains unclear.AimsWe aimed to explore variability in anti-NP and anti-RBD antibody detectability following mild symptomatic or asymptomatic SARS-CoV-2 infection and analyse antibody response for correlation with symptomatology.MethodsA multicentre prospective cross-sectional study was undertaken (April–July 2020). Paired serum samples were tested for anti-NP and anti-RBD IgG antibodies and reactivity expressed as binding ratios (BR). Multivariate linear regression was performed analysing age, sex, time since onset, symptomatology, anti-NP and anti-RBD antibody BR.ResultsWe included 906 adults. Antibody results (793/906; 87.5%; 95% confidence interval: 85.2–89.6) and BR strongly correlated (ρ = 0.75). PCR-confirmed cases were more frequently identified by anti-RBD (129/130) than anti-NP (123/130). Anti-RBD testing identified 83 of 325 (25.5%) cases otherwise reported as negative for anti-NP. Anti-NP presence (+1.75/unit increase; p < 0.001), fever (≥ 38°C; +1.81; p < 0.001) or anosmia (+1.91; p < 0.001) were significantly associated with increased anti-RBD BR. Age (p = 0.85), sex (p = 0.28) and cough (p = 0.35) were not. When time since symptom onset was considered, we did not observe a significant change in anti-RBD BR (p = 0.95) but did note decreasing anti-NP BR (p < 0.001).ConclusionSARS-CoV-2 anti-RBD IgG showed significant correlation with anti-NP IgG for absolute seroconversion and BR. Higher BR were seen in symptomatic individuals, particularly those with fever. Inter-assay variability (12.5%) was evident and raises considerations for optimising seroprevalence testing strategies/studies.

&lt;i&gt;In vivo&lt;/i&gt; and &lt;i&gt;In vitro&lt;/i&gt; Mitigation Effects of Lactic Acid Bacteria Derived from Fresh Vegetables on Aflatoxins

Aflatoxins (AFs) are known to be oncogenic mycotoxins. This study investigated the mitigation effects of lactic acid bacteria (LAB) isolated from four types of vegetable, cucumber, Chinese cabbage, Japanese radish and eggplant, which are used to make Japanese traditional fermented pickles, on AFs. Using aflatoxin M1 (AFM1) binding assay for screening, four representative strains were selected (one from each vegetable) from total 94 LAB strains, based on the highest binding ratio. The ranges of the binding ratio of these representative strains to aflatoxin B1 (AFB1), aflatoxin B2, aflatoxin G1, aflatoxin G2 and AFM1 were 57.5%-87.9% for the LAB strain derived from cucumber, 18.9%-43.9% for the LAB strain derived from Chinese cabbage, 26.4%-41.7% for the LAB strain derived from Japanese radish, and 15.0%-42.6% for the LAB strain derived from eggplant. The strains isolated from cucumber, Chinese cabbage, Japanese radish and eggplant were identified as Lactococcus lactis subsp. lactis, Weissella cibaria, Leuconostoc mesenteroides and Leu. mesenteroides, respectively. An in vitro binding assay of the four strains under acidic conditions showed that the number of living bacteria decreased, while the binding ratio increased in some strains, suggesting that the LAB maintained their capacity to bind aflatoxins even in an environment that imitated the stomach. An in vivo experiment using L. lactis subsp. lactis derived from cucumber revealed that the bacteria significantly inhibited the absorption of AFB1 into blood. These results showed that the LAB used for Japanese vegetable pickles was an effective binding agent of AFs and suggested that they might play a role in mitigating AF absorption.

In vivo and in vitro studies of Banhahoobak-tang tablets using UPLC-ESI-MS/MS with polarity switching

Banhahoobak-tang is the most prescribed herbal drug in East Asia when individuals experience sudden symptoms such as sore throat or neurological symptoms. The low toxicity and high in-vivo safety of this herbal medicine has made it more attractive to patients, and it has recently been formulated as tablets. In addition, Banhahoobak-tang tablets are registered as health insurance drugs in South Korea, and clinical prescriptions and demand are increasing. However, there are very few clinical trial data as well as very little accurate content analysis and results for Banhahoobak-tang tablets. The purpose of this study was to perform in-vitro and in-vivo studies on Banhahoobak-tang tablets, including content analysis, pharmacokinetics in humans, and plasma protein binding. For this study, a UPLC-ESI-MS/MS method with polarity switching was developed for simultaneous analysis of 18 components of Banhahoobak-tang. To separate the analytes, a C8 reverse-phase column was used as the stationary phase, 0.1 % aqueous formic acid and acetonitrile as the mobile phase, and ionization and multiple reaction monitoring for quantification. The developed method was able to isolate and quantify the 18 components with good sensitivity and selectivity and was fully validated according to international analytical standards. Stability tests were also conducted on the analytes. Finally, the method was applied to in-vitro and in-vivo studies of Banhahoobak-tang tablets, and the tablet components were 52.49 ng/g to 91.00 μg/g on average. The detected components showed rapid oral absorption in humans as well as high plasma protein binding ratio overall. These results and methods can be useful not only for effectiveness and safety evaluation but also for quality control of Banhahoobak-tang tablets.

Effects of concentration of flavor compounds on interaction between soy protein isolate and flavor compounds

Headspace-solid-phase microextraction/gas chromatography-mass spectrometry, static headspace/gas chromatography, fluorescence spectroscopy, zeta potential, particle size, and surface hydrophobicity were used to obtain comprehensive and precise information on binding properties. Klotz and Hill binding models were compared at low and high concentration of flavors binding to soy protein isolate (SPI) to estimate binding parameters in aqueous solution. These results show the low molar binding ratio of flavor bind to SPI induced almost no conformational changes in SPI, which indicates linear binding; the high molar binding ratio induced conformational changes in SPI (citronellal > citronellyl acetate > citronellyl propionate > citronellol), indicating nonlinear binding; salting out of α–pinene, and terpinene were analyzed at low and high molar ratios of flavors binding to SPI. These results indicate the necessity of multiple analytical methods to accurately assess interactions between flavor compounds and proteins in food and contribute to current knowledge regarding flavor profiles.

Preparation of molecularly imprinted copoly(acrylic acid-divinylbenzene) for extraction of environmentally relevant sertraline residues

Aiming to improve the sensitivity and specificity of environmental analysis of sertraline residues, we investigated the use of molecularly imprinted polymers (MIPs) as the solid phase extraction sorbents for rebinding of the parent drug, its metabolite norsertaline and its transformation product (TP) sertraline ketone. MIPs were synthesized using the antidepressant sertraline as the target, where two polymerisation approaches were used: polymerisation in solvent and two-stage polymerisation. Leaching of sertraline from the imprinted material presented a major problem in case of MIPs polymerised in solvent. On the contrary, by synthetizing MIPs with the two-stage polymerisation approach, sertraline was removed to below the limit of quantification (LOQ). Two MIPs were selected that showed the best binding characteristics during the solid phase extraction rebinding experiments: one was prepared by polymerisation in solvent and another one by two-stage polymerisation. The former bound norsertraline and sertraline ketone in a high binding ratio compared to its non-imprinted analogue, while the latter had the highest imprinting factor for sertraline. Based on the proved cross-selectivity of MIPs, we propose their use in enrichment, purification and isolation of transformation products, including novel compounds that have before not been recognised.

Investigation of manganese(II)-insulin complexes using electrospray ionization mass spectrometry

Manganese is a trace element in human nutrition. It is involved in many enzymes, proteins and biological activities. Mn(II) ion has the capable of binding to protein or peptides. Insulin is a blood glucose-lowering peptide hormone and it is secreted by the pancreatic β-cells. In this study, the binding capability of Mn(II) ions to insulin was studied using ESI-MS, nano-ESI-MS and MS/MS methods. The binding of Mn(II) ions to insulin molecule was studied by examining the effect of pH, the molar ratio of Mn(II) ions to insulin, the flow rate with nano-ESI system and MS/MS spectrometry. The ESI-MS measurements showed that the Mn(II)-insulin complexation mostly produces ML and M2L type complexes. The highest binding ratio was found at pH 7. The complex formation equilibrium constants of Mn(II)-insulin were calculated as Kf1: 1.03 ± 0.12 × 104 and Kf2: 1.93 ± 0.17 × 103. The nano-ESI-MS and MS/MS measurements exhibited strong and specific binding of Mn(II) ions to insulin molecule. It was concluded from all the ESI-MS measurements that Mn(II) ion has a high affinity to insulin molecule to form stable complexes.

Undulating the Lamellar Interface of Polymer–Surfactant Complex by Dendrimer

Self-assembly of the supramolecules formed by the complexation between poly(amidoamine) (PAMAM) generation four (G4) dendrimer and the surfactant, dodecylbenzenesulfonic acid (DBSA), generated a hexagonal columnar phase and a typical flat lamellar phase at low and high surfactant binding ratio, respectively, due to the dominance of the dendrimer to attain its natural curvature and the surfactant alkyl tails to reduce their hydrophobic interaction energy and packing frustration. Most strikingly, the delicate balance between these two free energy components at the intermediate binding ratios resulted in an undulated lamellar structure characterized by the centered rectangular unit cell. The finding demonstrates the power of dendrimer as a building block for expanding the morphological window of polymeric assemblies by modulating the interfacial curvature of microphase-separated structure.

Effective binding of perhalogenated closo-borates to serum albumins revealed by spectroscopic and ITC studies

The interactions of boron cluster compounds closo-borates with biomolecules are widely studied due to their efficiency as agents for boron neutron capture therapy of cancer. In present work the binding abilities of anionic halogen closo-borates [B10Hal10]2− (Hal = Cl, Br, I) and [B12Hal12]2− (Hal = Cl, I) towards bovine and human serum albumins were investigated by spectroscopic and isothermal titration calorimetry (ITC) methods. The protein fluorescence quenching method and ITC studies confirmed the complex formation. The degree of protein fluorescence quenching increased from chlorine to iodine boron derivatives that is attributed to external heavy atom effect. The ITC data point on the existence in the protein structure of two types of binding sites: with higher and lower affinity to closo-borates. Albumin-closo-borate complex binding ratio, n (4–5 anions per protein molecule) is higher than for the parent hydrogen closo-borates (2 anions per protein molecule). Binding constants estimated by fluorescent and ITC methods indicate higher affinity of halogen closo-borates to albumins (K in the range of 104–106 M−1) comparing to that of the hydrogen closo-borate (K about 103 M−1). Due to their high affinity and high binding ratio to albumins halogen closo-borates are proposed for further studies as agents for boron neutron capture therapy.

Development of a series of novel carbon-11 labeled PDE10A inhibitors.

Phosphodiesterase 10A (PDE10A) is a member of the PDE family of enzymes that degrades cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP). Our aim was to label a series of structurally related PDE10A inhibitors with carbon-11 and evaluate them as potential positron emission tomography (PET) radioligands for PDE10A using nonhuman primates. The series consisted of seven compounds based on the 3-(1H-pyrazol-5-yl)pyridazin-4(1H)-one backbone. These compounds were selected from the initial larger library based on a number of parameters such as affinity, selectivity for hPDE10A in in vitro tests, lipophilicity, and on the results of multidrug resistance protein 1 (MDR1)-LLCPK1 and the parallel artificial membrane permeability assays. Seven radioligands (KIT-1, 3, 5, 6, 7, 9, and 12) were radiolabeled with carbon-11 employing O-methylation on the hydroxyl moiety using [(11)C]methyl triflate. In vivo examination of each radioligand was performed using PET in rhesus monkeys; analysis of radiometabolites in plasma also was conducted using HPLC. All seven radioligands were labeled with high (>90%) incorporation of [(11)C]methyl triflate into their appropriate precursors and with high specific radioactivity. Carbon-11 labeled KIT-5 and KIT-6 showed high accumulation in the striatum, consistent with the known anatomical distribution of PDE10A in brain, accompanied by fast washout and high specific binding ratio. In particular [(11)C]KIT-6, named [(11)C]T-773, is a promising PET tool for further examination of PDE10A in human brain.

Interaction Between Surfactin and Bovine Serum Albumin

The interaction of surfactin, a typical biosurfactant, with bovine serum albumin (BSA) was investigated by surface tension, fluorescence, freeze-fractured transmission electron microscopy (FF-TEM) and circular dichroism (CD) measurements. The surface tension curves of pure surfactin solution and surfactin/BSA solutions have different phenomena, where two obvious inflections determined as the critical aggregation concentration (cac) and the critical micelle concentration (cmc) appear for surfactin/BSA solutions. The higher BSA concentration, the higher cac and cmc values for surfactin/BSA solution. Fluorescence spectra show that the structure change of BSA is dependent on both surfactin and BSA concentration. The micropolarity, FF-TEM and CD results further demonstrate the interaction between BSA and surfactin. The excess free energy (ΔG0) of surfactin/BSA interactions have been obtained as −6.13 and 5.32 kJ/mol for 1.0 × 10−6 and 3.8 × 10−6 mol/L BSA concentration, respectively. The binding ratio (R) determined for surfactin/BSA systems are higher than that reported for dirhamnolipid to BSA. Above all, it can be concluded that the hydrophobic interaction and the hydrogen bonds between surfactin and BSA play the key role for the high binding ratio for surfactin to BAS.

P3-129 - The Effects of Adding Loxoprofen to the Treatment with Fluorouracil Anticancer Drug for the Patients Taking Warfarin

ABSTRACT Preface Warfarin is widely used as a preventive and therapeutic medicine for thromboembolism, and there are not a few cases in which the patients who are to undergo anticancer drug treatment are taking it. Warfarin interacts with many medicines and one should take precautions in the treatment with fluorouracil anticancer drug against the situations where the anticoagulant effect is strengthened and bleeding complications appear. I experienced a few cases in which the patients who were carefully achieving good control of combined application had serious bleeding complications when they were given Loxoprofen additionally, on which I will report. Background The factors of bleeding complications are ‘combined application of loxoprofen, which has a high protein binding ratio, inhibits the metabolism of warfarin and strengthens the anticoagulant effect’, ‘the platelet aggregation inhibition effect of loxoprofen’, ‘furthering the bleeding tendency by loxoprofen's inhibiting COX-1 such as gastrointestinal tract ulcer’, etc. The course of the cases The regimen is mFOLFOX, FOLFIRI and XELOX. In every case, the patient was taking warfarin orally and then added with the treatment with fluorouracil anticancer drug and the dose was changed on a timely basis, which had favorable outcome of anticoagulation treatment, and it was to such condition where loxoprofen was added. The serious bleeding occurred 12 days after taking Loxoprofen for the early cases and 18 days for the late cases, and some needed blood transfusion as PT-INR was 15.42. Issue Loxoprofen is frequently used to control cancer-related pain and it is highly likely that patients undergoing anticancer drug treatment are administered with it. In addition to cancer-related pain, it can be also prescribed in the areas of dentistry and orthopedics. It is necessary to actively disseminate such interaction information to medical staff and patients.

Determination of the toluene diisocyanate binding sites on human serum albumin by tandem mass spectrometry

Diisocyanates are highly reactive chemical compounds widely used in the manufacture of polyurethanes. Although diisocyanates have been identified as causative agents of allergic respiratory diseases, the specific mechanism by which these diseases occur is largely unknown. To better understand the chemical species produced when diisocyanates react with protein, tandem mass spectrometry was employed to unambiguously identify the binding sites of the industrially important isomers, 2,4- and 2,6-toluene diisocyanate, on human serum albumin at varying diisocyanate/protein ratios. The 2,4-isomer results in approximately 2-fold higher conjugation product ion abundances than does the 2,6-isomer, suggesting that the 2,4-isomer has a higher reactivity toward albumin. Both isomers preferentially react with the N-terminal amine of the protein and the ε-NH 2 of lysine. At a low (1:2) diisocyanate/protein ratio, five binding sites are identified, whereas at a high (40:1) ratio, near-stoichiometric conjugation is observed with a maximum of 37 binding sites identified. Binding sites observed at the lowest conjugation ratios are conserved at higher binding ratios, suggesting a subset of 5–10 preferential binding sites on albumin. Diisocyanate–protein conjugation results in a variety of reaction products, including intra- and intermolecular crosslinking, diisocyanate self-polymerization, and diisocyanate hydrolysis.

Pharmacological Evaluation of Halogenated and Non-halogenated Arylpiperazin-1-yl-ethyl-benzimidazoles as D2 and 5-HT2A Receptor Ligands

Five groups of previously synthesized and initially screened non-substituted and 4-halogenated arylpiperazin-1-yl-ethyl-benzimidazoles were estimated for their in-vitro binding affinities at the rat D(2) , 5-HT(2A) , and α(1) -adrenergic receptors. Among all these compounds, 2-methoxyphenyl and 2-chlorophenyl piperazines demonstrate the highest affinities for the tested receptors. The effects of 4-halogenation of benzimidazoles reveal that substitution with bromine may greatly increase the affinity of the compounds for the studied receptors, while the effect of substitution with chlorine is less remarkable. Most of the tested components show 5-HT(2A)/D(2) pK(i) binding ratios slightly above or less than 1, while only 4-chloro-6-(2-{4-[3-(trifluoromethyl)phenyl]piperazin-1-yl}ethyl)-1H-benzimidazole expresses an appropriate higher binding ratio (1.14), which was indicated for atypical neuroleptics. This compound exhibits a non-cataleptic action in rats and prevents d-amphetamine-induced hyperlocomotion in mice, which suggest its atypical antipsychotic potency.

Pharmacokinetics of [18F]fluoroalkyl derivatives of dihydrotetrabenazine in rat and monkey brain

The specific binding and regional brain pharmacokinetics of new fluorine-18 ([(18)F])-labeled radioligands for the vesicular monoamine transporter (VMAT2) were examined in the rat and primate brain. In the rat, 9-[(18)F]fluoropropyl-(+/-)-9-O-desmethyldihydrotetrabenazine ([(18)F]FP-(+/-)-DTBZ) showed better specific binding in the striatum than either (+)-[(11)C]dihydrotetrabenazine ((+)-[(11)C]DTBZ) or 9-[(18)F]fluoroethyl-(+/-)-9-O-desmethyldihydrotetrabenazine ([(18)F]FE-(+/-)-DTBZ). Using microPET, the regional brain pharmacokinetics of [(18)F]FE-(+/-)-DTBZ, [(18)F]FP-(+/-)-DTBZ and (+)-[(11)C]DTBZ were examined in the same monkey brain. (+)-[(11)C]DTBZ and [(18)F]FP-(+/-)-DTBZ showed similar brain uptakes and pharmacokinetics, with similar maximum striatum-to-cerebellum ratios (STR/CBL=5.24 and 5.15, respectively) that were significantly better than obtained for [(18)F]FE-(+/-)-DTBZ (STR/CBL=2.55). Striatal distribution volume ratios calculated using Logan plot analysis confirmed the better specific binding for the fluoropropyl compound [distribution volume ratio (DVR)=3.32] vs. the fluoroethyl compound (DVR=2.37). Using the resolved single active isomer of the fluoropropyl compound, [(18)F]FP-(+)-DTBZ, even better specific to nonspecific distribution was obtained, yielding the highest distribution volume ratio (DVR=6.2) yet obtained for a VMAT2 ligand in any species. The binding of [(18)F]FP-(+)-DTBZ to the VMAT2 was shown to be reversible by administration of a competing dose of unlabeled tetrabenazine. Metabolic defluorination was slow and minor for the [(18)F]fluoroalkyl-DTBZ ligands. The characteristics of high specific binding ratio, reversibility, metabolic stability and longer half-life of the radionuclide make [(18)F]FP-(+)-DTBZ a promising alternative VMAT2 radioligand suitable for widespread use in human positron emission tomography studies of monoaminergic innervation of the brain.

A Polarized-Light Spectroscopy Study of Interactions of a Hairpin Polyamide with DNA

We here study the interactions of a polyamide with large DNA, and compare to those of minor groove binder distamycin (DST), including high ligand/DNA binding ratios. Specific as well as nonspecific binding is probed using polarized-light spectroscopy combined with singular value decomposition analysis. Circular and linear dichroism data confirm binding geometries consistent with minor groove binding for both of the ligands. Interestingly, at high and intermediate ligand/DNA ratios the polyamide exhibits no significant sequence discrimination between mixed-sequence (calf thymus) and AT DNA as compared to DST. Each ligand is concluded to exhibit two different binding modes depending upon ligand/DNA ratio and nucleo-base sequence. At high binding ratios, distinct differences between the ligands are observed: circular dichroism spectra exciton effects provide evidence of bimolecular interactions of the polyamide when bound to AT-DNA, whereas no effects are seen with DST or mixed-sequence DNA. Also linear dichroism indicates that a change in binding geometry occurs at high polyamide/AT ratios, and that the effect occurs only with polyamide in contrast to DST. Since the effect is insignificant with DST, or with calf thymus DNA, it is concluded that it relates to the sizes of the ligands and the minor grooves, becoming critical in the limit of crowding.

Selection of the alkylamino group introduced into the polymer chain grafted onto a porous membrane for the impregnation of an acidic extractant

Bis(2-ethylhexyl)phosphate (HDEHP) as an acidic extractant selective for lanthanides was impregnated into an alkylamino group introduced into a polymer chain grafted onto the pore surface of a porous hollow-fiber membrane. The alkyl chain length was varied from 2 to 18 carbons by reacting the glycidyl methacrylate (GMA)-grafted hollow fiber with various alkylamines. The HDEHP-impregnated porous hollow-fiber membrane originating from the GMA-grafted hollow fiber prepared with methanol used as a solvent for GMA had the highest pure water flux among that prepared with methanol, ethanol, 1-propanol, or 1-butanol. The HDEHP impregnated into the dodecyl- or octadecylamino group of the graft chain exhibited the absence of leakage in a high ionic strength medium as well as a high binding ratio of yttrium ion to the impregnated HDEHP. The equilibrium binding capacity of the HDEHP-impregnated porous hollow-fiber membrane for yttrium ion was 0.6 mol/kg of the GMA-grafted hollow-fiber membrane. The HDEHP impregnated into the dodecylamino group was found to efficiently bind yttrium ion in a permeation mode.