[18F]-florbetaben PET/CT Imaging in the Alzheimer's Disease Mouse Model APPswe/PS1dE9.

BACKGROUND: Positron-emission-tomography (PET) using 18F labeled florbetaben allows noninvasive in vivo-assessment of amyloid-beta (Aß), a pathological hallmark of Alzheimer's disease (AD). In preclinical research, [18F]-florbetaben-PET has already been used to test the amyloid-lowering potential of new drugs, both in humans and in transgenic models of cerebral amyloidosis. The aim of this study was to characterize the spatial pattern of cerebral uptake of [18F]-florbetaben in the APPswe/ PS1dE9 mouse model of AD in comparison to histologically determined number and size of cerebral Aß plaques. METHODS: Both, APPswe/PS1dE9 and wild type mice at an age of 12 months were investigated by smallanimal PET/CT after intravenous injection of [18F]-florbetaben. High-resolution magnetic resonance imaging data were used for quantification of the PET data by volume of interest analysis. The standardized uptake values (SUVs) of [18F]-florbetaben in vivo as well as post mortem cerebral Aß plaque load in cortex, hippocampus and cerebellum were analyzed. RESULTS: Visual inspection and SUVs revealed an increased cerebral uptake of [18F]-florbetaben in APPswe/ PS1dE9 mice compared with wild type mice especially in the cortex, the hippocampus and the cerebellum. However, SUV ratios (SUVRs) relative to cerebellum revealed only significant differences in the hippocampus between the APPswe/PS1dE9 and wild type mice but not in cortex; this differential effect may reflect the lower plaque area in the cortex than in the hippocampus as found in the histological analysis. CONCLUSION: The findings suggest that histopathological characteristics of Aß plaque size and spatial distribution can be depicted in vivo using [18F]-florbetaben in the APPswe/PS1dE9 mouse model.

Targeting myelin lipid metabolism as a potential therapeutic strategy in a model of CMT1A neuropathy.

In patients with Charcot-Marie-Tooth disease 1A (CMT1A), peripheral nerves display aberrant myelination during postnatal development, followed by slowly progressive demyelination and axonal loss during adult life. Here, we show that myelinating Schwann cells in a rat model of CMT1A exhibit a developmental defect that includes reduced transcription of genes required for myelin lipid biosynthesis. Consequently, lipid incorporation into myelin is reduced, leading to an overall distorted stoichiometry of myelin proteins and lipids with ultrastructural changes of the myelin sheath. Substitution of phosphatidylcholine and phosphatidylethanolamine in the diet is sufficient to overcome the myelination deficit of affected Schwann cells in vivo. This treatment rescues the number of myelinated axons in the peripheral nerves of the CMT rats and leads to a marked amelioration of neuropathic symptoms. We propose that lipid supplementation is an easily translatable potential therapeutic approach in CMT1A and possibly other dysmyelinating neuropathies.

Decitabine demonstrates antileukemic activity in B cell precursor acute lymphoblastic leukemia with MLL rearrangements.

BACKGROUND: Promotor hypermethylation of CpG islands is common in B cell precursor acute lymphoblastic leukemia (BCP-ALL) with mixed lineage leukemia (MLL) gene rearrangements. Hypomethylating agents (HMA) such as azacitidine (AZA) and decitabine (DEC) reduce DNA hypermethylation by incorporation into DNA and were successfully introduced into the clinic for the treatment of myeloid neoplasias. METHODS: Here, we investigated whether HMA induce comparable biological effects in MLL-positive BCP-ALL. Further, efficacy of HMA and concomitant application of cytostatic drugs (cytarabine and doxorubicin) were evaluated on established SEM and RS4;11 cell lines. In addition, promising approaches were studied on BCP-ALL cell line- and patient-derived xenograft models. RESULTS: In general, DEC effects were stronger compared to AZA on MLL-positive BCP-ALL cells. DEC significantly reduced proliferation by induction of cell cycle arrest in G0/G1 phase and apoptosis. Most sensitive to HMA were SEM cells which are characterized by a fast cell doubling time. The combination of low-dose HMA and conventional cytostatic agents revealed a heterogeneous response pattern. The strongest antiproliferative effects were observed when ALL cells were simultaneously exposed to HMA and cytostatic drugs. Most potent synergistic effects of HMA were induced with cytarabine. Finally, the therapeutic potential of DEC was evaluated on BCP-ALL xenograft models. DEC significantly delayed leukemic proliferation in xenograft models as demonstrated longitudinally by non-invasive bioluminescence as well as 18F-FDG-PET/CT imaging. Unexpectedly, in vivo concomitant application of DEC and cytarabine did not enhance the antiproliferative effect compared to DEC monotherapy. CONCLUSIONS: Our data reveal that DEC is active in MLL-positive BCP-ALL and warrant clinical evaluation.

Terrestrial toxicity evaluation of decabromodiphenyl ethane on organisms from three trophic levels.

Decabromodiphenyl ethane (DBDP-Ethane) was evaluated for its potential to effect sewage sludge respiration, soil nitrification, survival and reproduction in Eisenia fetida, and seedling emergence and growth in Zea mays, Lolium perenne, Glycine max, Allium cepa, Lycopersicon esculentum, and Cucumis sativa. The no observed effect concentrations (NOECs) were identified at the limit concentration level for sewage sludge respiration (>10 mg DBDP-Ethane/kg dry soil), >2500 mg/kg dry soil for soil nitrification, >3720 mg/kg dry soil for earthworm survival, and >6250 mg/kg dry soil for seedling emergence and growth in Z. mays, L. perenne, and G. max . Treatment-related effects were identified for E. fetida reproduction, C. sativa survival, and L. esculentum and A. cepa height and dry weight. The most sensitive endpoints were decreased height and dry weight for A. cepa and decreased reproduction for E. fetida with NOECs of 1563(nominal) (1540(measured)) and 2210(nominal) (1907(mean measured)) mg/kg dry soil. The NOEC for soil nitrification and the lowest NOEC identified for soil (i.e., A. cepa) were used to derive predicted no effect concentrations (PNEC) values of 2500 mg/kg for sewage sludge and 156 mg/kg for soil. The calculated PNECs indicate DBDP-Ethane presents little risk to organisms in the sewage sludge and soil compartments.

Preparation and characterization of dialkylphosphoryl-obidoxime conjugates, potent anticholinesterase derivatives that are quickly hydrolyzed by human paraoxonase (PON1192Q).

The potential of the most active pyridinium-4-aldoximes, such as obidoxime and trimedoxime, to reactivate phosphorylated acetylcholinesterase is not fully exploited because of inevitable formation of phosphoryloximes (POXs) with extremely high anticholinesterase activity. Hence, a topochemical equilibrium is expected at the active site, with the freshly reactivated enzyme being rapidly re-inhibited by POX produced during reactivation. In the present study, dimethylphosphoryl-, diethylphosphoryl-, and diisopropyl-obidoxime conjugates were generated and isolated in substance. Their inhibition rate of acetylcholinesterase from human red cell membranes was by a factor of 2250, 480 and 600 higher than that observed with paraoxon-methyl, paraoxon-ethyl, and diisopropyl phosphorofluoridate, respectively. All three POXs were hydrolyzed by human paraoxonase (PON1), with the alloenzyme PON1192Q being about 50-fold more active than PON1192R. The rate of hydrolysis, yielding obidoxime, was 1:6:0.03 for the three POXs, respectively. The rate of non-enzymic degradation, yielding obidoxime mononitrile, was similar with the three POXs and showed a high dependency on the reaction temperature (activation energy 83 kJ/mol), while enzymic hydrolysis required less energy (16 kJ/mol). To determine POX-hydrolase activity, we preferred a reaction temperature of 20 degrees C to reduce the noise of spontaneous degradation. A plot of POX-hydrolase versus salt-stimulated paraoxonase activity showed a highly discriminating power towards the PON1Q192R alloenzymes, which may be based on repulsive forces of the quaternary nitrogen atoms of the protonated arginine subtype and the bisquaternary POXs. It is concluded that the pharmacogenetic PON1Q192R polymorphism may be another contributor to the large variability of susceptible subjects seen in obidoxime-treated patients.

Hyperoxic lung injury in Fischer-344 and Sprague-Dawley rats in vivo.

Supplemental oxygen remains an important therapy for pulmonary insufficiency, despite the potential adverse effects of hyperoxic exposures. Recently, He et al. reported that hyperoxic ventilation more readily damaged isolated perfused lungs from Fischer-344 rats than from Sprague-Dawley rats (Am. J. Physiol. 259:L451), which correlates with the previously reported strain differences in hepatic responses to diquat-induced oxidant stress in vivo (J. Pharmacol. Exp. Ther. 235:172). We therefore examined the differences in hyperoxic lung injury in Fischer-344 and Sprague-Dawley rats in vivo. Adult male rats were exposed to > 95% O2 and were sacrificed after 24, 48, or 60 h. Control animals were maintained in room air. Dramatically greater increases in pleural effusions and bronchoalveolar lavage protein concentrations in response to hyperoxia were observed in the Fischer-344 rats than in the Sprague-Dawley rats (p < .05 at both 48 and 60 h for both measurements). Additionally, the glutathione concentrations in alveolar lining fluid decreased from 800 microM to 115 microM in Fischer-344 rats after 60 h of > 95% O2, but did not change in Sprague-Dawley rats. We conclude that the greater susceptibility of Fischer-344 than of Sprague-Dawley rats to hyperoxic lung injury in vitro reported previously also is observed in vivo and that this strain difference offers unique opportunities to study mechanisms of hyperoxic lung injury.

Thieno-triazolo-1,4-diazepines as antagonists of platelet-activating factor: present status.

The platelet-activating factor, 1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine, (PAF) antagonistic activity of thienotriazolodiazepines has recently been described. The lead compound in this series was brotizolam, which also exhibits sedative and hypnotic effects. By combination of brotizolam with the benzodiazepine receptor antagonist RO 15-1788, PAF antagonistic and central nervous system (CNS) activities could be segregated. Systematic structure variation has led to potent and selective PAF antagonists without CNS effects. WEB 2086 and its analogues WEB 2170 and STY 2108 are representative examples of this structural type and have shown a high potency and selectivity in PAF-induced and PAF-dependent in vitro tests and in experimental models. Studies in healthy volunteers have demonstrated potent pharmacological activity and good safety and tolerance of oral, intravenous or inhaled WEB 2086 in man. These agents should therefore prove useful for the further elucidation of the pathophysiological role of PAF and provide an opportunity for therapeutic applications in diseases in which the involvement of PAF has been implicated.

Pharmacologic activity of bepafant (WEB 2170), a new and selective hetrazepinoic antagonist of platelet activating factor.

The hetrazepine WEB 2170 (international nonproprietary name: bepafant), a thieno-triazolodiazepine that is structurally related to the recently described platelet activating factor (PAF) antagonist WEB 2086, is a potent and selective PAF antagonist both in vitro and in vivo. WEB 2170 inhibited PAF-induced human platelet and neutrophil aggregation in vitro (IC50 values: 0.3 and 0.83 microM, respectively) but had little or no inhibitory action against aggregation induced by other agonists. The potency in vitro was comparable to that described recently for WEB 2086 (Casals-Stenzel, J., Muacevic, G. and Weber, K.H.: J. Pharmacol. Exp. Ther. 241: 974-981, 1987). When guinea pigs were given i.v. infusions of PAF at 30 ng x kg-1 x min-1, oral (0.005-0.5 mg/kg) as well as intravenous (0.005-0.05 mg/kg) treatment with WEB 2170 abrogated the intrathoracic accumulation of 111In-labeled platelets, the bronchoconstriction and the hypotension as well as the finally occurring death in a dose-dependent fashion. Oral (0.01-1 mg/kg) and intravenous (0.005-0.1 mg/kg) WEB 2170 shared with the beta 2 agonist fenoterol and the steroid dexamethasone the property of protecting elderly NMRI mice from the lethal effect of PAF. In anesthetized rats, intravenous (0.001-0.1 mg/kg) and oral (0.05-1 mg/kg) WEB 2170 inhibited PAF-induced hypotension in a dose-related manner. Coadministration of WEB 2170 inhibited PAF-induced increase of vascular permeability in rat skin very effectively. The half-time of duration of action in the rat was estimated to be about 5 to 6 h after oral administration and about 1.1 to 2.3 h after intravenous application. In conclusion, the hetrazepine WEB 2170 is a strong and selective PAF antagonist, which is in vitro more or less equipotent to WEB 2086. In contrast, in vivo oral WEB 2170 is--depending on the species and considered parameter--about 5 to 40 times more potent against exogenous PAF-induced alterations than the recently described hetrazepine WEB 2086. Particularly in mice and rats, oral WEB 2170 is by far superior to WEB 2086.

Percutaneous central venous catheterization in a pediatric intensive care unit: a survival analysis of complications.

We investigated the relationship between the duration of percutaneous central venous catheterization and the occurrence of catheter-related complications in critically ill children by survival analysis techniques. Data were collected prospectively and analyzed for infectious and noninfectious complications from 379 pediatric patients in whom central venous catheters had been placed in the pediatric ICU over a 45-month period. Cumulative survival rate analysis revealed a linear decrease in the number of complication-free catheters with time. The median duration of complication-free catheter survival was projected to be 23.3 days. The risk of catheter complication did not increase with increasing daily duration of catheter use as demonstrated by probability density function: catheter complication rates were similar on the first day after insertion (1.06 +/- 0.5%), the seventh day (4.27 +/- 1.6%), and the 24th day (2.48 +/- 2.4%). Therefore, in this population, routine catheter replacement would not be expected to lower the incidence of catheter-related complications, but may unnecessarily increase the number of insertion-related complications.

Percutaneous femoral venous catheterizations: a prospective study of complications.

In a prospective, 45-month study, we compared the complication rates of percutaneously placed femoral and nonfemoral central venous catheters in critically ill pediatric patients. Forty-one percent of the 395 central venous catheters placed during this interval were femoral. Noninfectious complications were recognized for 2.5% of femoral catheters and 2.1% of nonfemoral catheters. Only three complications occurred with catheter insertion, all during nonfemoral attempts. Systemic infections that were possibly attributable to the central venous catheter were found in 3.7% of patients with femoral catheters and 7.3% of those with nonfemoral catheters. Femoral venous catheterization offers several practical advantages for central access over other sites. The low incidence of complications documented in this study suggests that the femoral vein is the preferred site in most critically ill children when central venous catheterization is indicated.

Influence of nitrendipine and verapamil on plasma levels, urinary excretion, and beta-blocking effect of metoprolol.

Following randomized allocation eight healthy volunteers were treated for 1 week each with metoprolol alone (100 mg twice daily), verapamil 80 mg three times a day plus metoprolol 100 mg twice daily, and with nitrendipine 20 mg twice daily. Plasma levels and urinary recovery of the beta-blocker, antipyrine clearance, and heart rate on exercise were measured. Verapamil and nitrendipine slightly prolonged elimination half-life of metoprolol. The urinary recovery of the parent beta-blocker and of its alpha-hydroxy metabolite was elevated by both calcium antagonists (verapamil and nitrendipine). Exercise tachycardia (150 beats/min without drugs) was inhibited more pronounced on the combination therapies than under metoprolol administration alone. Results of the present study indicate that calcium antagonists enhance inhibition of exercise tachycardia caused by metoprolol, possibly due to their binding to myocardial beta-adrenergic receptors which is known from the literature. As both calcium antagonists did not increase plasma levels of metoprolol, in the present study a kinetic interaction between the beta-blocker and the calcium channel blockers investigated does not appear to be responsible for the pharmacodynamic effects observed.

Effect of the PAF-antagonist WEB 2086 on anaphylactic lung reaction: comparison of inhalative and intravenous challenge.

The specific and potent antagonist of platelet activating factor (PAF), WEB 2086, has been used to investigate the putative role of PAF in anaphylaxis in the guinea-pig. Intravenous (i.v.) challenge with ovalbumin has been compared to inhalative provocation. When actively sensitized guinea-pigs were challenged by the i.v. route, in the presence of mepyramine (5 micrograms/kg i.v.), oral WEB 2086 (0.05-0.5 mg/kg) inhibited the anaphylactic bronchoconstriction but not the hypotension. In passively sensitized guinea-pigs, i.v. WEB 2086, in the presence of mepyramine, protected the animals from both anaphylactic bronchoconstriction and blood pressure changes. When the antigen was inhaled by actively sensitized guinea-pigs, WEB 2086 inhibited the anaphylactic reaction without the administration of any additional antihistamine. These results suggest that PAF plays an important role in anaphylaxis. PAF may be (a) more important in passive than in active anaphylaxis and (b) may be involved during inhalative challenge in active anaphylaxis.

Aldosterone antagonists. 2. New 7 alpha-(acetylthio)-15,16-methylene spirolactones.

Some 15,16-methylene derivatives of the aldosterone antagonist spironolactone were synthesized with the purpose of increasing the antialdosterone potency and reducing the endocrinological effects of this standard compound. By introduction of a 1,2-double bond and a 15 beta,16 beta-methylene ring in the spironolactone molecule both goals were achieved. In animal studies mespirenone exhibited a threefold-greater antialdosterone potency and less than 10% of the antiandrogenic activity of spironolactone.

Bronchial and vascular effects of Paf in the rat isolated lung are completely blocked by WEB 2086, a novel specific Paf antagonist.

1 The effect of the platelet-activating factor (Paf) antagonist, WEB 2086, on Paf-induced increase of pulmonary artery perfusion pressure (Pp), bronchial inflation pressure (Pi) and wet-to-dry lung weight ratios (W/D) was investigated in the rat isolated lung. 2 Lungs were perfused with Krebs-Ringer solution (KRS) as controls or with KRS containing WEB 2086 (0.1, 1.0, 10.0 or 100 micrograms ml-1) and then injected with a bolus of 20 micrograms Paf. 3 A dose-related inhibition of the Paf-induced increase of Pp, Pi and W/D was observed, being almost maximal for the 10.0 micrograms ml-1 and complete for the 100 micrograms ml-1 doses of WEB 2086 when compared to controls. 4 It is concluded that WEB 2086 is a highly effective and specific Paf antagonist in the pulmonary vasculature and bronchial tract.

Pharmacological actions of WEB 2086, a new specific antagonist of platelet activating factor.

WEB 2086, a thieno-triazolodiazepine, is a potent and specific antagonist of platelet activating factor (PAF) in vitro and in vivo. This compound inhibits PAF-induced human platelet and neutrophil aggregation in vitro (IC50 = 0.17 and 0.36 microM, respectively) but has little or no effect on the action of other platelet aggregating agents. In comparison with kadsurenone, ketotifen or thiazinamium chloride, WEB 2086 was 26 to 200 times more potent in the PAF-induced platelet aggregation. In anesthetized guinea pigs, pretreatment with 0.1 to 2.0 mg/kg p.o. or 0.01 to 0.5 mg/kg i.v. of WEB 2086 inhibits dose-dependently the accumulation and aggregation of 111Indium labeled platelets, bronchoconstriction, systemic hypotension and also the lethal effect due to an i.v. PAF infusion [30 ng/(kg X min)] or intratracheal instillation of PAF (300 micrograms/kg). Under the same experimental conditions in guinea pigs, WEB 2086 given by inhalation achieved a similar anti-PAF activity. In anesthetized rats, the hypotension induced by an i.v. PAF infusion was also reversed (ED50 = 0.052 mg/kg i.v.). The increase in cutaneous vascular permeability due to intradermal PAF (25 ng/site) was inhibited dose-dependently by WEB 2086 (0.025-2 micrograms/site) in rats. Because of its structural relationship to triazolodiazepines, WEB 2086 was examined for anticonvulsant and sedative action. Up to doses of 300 and 800 mg/kg p.o., respectively, no effects were found. In conclusion, WEB 2086 is a potent and specific PAF antagonist with triazolodiazepine structure but without sedative activity.

Effects of WEB 2086, a novel antagonist of platelet activating factor, in active and passive anaphylaxis.

WEB 2086, a novel specific platelet activating factor (PAF) antagonist derived from triazolodiazepines, inhibited in a dose-related manner the PAF-dependent component of anaphylaxis as well as PAF-induced effects in mice and guinea pigs. In mice a lethal anaphylactic shock and a PAF-induced (100 micrograms/kg i.v.) death was inhibited by i.v. WEB 2086. The ED50 values were 13.6 and 0.37 mg/kg i.v., respectively. In actively sensitized guinea pigs, the anaphylactic lung reaction (bronchoconstriction), but not the corresponding hypotension, was prevented by oral (0.05-0.5 mg/kg) doses of WEB 2086. In contrast, in passively sensitized animals a dose-dependent inhibition of the pulmonary (bronchoconstriction) and blood pressure (hypotension) reaction due to anaphylaxis was achieved by i.v. WEB 2086. Similarly, oral (0.05-0.5 mg/kg) and i.v. (0.005-0.05 mg/kg) WEB 2086 inhibited PAF-induced reduction in respiratory flow (bronchoconstriction) and hypotension in guinea pigs. The ED50 values were 0.070 and 0.066 mg/kg p.o., and 0.017 and 0.015 mg/kg i.v., respectively. In conclusion, PAF seems to play a more major role in passive than in active anaphylaxis in guinea pigs. These results provide further evidence for an important role of PAF in anaphylaxis and support the hypothesis that PAF is involved in asthma and other allergic diseases.

Triazolodiazepines are potent antagonists of platelet activating factor (PAF) in vitro and in vivo.

The triazolodiazepines brotizolam, triazolam and alprazolam inhibited PAF-induced human platelet aggregation in vitro (IC50 = 0.54, 7.6 and 13.7 microM, respectively) but showed only a weak or no effect against other aggregating agents (ADP, adrenaline, collagen, serotonin, arachidonic acid). In comparison, flunitrazepam and diazepam, two diazepines without the triazole ring, showed IC50-values of 42 and 260 microM, respectively. Flunitrazepam does not possess the specificity shown by the other compounds. Brotizolam and triazolam also inhibited PAF-induced human neutrophil aggregation in vitro, with IC50-values 0.21 and 6.6 microM, respectively. In anesthetized guinea pigs, brotizolam (2.5 to 10 mg/kg p.o. or 0.1 to 0.5 mg/kg i.v.) or triazolam (20 to 100 mg/kg p.o.) inhibited dose-dependently the intrathoracic accumulation and aggregation of 111Indium labelled platelets induced by an i.v. infusion of PAF (30 ng/kg X min). Brotizolam at doses of 1 to 10 mg/kg p.o. and 0.1 to 0.5 mg/kg i.v. inhibited dose-dependently the reduction in tidal volume (bronchoconstriction), the systemic hypotension and the lethal effect due to i.v. PAF in guinea pigs. Triazolam inhibited these effects of PAF at doses of 50 to 200 mg/kg p.o. PAF-induced systemic hypotension in rats can be reversed by cumulative i.v. doses (0.05 to 1.0 mg/kg) of brotizolam. In conclusion, these results show that triazolodiazepines, like brotizolam and triazolam, are potent inhibitors of PAF-induced effects in vitro and in vivo.