Pimavanserin, a 5HT2A receptor inverse agonist, rapidly suppresses Aß production and related pathology in a mouse model of Alzheimer's disease.

Amyloid-ß (Aß) peptide aggregation into soluble oligomers and insoluble plaques is a precipitating event in the pathogenesis of Alzheimer's disease (AD). Given that synaptic activity can regulate Aß generation, we postulated that 5HT2A -Rs may regulate Aß as well. We treated APP/PS1 transgenic mice with the selective 5HT2A inverse agonists M100907 or Pimavanserin systemically and measured brain interstitial fluid (ISF) Aß levels in real-time using in vivo microdialysis. Both compounds reduced ISF Aß levels by almost 50% within hours, but had no effect on Aß levels in 5HT2A -R knock-out mice. The Aß-lowering effects of Pimavanserin were blocked by extracellular-regulated kinase (ERK) and NMDA receptor inhibitors. Chronic administration of Pimavanserin by subcutaneous osmotic pump to aged APP/PS1 mice significantly reduced CSF Aß levels and Aß pathology and improved cognitive function in these mice. Pimavanserin is FDA-approved to treat Parkinson's disease psychosis, and also has been shown to reduce psychosis in a variety of other dementia subtypes including Alzheimer's disease. These data demonstrate that Pimavanserin may have disease-modifying benefits in addition to its efficacy against neuropsychiatric symptoms of Alzheimer's disease. Read the Editorial Highlight for this article on page 560.

Effect of escitalopram on Aß levels and plaque load in an Alzheimer mouse model.

BACKGROUND: Several neurotransmitter receptors activate signaling pathways that alter processing of the amyloid precursor protein (APP) into ß-amyloid (Aß). Serotonin signaling through a subset of serotonin receptors suppresses Aß generation. We proposed that escitalopram, the most specific selective serotonin reuptake inhibitor (SSRI) that inhibits the serotonin transporter SERT, would suppress Aß levels in mice. OBJECTIVES: We hypothesized that acute treatment with escitalopram would reduce Aß generation, which would be reflected chronically with a significant reduction in Aß plaque load. METHODS: We performed in vivo microdialysis and in vivo 2-photon imaging to assess changes in brain interstitial fluid (ISF) Aß and Aß plaque size over time, respectively, in the APP/presenilin 1 mouse model of Alzheimer disease treated with vehicle or escitalopram. We also chronically treated mice with escitalopram to determine the effect on plaques histologically. RESULTS: Escitalopram acutely reduced ISF Aß by 25% by increasing α-secretase cleavage of APP. Chronic administration of escitalopram significantly reduced plaque load by 28% and 34% at 2.5 and 5 mg/d, respectively. Escitalopram at 5 mg/kg did not remove existing plaques, but completely arrested individual plaque growth over time. CONCLUSIONS: Escitalopram significantly reduced Aß in mice, similar to previous findings in humans treated with acute dosing of an SSRI.

Rapid in vivo measurement of ß-amyloid reveals biphasic clearance kinetics in an Alzheimer's mouse model.

Findings from genetic, animal model, and human studies support the observation that accumulation of the ß-amyloid (Aß) peptide in the brain plays a central role in the pathogenic cascade of Alzheimer's disease (AD). Human studies suggest that one key factor leading to accumulation is a defect in brain Aß clearance. We have developed a novel microimmunoelectrode (MIE) to study the kinetics of Aß clearance using an electrochemical approach. This is the first study using MIEs in vivo to measure rapid changes in Aß levels in the brains of living mice. Extracellular, interstitial fluid (ISF) Aß levels were measured in the hippocampus of APP/PS1 mice. Baseline levels of Aß40 in the ISF are relatively stable and begin to decline within minutes of blocking Aß production with a γ-secretase inhibitor. Pretreatment with a P-glycoprotein inhibitor, which blocks blood-brain barrier transport of Aß, resulted in significant prolongation of Aß40 half-life, but only in the latter phase of Aß clearance from the ISF.

Highly efficient diastereoselective reduction of alpha-fluoroimines.

A highly selective reduction of alpha-fluoroimines to the corresponding beta-fluoroamines has been developed utilizing trichlorosilane as the reductant. The key aspect of this reaction is the ability of fluorine and nitrogen to activate organosilanes leading to high diastereoselectivity (>100:1) in the product distribution. This new method provides a new avenue for the diastereoselective synthesis of beta-fluorinated amines in good yields and selectivity.

Rate constants for peroxidation of polyunsaturated fatty acids and sterols in solution and in liposomes.

Rate constants for autoxidation propagation of several unsaturated lipids in benzene solution at 37 degrees C and in phosphatidylcholine liposomes were determined by a linoleate radical clock. This radical clock is based on competition between hydrogen atom abstraction by an intermediate peroxyl radical derived from linoleic acid that leads to a trans,cis-conjugated hydroxyoctadecadienoic product and beta-fragmentation of the same peroxyl that gives the trans,trans-product hydroxyoctadecadienoic acid. Rate constants determined by this approach in solution relative to linoleic acid (k(p) = 62 M(-1) s(-1)) were: arachidonic acid (k(p) = 197 +/- 13 M(-1) s(-1)), eicosapentaenoic acid (k(p) = 249 +/- 16 M(-1) s(-1)), docosahexaenoic acid (k(p) = 334 +/- 37 M(-1) s(-1)), cholesterol (k(p)= 11 +/- 2 M(-1) s(-1)), and 7-dehydrocholesterol (k(p)= 2260 +/- 40 M(-1) s(-1)). Free radical oxidations of multilamellar and unilamellar liposomes of various mixtures of glycerophosphatidylcholine molecular species were also carried out. In some experiments, cholesterol or 7-dehydrocholesterol was incorporated into the lipid mixture undergoing oxidation. A phosphatidylcholine bearing a linoleate ester at sn-2 was a component of each liposome peroxidation reaction and the ratio of trans,cis/trans,trans (t,c/t,t)-conjugated diene oxidation products formed from this phospholipid was determined for each oxidation reaction. This t,c/t,t-product ratio from linoleate was used to "clock" liposome constituents as hydrogen atom donors in the lipid bilayer. Application of this lipid bilayer radical clock gives relative autoxidation propagation rate constants of arachidonate (20:4), eicosapentaenoate (20:5), docosahexaenoate (22:6), and 7-dehydrocholesterol to be 115 +/- 7, 145 +/- 8, 172 +/- 13, and 832 +/- 86, respectively, a reactivity trend that parallels the one in solution. We also conclude from the liposome oxidations that linoleate peroxyl radicals at different positions on the eighteen-carbon chain (at C-9 and C-13) have different kinetic properties. This is in contrast to the results of solution oxidations of linoleate in which the C-9 and C-13 peroxyl radicals have similar reactivities. We suggest that peroxyl radical beta-scission depends on solvent polarity and the polarity of the local environment of peroxyl radicals in liposomal oxidations depends on the position of the peroxyl radical on the 18-carbon chain.

In vivo and in vitro lipid peroxidation of arachidonate esters: the effect of fish oil omega-3 lipids on product distribution.

The effect of lipid composition on the distribution of free radical oxidation products derived from arachidonic acid (20:4) esters has been studied in vitro and in vivo. Pro-inflammatory prostaglandin (PG) F2-like compounds, termed F2-isoprostanes (IsoPs), are produced in vivo and in vitro by the free radical-catalyzed peroxidation of arachidonic acid. Controlled free radical oxidation of mixtures of fatty acid esters in vitro showed that the formation of IsoPs from arachidonate is dramatically influenced by the presence of other fatty acid esters in the reaction mixture. Thus, three lipid mixtures containing the same arachidonate concentration but different amounts of other fatty esters (16:0; 18:1; 18:2; 20:5, and 22:6) were oxidized, and the product yields were determined by GC and LC/MS/MS analysis. The yield of F2-IsoP formed after 1 h of oxidation was 18% (based on arachidonate consumed) for mixtures containing arachidonate as the only oxidizable PUFA, but yields of these biologically active compounds dropped to 6% in polyunsaturated fatty acid (PUFA) mixtures typical of those found in tissues of fish oil-fed animals. F2-IsoP levels were also monitored in the livers of mice on diets supplemented with eicosapentaenoic acid (C20:5 omega-3; EPA), the PUFA most abundant in fish oil. While the level of arachidonic acid present in livers was not significantly different from that in control animals, levels of IsoPs in the liver were reduced in the EPA-fed mice compared to those in controls under conditions of oxidative stress (60 +/- 25% reduction, n = 5) or at baseline (48 +/- 14% reduction, n = 5). These results suggest that dietary omega-3 PUFAs may influence the formation of bio-active peroxidation products derived from omega-6 PUFAs by channeling the free radical pathway away from the F2-IsoPs.

Disposition of 9-nitrocamptothecin and its 9-aminocamptothecin metabolite in relation to ABC transporter genotypes.

PURPOSE: The source of the pharmacokinetic variability of 9-nitrocamptothecin (9NC) and its 9-aminocamptothecin (9AC) metabolite is unknown. ATP-binding cassette (ABC) transporters have been reported to modulate camptothecin analogues, are associated with camptothecin resistance, and might also affect 9NC and 9AC pharmacokinetics. The aim of this study was to evaluate the functional consequence of known single nucleotide polymorphisms in the transporter genes ABCB1, ABCC2, and ABCG2 on the pharmacokinetic disposition of 9NC and 9AC. EXPERIMENTAL DESIGN: Pharmacokinetic and genotyping studies were performed in 55 patients as part of two phase I studies of 9NC in patients with refractory solid tumors, a phase II study of 9NC in patients with advanced colon cancer, and a study evaluating the disposition of 9NC after administration of a single dose under fasting conditions. DNA was isolated from plasma and analyzed for variants in ABCB1, ABCC2, and ABCG2 genes. The ABCB1 1236C>T (n = 43), ABCB1 2677G>T/A (n = 43), ABCB1 3435C>T (n = 43), ABCC2 3972C>T (n = 39), and ABCG2 421C>A (n = 42) variants were analyzed using Pyrosequencing. RESULTS: The ABCG2 421C>A genotype significantly affected the pharmacokinetics of 9AC. The mean 9AC lactone AUC/dose for wild-type (n = 25) and heterozygous (n = 2) patients were 14.3 ng/mL x h and 51.1 ng/mL x h, respectively (P = 0.032). The mean +/- SD 9AC total AUC/dose for wild-type (n = 39) and heterozygous (n = 3) patients were 91.9 +/- 78.3 ng/mL x h and 129.0 +/- 90.5 ng/mL x h, respectively (P = 0.40). 9NC and 9AC disposition were not significantly influenced by variants in ABCB1, ABCC2, and ABCG2, and ABCB1 and ABCC2, respectively (P > 0.05). CONCLUSION: These findings suggest that inter-individual variability in 9AC disposition, but not 9NC, may be influenced, in part, by ABCG2 genotype. In contrast, there was no evidence for a relationship between ABCG2 and the disposition of 9NC, or for relationships between ABCB1 and ABCC2 genotypes and the disposition of 9NC or 9AC.

Chelation-controlled diastereoselective reduction of alpha-fluoroketones.

The effect of Ti-based Lewis acids on the reduction of alpha-fluoropropiophenone was examined to determine whether chelation control could be used to direct the diastereoselectivity of conversion to an alpha-fluoro alcohol. Pretreatment of alpha-fluoropropiophenone with TiCl4 followed by reduction with LiBH4 in diethyl ether or methylene chloride provided the syn diastereomer predominantly, while use of Ti(OiPr)4 under identical conditions provided the anti diastereomer as the major product. The products are consistent with a chelation-controlled mechanistic pathway in the former reduction and a nonchelation pathway in the latter case. Detailed 1H, 13C, and 19F NMR studies were consistent with chelation between TiCl4 and alpha-fluoropropiophenone under the reaction conditions utilized in this study. Reduction of other alpha-fluoroketones in the presence of TiCl4 also provided a high degree of diastereoselectivity in the conversion to alpha-fluoro alcohols, showing the generality of this approach.

Reduction of beta-hydroxyketones by SmI2/H2O/Et3N.

Reduction of a series of beta-hydroxyketones by SmI2/H2O/Et3N provided 1,3-diols in quantitative yields. The reactions were exceedingly clean with no byproduct formation, negating the need for further purification. Most reactions provided moderate to excellent diastereoselectivity with syn-diols as the major isomer in most instances.

Solvent-dependent diastereoselectivities in reductions of beta-hydroxyketones by SmI2.

The reductions of a series of beta-hydroxyketones by SmI(2) were examined in THF, DME, and CH(3)CN using methanol as a proton source. Reductions in THF and DME typically lead to the syn diastereomer with DME providing higher diastereoselectivities. Reductions in CH(3)CN provided the anti diastereomer predominantly. This study reveals that solvation plays an important role in substrate reduction by SmI(2). [reaction: see text]