Sphingosine kinases (SphK) 1 and 2 phosphorylate sphingosine to generate sphingosine-1-phosphate (S1P), a pluripotent lipophilic mediator implicated in a variety of cellular events. Here we show that the activity of β-site APP cleaving enzyme-1 (BACE1), the rate limiting enzyme for amyloid-β peptide (Aβ) production, is modulated by S1P in neurons. Treatment by SphK inhibitor, RNAi knockdown of SphK or overexpression of S1P degrading enzymes decreased BACE1 activity to reduce Aβ production. S1P specifically bound to full-length BACE1 and increased its proteolytic activity, suggesting that the cellular S1P directly modulates BACE1 activity. Notably, the relative activity of SphK2 was upregulated in the brains of patients with Alzheimer disease. The unique modulatory effect of cellular S1P on BACE1 activity is a novel potential therapeutic target for Alzheimer disease.

Amyloid-b peptide ending at the 42nd residue (Ab42) is implicated in the pathogenesis of Alzheimer's disease (AD). Small compounds that exhibit selective lowering effects on Ab42 production are termed c-secretase modulators (GSMs) and are deemed as promising therapeutic agents against AD, although the molecular target as well as the mechanism of action remains controversial. Here, we show that a phenylpiperidine-type compound GSM-1 directly targets the transmembrane domain (TMD) 1 of presenilin 1 (PS1) by photoaffinity labelling experiments combined with limited digestion. Binding of GSM-1 affected the structure of the initial substrate binding and the catalytic sites of the c-secretase, thereby decreasing production of Ab42, possibly by enhancing its conversion to Ab38. These data indicate an allosteric action of GSM-1 by directly binding to the TMD1 of PS1, pinpointing the target structure of the phenylpiperidine-type GSMs.

BackgroundAmyloid-β peptide ending at 42nd residue (Aβ42) is believed as a pathogenic peptide for Alzheimer disease. Although γ-secretase is a responsible protease to generate Aβ through a processive cleavage, the proteolytic mechanism of γ-secretase at molecular level is poorly understood.ResultsWe found that the transmembrane domain (TMD) 1 of presenilin (PS) 1, a catalytic subunit for the γ-secretase, as a key modulatory domain for Aβ42 production. Aβ42-lowering and -raising γ-secretase modulators (GSMs) directly targeted TMD1 of PS1 and affected its structure. A point mutation in TMD1 caused an aberrant secretion of longer Aβ species including Aβ45 that are the precursor of Aβ42. We further found that the helical surface of TMD1 is involved in the binding of Aβ45/48 and that the binding was altered by GSMs as well as TMD1 mutation.ConclusionsBinding between PS1 TMD1 and longer Aβ is critical for Aβ42 production.

Substrate-selective inhibition or modulation of the activity of γ-secretase, which is responsible for the generation of amyloid-β peptides, might be an effective strategy for prevention and treatment of Alzheimer's disease. We have shown that helical β-peptide foldamers are potent and specific inhibitors of γ-secretase. Here we report identification of target site of the foldamers by using a photoaffinity probe. The photoprobe directly and specifically labeled the N-terminal fragment of presenilin 1, in which the initial substrate docking site is predicted to be located. We also optimized the foldamer structure by preparing a variety of derivatives and obtained two highly potent foldamers by incorporation of a hydrophilic and neutral functional group into the parent structure. The class of side chain functional group and the position of incorporation were both important for γ-secretase-inhibitory activity. The substrate selectivity of the inhibitory activity was also quite sensitive to the class of side chain group incorporated.

Lycopodium alkaloids have attracted the attention of synthetic chemists because of their intriguing structural features and the remarkable bioactivities. [1] Among them is (À)lycoposerramine-S (1; Figure 1), which was isolated from Lycopodium serratum by Takayama and co-workers in 2002 [2] and has a unique structure including a highly fused tetracyclic skeleton with two nitrogen atoms and one quaternary carbon center. Although synthetic studies of (À)-lycoposerramine-S were reported by Elliott and co-workers, [3] its total synthesis has not been reported to date. Although the biological activity of this molecule has not yet been determined, we were interested in exploring this unique molecule and its derivatives for drug discovery. Herein, we report the first total synthesis of (À)-lycoposerramine-S (1). Key to the straightforward construction of the tetracyclic skeleton was the unexpected stereoselectivity in the intramolecular cycloaddition of an azomethine ylide.Our retrosynthesis is shown in Scheme 1. Formation of the nine-membered ring and the cyclopentane ring could be achieved by alkylation of a nosyl amide and radical cyclization, respectively. The requisite precursor 2 would be derived from the ketoester 3. The bicyclic system of 3 could in turn be constructed by an intramolecular cycloaddition of the azomethine ylide 4. [4] Our synthesis commenced with preparation of the precursor of the azomethine ylide (Scheme 2). Reaction of the alkyl iodide 5 [5] with an anion derived from the terminal alkyne 6 afforded the alkyne 7, which was subjected to hydrozirconation and subsequent addition of iodine to give the alkenyl iodide 8. [6] Halogen-lithium exchange of 8 generated the corresponding alkenyllithium species, which was reacted with the known lactone 9 [7] to afford 10. Subsequent oxidation of the hydroxy group in 10 furnished the ketoaldehyde 11, which was used for the cycloaddition of the azomethine ylide.Treatment of 11 with N-benzylglycine ethyl ester in refluxing toluene formed an azomethine ylide, which underwent cycloaddition to afford the product as a 4:3 mixture of two diastereomers which differed in the relative configuration at C15. This result indicated that the distant methyl group on C15 did not control the facial selectivity of the cycloaddition. Therefore we decided to employ chiral amino esters as reagents for the cycloaddition. Using (5S,6R)-5,6-diphenylmorpholin-2-one, [8] the cycloaddition proceeded stereoselectively, albeit in low yield. After intensive screening of chiral amino esters, we found that the morpholinone 12 gave the best selectivity and yield. [9] Thus, heating 11 with 12 in toluene furnished the adduct 13 in 86 % yield as a single isomer Figure 1. Structure of lycoposerramine-S. Scheme 1. Retrosynthesis. Scheme 2. Preparation of a precursor for the azomethine ylide. a) nBuLi, HCC(CH 2 ) 3 OTBS (6), THF/DMPU, À78 8C to RT, 87 %; b) [Cp 2 ZrCl 2 ], DIBAL, THF, 0 8C to RT; I 2 , À78 8C, 87 %; c) nBuLi, Et 2 O, À78 8C; 9, À78 8C, 66 %; d) TPAP, NMO, 4 M.S., CH 2 Cl 2 , RT, 7...