Gibberellins are phytohormones first discovered at the University of Tokyo in 1938. When I was young, I worked for 9 years since 1959 to synthesize gibberellin A_4 in Prof. M. Matsui's laboratory at the University of Tokyo. When I finished the work in 1968, a'famous microbiologist Prof. K. Arima said to me, "Congratulations, Dr. Mori on the completion of the gibberellin synthesis. But you spent 9 years of your life to do it. Don't forget that the fungus Gibberella fujikuroi makes the gibberellins within a couple of days." This criticism made me to think that we chemists can be respected by biologists only when we synthesize those compounds which are difficult to be prepared by biological systems. Accordingly, I started my pheromone synthesis in 1973. As a tool to prepare enantiomerically pure pheromones, I employed biocatalysis for the preparation of chiral and nonracemic building blocks. The latest pheromone work of ours was the synthesis and determination of the absolute configuration of the male aggregation pheromone of the stink bug, Erysarcoris lewisi. The natural pheromone was shown to be (2Z, 6R, 1'S, 5'S)-2-methyl-6-(4'-methylenebicyclo[3.1.0]hexyl) hept 2-en-l-ol by employing both biocatalytic and chemical methods. In the coming decades, clarification in the molecular level of the interaction between a small ligand molecule and a large receptor protein will be one of the most important subjects in natural products chemistry. In our immunological studies to find out medicinally useful ligands for CD1d protein, we inspected carefully the published X-ray crystal structure of human CD1d-ligand (KRN7000)-T cell receptor complex. This effort resulted in the invention of new ligands such as RCAI-56 and RCAI-61 with modified α-D-galactosyl parts. So-called "structure-based design of ligands" was successful in our case to make unusually potent ligands to induce interferon-γ production by natural killer T cells.

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In the first half of the presentation, we attempt to reproduce the talk "Synthetic Study on Tetrodotoxin and Related Compounds" given at the Thirteenth Symposium on the Chemistry of Natural Products (Nagoya, 1971). Using the synthesis of the protected tetrodamine as an example, we discuss stereochemistry control based on the convex-concave concept. In the second half of the presentation, we focus on stereochemistry control via the concept of acyclic stereocontrol. For this illustration, we use the synthesis of the C1-C7 moiety of monensin as an example.

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Matatabi (Actinidia polygamy Miq., Actinidiaceae) is widely distributed in this country. It is well known from early times that the leaf, the fruit, and the root of this plant show special biological activity in Felidae animals. Although significant chemical studies were carried out, the isolation and structure elucidation of the active principles were not reported until 1959. We isolated two active principles from the methanol extract of the leaf and the fruit of matatabi, a lactone C_<10>H_<16>O_2 named matatabilactone (1), and a base C_<10>H_<13>N named actinidine (2), in 0.0035 and 0.12% yield, respectively. Herein the structure study and the synthesis of 1 and 2 will be discussed.

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We have been involved in design of asymmetric catalysts. development of catalytic asymmetric reactions and their application of the synthesis of bioactive molecules. In this presentation, development to catalytic asymmetric carbon-carbon bond-forming reactions and their application to the synthesis of natural products are mainly discussed. 1) We developed a very practical catalytic asymmetric Michael reaction using enones and malonates. Using this reaction as a key step, we achieved a catalytic asymmetric synthesis of strychnine 9. 2) We also developed an efficient catalytic asymmetric cyanosilylation of ketones. This reaction made possible a catalytic asymmetric synthesis of camptothecin 15 as well as fostriecin 33. In addition to these catalytic asymmetric syntheses, we also achieved catalytic asymmetric syntheses of epothilones 34, lactacystin 35, aeruginosin 298-A 36 and cylindricine C 37.

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In cytotoxic conditions such as hypoxia, nutrient deprivation and low pH, unfolded proteins accumulate in cells, leading to ER stress. Poorly vascularized solid-tumor cells are considered to adapt to ER stress by activating a transcription factor XBP1, resulting in tumor survival. Therefore, inhibitors of XBP1 activation would be a new type of anticancer drugs. To screen for inhibitors of ER stress-induced XBP1 activation, we established a screening system (Fig. 1), and isolated novel triene-ansamycin group compounds trierixin and quinotrierixin from culture broths of Streptomyces sp. AC654 and Streptomyces sp. PAE37, respectively (Fig. 3). The planar structures of trierixin and quinotrierixin were elucidated on the basis of the spectroscopical properties (Fig. 2). Trierixin inhibited ER stress-induced XBP1 activation and cell growth of HeLa cells with IC_<50> of 28nM and 10nM, respectively. Quinotrierixin also showed both inhibitory activities with the same dose range. Therefore, we proposed a hypothesis that triene-ansamycin group compounds inhibited tumor cell growth via inhibition of XBP1 activation. To confirm this possibility, we isolated six additional triene-ansamycin group compounds (including three novel compounds) from the culture broth of Streptomyces sp. PAE37 by referring to the characteristic UV spectra (λ_<max> 260, 270 and 280nm), and prepared four derivatives by chemical reactions on the natural products (Fig. 3, Fig. 4). Using these compounds, we performed SAR study of triene-ansamycin group compounds. As a result, there was high correlation between inhibitory effects of triene-ansamycin group compounds against XBP1 activation and those against tumor cell growth (Fig. 5).

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Dinoflagellates are widely known as the rich source of biologically active and structural unique secondary metabolites. Among them, large polyols and polyethers composed of a long carbon backbone that is highly functionalized by oxygen, as we say "super-carbon-chain compounds (SCCs)", are some of the most unusual compounds. In our quest for SCCs from marine dinoflagellates, we isolated a novel spiroketal compound, symbiospirol A, from Symbiodinium sp. that was the same strain which produced symbiodinolide. Its planar structure and partial relative stereochemistries were elucidated based on 1D and 2D-NMR and degradation reaction. Symbiospirol A was shown to inhibit PKC activation induced by phosphatidylserine. Symbiodinolide is the typical SCC with a molecular weight of 2,860, which we isolated from the dinoflagellate Symbiodinium sp. The chemical degradation reaction using the 2nd-generation Hoveyda-Grubbs catalyst was employed for the stereochemical assignment at 62-membered macrolactone of symbiodinolide. The absolute configurations at several chiral centers were determined by spectroscopic analysis of MTPA diesters. Further chemical degradation studies using the 2nd-generation Grubbs' catalyst have performed new reaction. In addition, the complete stereochemical assignment of symbiodinolide is in progress. In our continuing search for novel bioactive metabolites from symbiotic marine dinoflagellates, we found that the water layer from the symbiotic dinoflagellate with jellyfish Mastigias papua displayed considerable activity against the expression of vascular cell adhesion molecule-1 (VCAM-1) in human umbilical vein endothelial cells (HUVEC). Further purification of the water layer guided by the bioassay and mass measurement led to the isolation of an unknown bioactive SCC, which we named symbiopolyol, possessing the molecular weight of 1,220. Studies on the isolation, structure elucidation, and biological activities of symbiopolyol will be described.

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Malaria caused by parasites of the genus Plasmodium is one of the leading infectious diseases in many tropical and some of temperate regions. During our studies on new lead substances against malaria from medicinal plants, cassiarins A (1) and B (2), novel aromatic alkaloids with an unprecedented tricyclic skeleton and potent antiplasmodial activity, and chrobisiamone A (3), 5-acetonyl-7-hydroxy-2- methylchromone (4), and anhydrobarakol (5) have been isolated from the leaves of Cassia siamea (Leguminosae), which have been widely used in traditional medicine, particularly for the treatment of periodic fever and malaria. Cassiarin A (1) might be derived through an imine intermediate of 5-acetonyl-7-hydroxy-2-methylchromone followed by cyclization with ketone of chromone. Treatment of 5-acetonyl-7-hydroxy-2-methylchromone (4) with ammonium acetate as the nitrogen source caused ring cyclization giving cassiarin A (1). Cassiarin A (1) showed promising in vitro antiplasmodial activity against Plasmodium falciparum (IC_<50> 0.005ug/ml). The first total synthesis of cassiarin A (1) was achieved via sequential alkynylation of arenes with Sonogashira coupling and 6-endo-dig-cyclization of phenolic oxygens to the resulting alkynes. This synthesis afforded cassiarin A (1) in 51% overall yield in seven steps from the readily accessible known starting material. Chrobisiamone A (3) was also synthesized from 5-acetonyl-7-hydroxy-2-methylchromone (4) by Michael addition of the chromone carbanion of C-14 to C-2' of a second chromone.

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Plants of the Kopsia genus (Apocynaceae) ranging from evergreen trees to shrubs are well known as fertile sources of structurally diversified indole alkaloids. There are 23 species belonging to this genus, which has a widespread distribution from southern China and Burma to northern Australia and Vanuatu and is most diverse in Peninsular Malaysia and Borneo. Among them, Kopsia officinalis Tsiang et P.T. Li (this species was amended to Kopsia arborea in the view of Dr. Middleton's latest revision) indigenous to Yunnan province in southwest of China has been reported for medicinal uses for the treatment of rheumatoid arthritis, dropsy, and tonsillitis. As a part of our ongoing program to search for novel and bioactive indole alkaloids, phytochemical research on this plant has led to the isolation of more than 40 alkaloids. Of these, several, such as arboloscine (10) and mersicarpine (11) are interesting for incorporating intriguing molecular skeletons. In this symposium, we present the structure elucidation of six unusual indole alkaloids, i.e.: the novel dimeric indole alkaloid kopsiyunnanine A (1), the novel rearranged oxindole alkaloid kopsiyunnanine B (2), three novel rhazinilam analogues kopsiyunnanine Cl (4), kopsiyunnanine C2 (5) and kopsiyunnanine C3 (6), and an unusual quebrachamine derivative kopsiyunnanine D (8) (Figure 1). In addition, one another novel alkaloid, named kopsiyunnanine E (9), which possesses a particular notable pentacyclic carbon skeleton incorporating a lactone ring, will be also introduced.

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The juvenile hormone (JH) is one of the key insect hormones. Here we describe the novel structure of the heteropteran JH, named JHSB_3. We hypothesized four candidate structures 1-4 as a result of the following experimental results: 1) The in vitro product of the corpus allatum (CA) of a stink bug, Plautia crossota stali, analyzed by gas chromatography/chemical ionization-mass spectrometry (GC/MS) showed a molecular mass ion peak at m/z 300 [M+NH_4]^+, 2) The high resolution fast atom bombardment mass spectrometry of the same sample gave the exact mass ion peak [M+H]^+ of 283.1885 suggesting a molecular formula, C_<16>H_<27>O_4, identical to that of JHB_3, 3) Enhanced JH production by CA in vitro by the addition of excess E, E-farnesol to the medium, indicated an E,E-farnesol skeleton for the heteropteran JH. The four candidates were stereoselectively synthesized in an enantiomeric pure form via Katsuki-Sharpless epoxidation and Sharpless dihydroxylation reactions. The juvenilizing effect of the bisepoxides 1-4 on metamorphosis of the bug showed that 1 and 2 were more potent than 3 and 4. When the CA product was subjected to a chiral GC/MS, the CA product and 2 had the same retention time. Thus, the structure of the native JH in P.c. stali was determined as 2.

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Sesquiterpenoid quinines and hydroquinones are representative marine natural products with unique structures and interesting biological activities. In our continuous search for bioactive compounds having unique structures from Okinawan marine sponges, we have isolated new sesquiterpenoid quinones, nakijiquinones E〜I (1〜5) and metachromins L-Q (6〜11), from three collections of Okinawan marine sponges. The structures of 1〜5 were elucidated on the basis of 2D NMR data, while those of 6〜11 were established by chemical correlations. Furthermore, biological activities for 1〜11 and synthetic metachromin analogs 15〜26 were examined. Nakijiquinones E (1) and F (2) were the first dimeric sesquiterpenoid quinones possessing a 3-aminobenzoate moiety, while nakijiquinones G〜I (3〜5) were new sesquiterpenoid quinones having a histamine unit, an agmatine unit, and 3-(methylsulfinyl) propan-1-amino group, respectively. Metachromins L〜Q (6〜11) were new sesquiterpenoid quinones containing a different amino group derived from amino acids. Most of nakijiquinones E〜I (1〜5), metachromins L〜Q (6〜11), and synthetic metachromin analogs (15〜26) showed modest cytotoxicity against murine leukaemia P388 and L1210 cells, and human epidermoid carcinoma KB cells (IC_<50>, 1〜10μg/mL). Inhibitory activity of 15〜26 against several tyrosine kinases are currently investigated.

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Ladder-shaped polyether (LSP) toxins represented by brevetoxins are thought to bind to transmembrane (TM) proteins. To elucidate the interactions of LSPs with TM proteins, we have synthesized artificial ladder-shaped polyethers (ALPs) containing 6/7/6/6 tetracyclic, 6/7/6/6/7/6/6 heptacyclic, and 6/7/6/6/7/6/6/7/6/6 decacyclic system, based on the convergent method via α-cyano ethers. The ALPs possessing the simple iterative structure with different number of rings would be useful for structure-activity relationship studies on the molecular length. Two series of ALPs were prepared, (i) both sides were functionalized as diols (A series), and (ii) one side remained as diol and the other side was protected as benzyl ethers (B series), and the interaction of these ALPs with TM proteins was evaluated, based on dissociation of glycophorin A (GpA) dimers into monomers. The heptacyclic ether (ALP7B) elicited the most potent activity in the presence of 2% SDS buffer, whereas the decacyclic ether (ALP10A) exhibited intriguing phenomenon to induce precipitation of GpA in a dose-dependent manner, under the low concentration of SDS (0.03%). The different activities among the ALPs can be accounted for by the concept of 'hydrophobic matching' i.e. lengths of the hydrophobic region including the side chains of ALP7B and ALP10A are ca. 25Å, which match the lengths of the hydrophobic region of α-helical TM proteins, as well as the hydrophobic thickness of lipid bilayer membranes.

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2-Deoxyoligosaccharides are structural units in many biological active natural products and play important roles in their biological activities. The synthesis of various deoxyoligosaccharides derivatives would be attractive and effective way for the synthesis of new biologically active small molecules. However, construction of 2-deoxyoligosaccharide is very difficult because the lack of electron-withdrawing substituent at C-2 makes the glycosidic linkage unstable under acidic condition. In this symposium, we describe effective α- and β-selective glycosylation for the synthesis of deoxyoligosaccharides and an application to the synthesis of a versipelostatin sugar-part. First, we describe β-selective glycosidation of 2-deoxyosugars via oxidative activation of glycosyl imidates with iodine and a catalytic amount of Et_3SiH. Using this method, β(1,4) tetraolivoside was prepared by a simple glycosidation and deprotection procedure. Next we report α-selective glycosylation based on the oxidative activation of the glycosyl imdiates. This method enables the direct and stereoselective synthesis of deoxyoligosaccharide involving both α- and β-glycosidic linkages. Finally, we describe the synthesis of 7-reduced-versipelostatin (VST) derivatives from a protected reduced aglycon. We also report the biological effect of the deoxusugar part on the biological activity, and structure revision of the deoxysugar part of VST.

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The inhibitors of Ca^<2+>-ATPases exemplified by artemisinin and transtaganolides possess highly oxygenated sesquiterpene frameworks. We focused our attention to the tricyclic system as a common structural motif of the Ca^<2+>-ATPase inhibitors aside from the differences in their stereochemical relationships of the ring junctions and oxygenated functionalities incorporated into the cyclic skeletons. To access varied molecular architectures related to the biologically active sesquiterpenes, we envisioned stereochemical diversification of the consecutive ring-junctions into three types involving trans-cis, cis-cis and trans-trans fused skeletons. Tandem ring-closing olefin metathesis reactions of three dienynes possessing distinct stereochemical relationships (anti-syn, syn-syn and anti-anti) would furnish the sets of tricyclic skeletons. With an aim of rapid and stereo-controlled assembly of dienynes into the three consecutive carbon centers on cyclohexane rings, six-membered oxonitrile 3 was designed as a versatile scaffold for installing the dienynes on the three sites (A-C). Based on this strategy, we have developed an expeditious and programmable synthetic process that controls not only the stereochemical relationships of ring-junctions but also the modes of cyclization, which entails formation of six types of the tricyclic skeletons in a systematic fashion. Furthermore, isomerization of internal dienes and subsequent photo-oxidation were demonstrated to construct a collection of artemisinin analogs and also increase three-dimensional structural diversity of the small molecule library. Preliminary results on the screening and SAR studies of anti-parasitic substances will be presented.

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Chitin, the second most abundant polysaccharide in nature, is a constituent of fungal cell walls, the exoskeletons of crustaceans and insects, and the microfilarial sheaths of parasitic nematodes. Accumulation of chitin by organisms is modulated by chitin synthase-mediated biosynthesis and by chitinases-mediated hydrolytic degradation. Consequently, chitinases have been expected to be specific target for antifungal, insecticidal and antiparasitic agents. Argifin (1) represents a family of peptide products, of natural microorganic origin, isolated from the cultured broth of a fungal strain, Gliocladium sp. FTD-0668, by our research group and characterized as a novel chitinase inhibitor, with strong inhibitory activity against Serratia marcescens chitinases (SmChi). Recently, X-ray crystallographic analysis of chitinases-argifin complexes has revealed that there are strong hydrogen-bond interactions between the N^ω-methylcarbamoyl-L-arginine moiety and the motif of the hydrolytic pocket of bacterial chitinases, resulting in nanomolar to micromolar range inhibition. We report the efficient solid phase total synthesis of arigifin, the molecular design of novel chitinase inhibitors derived from its structure, and application of in situ click chemistry, using an azide-bearing simplified analogue, to identify more potent chitinase inhibitors.

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Neobivsanins A (1) and B (2), novel tricyclic bistetrahydrofurans isolated from Viburnum awabuki, have been proved to enhance neurite-outgrowth of NGF-mediated PC12 cells by our group, and thus these are regarded as a seed compound to develop therapeutic agent for treatment of neurodegenerative disease such as Alzheimer disease. Therefore, we have launched synthetic study of neovibsanin A (1) and B (2), and its derivatives. Our synthetic plan is shown in Scheme 1. Tricyclic lactone 3 would be set up as the precursor of 1 and 2 or its derivatives and expected to be formed via succesive oxy-Michael addition and lactonization of 4. In order to construct three consecutive stereogenic centers of 4, the stereoselective alkylation and the regioselective intramolecular Diels-Alder reaction of tetraene 7 were envisaged. The intramolecular Diels-Alder reaction of 7, which was synthesized form geraniol in 3 steps, proceeded by heating to 200℃ in DMI for 3h to give lactone 6 in good yield. After conversion of lactone 6 into 11 via Baylis-Hillman reaction using triethylphosphine as a catalyst, alkylation of the resulted 11 with lithium ethyl propiolate afforded a mixture of 12a and 12b (1.3:1). By the treatment of reduced product of 12b with TBAF, successive oxy-Micael addition and lactoniaztion took place cleanly, giving rise to tricyclic lactone 15 in good yield. After deprotection of MOM group and TBS protection of the alcohol, the resulting 16 was treated with Tebbe reagent, and followed by a solvolysis of the enol ether with PPTS in methanol, leading acetal 17. TBS ether of 17 was removed with TBAF, and the resulting alcohol was oxidized with SO_3-pyridine, giving rise to unstable aldehyde 18 in 90% yield. Finally, a trapping lithium enolate prepared by treatment of 18 with KHMDS with 3,3-dimethylacryloyl chloride led to the complete syntheses of (±)-4, 5-bis-epi-neovibsanin A (19) and (±)-(8Z)-4, 5-bis-epi-neovibsanin A (20) in 90% yield (19/20 1:1). Furthermore, our synthetic procedure could be successfully applied to the preparation of a number of derivatives (31-39), and then the studies of structure activity relationships were carried out As results, it was revealed that 10μM of (±)-4, 5-bis-epi-neovibsanin A (19) promoted effectively neurite-outgrowth of NGF-mediated PC12 cells, comparable with neovibsanin B (2). Compounds 29-39 showed also activity, whereas lactone 29, bicyclic 31, and 37 and 38, which have longer side chains than propyl group at the acetal position, were found to be inactive.

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Bacilosarcins A (1) & B (2), potent plant seed germination inhibitors comparable to herbimycin A, were recently isolated from the culture broth of Bacillus subtilis TP-B0611 by Igarashi and co-workers. Their unique structures featuring a 3,4-dihydroisocoumarin ring system linked to unprecedented heterocyclic frameworks as well as the intriguing biological activity prompted us to embark on their total synthesis. In this symposium, we report the first enantioselective total synthesis of 1, 2, and a structurally-related cytotoxin (Sg17-1-4) Starting from a known ester (11), the right segment (14) was synthesized in 6 steps through Cordova's asymmetric epoxidation, Z-selective Horner-Emmons olefination and TEA-mediated hydroxylactonization. The segment (14) was condensed with the left segment (25) prepared via the Sharpless kinetic resolution, and deprotected to give 3. Selective hydrolysis of its 5-membered lactone ring gave AI-77-B (26), an antiulcerogenic substance. The total synthesis of 1 was accomplished by selective ammonolysis of the 5-membered lactone moiety of 3 followed by condensation with diacetyl. N-Alkylation of 3 was successfully achieved by treatment of 3 with acetoin in the presence of MgSO_4, and the product 30 obtained as an epimeric mixture was converted into 2・HCl (formed as a single diastereomer) or Sg17-1-4 (34, a cytotoxin) by ammonolysis or hydrolysis, respectively. We will discuss a plausible mechanism for the stereoconvergent formation of 2 from 30. Thus, the first total synthesis of 1 and 2 has been accomplished in 10 steps from 11.

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Cytotrienin A (1) is a microbial antitumor secondary metabolite, isolated from a fermentation broth of Streptomyces sp. RK95-74 from soil. It possesses a (E, E, E)-triene within a 21-membered cyclic lactam which also contains four chiral centers, common structural features of the ansamycin-class of natural products, including the mycotrienins (or ansatrienins), trienomycins, thiazinotrienomycins, and trierixin. Cytotrienin A, with its unusual aminocyclopropane carboxylic acid side chain, exhibits potent apoptosis-inducing activity on HL-60 cells, with an ED_<50> value of 7.7nM. To facilitate elucidation of its mechanism of action, the development of a method for its total synthesis and derivatization is highly desirable. The first asymmetric total synthesis of (+)-cytotrienin A has been achieved, and its absolute stereochemistry has been determined. There are several noteworthy features to this total synthesis: A practical diastereo- and enantio-selective aldol reaction using novel catalyst 10 under solvent-free conditions, highly diastereoselective construction of the three contiguous chiral centers, a deoxygenation reaction without positional or E/Z isomerization (from 22 to 23), desulfonylation using NaBH_4 (from 30 to 31), control of the absolute configuration at C_<13> by proline-mediated α-aminoxylation (from 32 to 33), and RCM for the formation of the 21-membered macrolactam (from 43 to 44).

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To investigate the function of oligosaccharide on protein, we have examined synthesis of homogeneous glycoproteins by use of combined chemical and E. coli expression system. In our strategy, a homogeneous glycopeptide thioester is synthesized by chemical method, while normal peptide chain is prepared by E. coli expression system. Then coupling between this glycopeptide thioester and a large polypeptide chain prepared by E. coli expression system, can be performed by Native Chemical Ligation (NCL). In this study, we selected erythropoietin (EPO) analogue, which is consist of 166-amino acid residues and display two N-linked complex type disialyloligosaccharide chains at the 24 and 30 positions on the polypeptide chain. EPO analogue was divided into two segments, EPO segment (1-32) 5 and EPO segment (33-166) 7. In order to synthesize glycopeptides thioester segment 5 having two cysteine residues at the 24 and 30 positions where would be incorporated with oligosaccharides, protected peptide chain (1-32) was prepared by Fmoc SPPS and then its C-terminal carboxylic acid was converted into ethyl-thioester to afford peptide thioester 3. A complex-type disialyloligosaccharyl-bromoacetamide was employed to modify the two free cysteine residues. This coupling reaction afforded the desired neoglycopeptide thioester 5 in good yield. The EPO polypeptide chain, comprising residues 33-166 (7) was purified from E. coli as a polyhistidine-tagged fusion protein with a methionine residue inserted between the tag and the EPO segment (33-166) 6. After isolation of the fusion protein 6 by nickel-affinity column, Histag-methionine segment was removed by CNBr treatment to afford desired EPO segment having cysteine residue at the N-terminus. Finally, we examined coupling reaction between these two segments by NCL and this reaction afforded whole EPO analogue in moderate yield. Finally, we examined folding experiment in order to have bioactive EPO analogue. In this presentation, we would like to discuss synthesis of EPO analogue 10, its folding experiments and bioactivity.

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In this conference, we report our efforts on the "library-directed" solid-phase synthesis of N-linked glycans, for the first step toward investigating their biological functions. We have efficiently prepared the Manβ (1-4) GlcNTroc fragment C by developing the new dual Lewis acid/cation trap reagent, TMSB(C_6F_5)_4. On the other hand, a highly α-selective sialylation of sialic acid N-phenyltrifluoroacetimidate was achieved by introducing the C-5 N-phthalylimide or the azide groups on the donor; the "fixed dipole moment effects" of these C-5 functional groups was proposed to explain the high reactivity and excellent α-selectivity. The microfluidic system was then applied to the present α-sialylation, which is amenable to large-scale synthesis of the Neu5Acα(2-6)Gal fragment D. The mono- and disaccharide fragments A-D thus prepared were efficiently glycosylated on the JandaJel^<TM> resin, by the use of the N-phenyltrifluoroacetimidate as the leaving group. In order to overcome the retarded reactivity on the solid-supports, especially for the glycosylation on the elongated sugar chain, the "reagent concentration effects" by using the fluorous solvent was successfully applied to the efficient glycosylation on the solid supports. A large-scale reductive cleavage of the benzylidene acetal group under the microfluidic conditions was also developed during the current N-glycan synthesis. The orchestration of these methods led to the efficient synthesis of the complex-type N-glycans, i.e., the core pentasaccharide as well as the octasaccharide, which contains the non-reducing end sialic acid, for the first time.

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Spiruchostatins A (1) and B (2), isolated from a culture broth Pseudomonas sp. by Shinya and co-workers in 2001, exhibit potent histone deacetylase (HDAC) inhibitor. A closely related depsipeptide FK228 (3), a powerful HDAC inhibitor, was previously isolated from Chromobacterium violaceum by Fujisawa Pharmaceutical Co. Ltd.. Therefore, these natural products are expected to be promising candidates for novel molecular-targeted anti cancer agents. The attractive biological activities and unique structural features prompted us to undertake the total syntheses of 1, 2 and 3. We envisioned that spiruchostatin A (1) would be elaborated through the disulfide bond formation of 4. Macrocyclic compound 4 can be synthesized through macrolactonization of seco-acid 5. The synthesis began with the preparation of intermediate 20, the substrate for the key macrolactonization. Coupling product 20 was efficiently prepared from 6 with 7 using a combination of HATU and HOAt at low temperature. After deprotection of the PMB and allyl groups, seco-acid 5 was obtained. The critical macrolactonization of 5 was best achieved by employing the Shiina protocol (MNBA and DMAP in CH_2Cl_2 at rt). Finally, disulfide bond formation by exposure to iodine followed by deprotection of TBS group completed the total synthesis of 1. Total synthesis of spiruchostatin B (2) has not yet been mentioned in the literature, and the stereochemistry at C5" of 2 has not clearly assigned. Therefore, we attempted determination of C5" configuration in 2; segment (5S)-22 would be produced through condensation of D-allo-isoleucine 21a with D-cysteine derivative. Finally, synthetic product 2 was prepared in a similar manner as 1 (22+7→→23→→2), and C5" stereochemistry of 2 was determined to be (S)-configuration. We envisioned that FK228 (3) would be constructed efficiently from the coupling reaction of two segments 24 with 25 followed by macrolactonization and disulfide bond formation. Initial attempts to achieve the pivotal macrolactonization of seco acid 15-epi-26 under usual Shiina conditions failed. After screening several reaction conditions, we solved this problem using Mitsunobu conditions, producing the desired 16-membered depsipeptide 27(26→27). Finally disulfide bond formation produced the target 3, whose spectroscopic properties were identical with those of natural product 3. In conclusion, we have accomplished total synthesis of spiruchostatin A (1), B (2), and FK228 (3) in a convergent manner. The C5" stereochemistry of 2 was determined to be (S)-configuration by the present synthesis.

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Polytheonamide B (1) is a potent cytotoxic natural product isolated from marine sponge and is by far the largest non-ribosomal peptide known to date. The 48 amino acid residues including a variety of non-proteinogenic amino acids have the sequence of alternating D- and L-chirality. These structural features permit the formation of a stable β-strand-type structure with all of the side chains on one side, thereby encouraging the formation of a helix (6.5 residues per turn). In the biological setting, cations can be transported across lipid bilayer of cells through the pore to exert its bioactivity. The unique and synthetically challenging structure of 1 together with the potent cytotoxicity motivated us to launch a project for their chemical construction. Here we report the first total synthesis of polytheonamide B The containing non-proteinogenic amino acids of 1 were prepared by multi-step syntheses from readily available starting materials. Then, four peptide fragments {Ncap-[1-11]-OH (A), Fmoc-[12-25]-OH (B), Fmoc-[26-32]-OH (C), Fmoc-[33-48]-OH} (D) were synthesized on solid phase under the carefully optimized conditions. Fragments A, B and C were derivatized to the corresponding thioester, and the union of these fragments was performed using AgNO_3 in the presence of HOOBt in a stepwise fashion, delivering the protected polytheonamide B. The last global deprotection was realized under the acidic conditions to provide polytheonamide B (1).

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Kedarcidin, isolated from the actinomycete strain, was identified as a 1:1 complex of an apoprotein and a nine-membererd ring enediyne chromophore. It exhibits potent in vivo antitumor activity and also shows pronounced activity against Gram-positive bacteria. The structural complexity of the kedarcidin chromophore, as well as its unique mode of action, makes it a challenging and fascinating target for the total synthesis. The structure of chromophore was originally assigned by Leet et al. in 1993. In 1997, we found that the kedarcidin chromophore has the β-azatyrosyl moiety, instead of the α-amino acid form. Consequently, the absolute configuration of the chromophore was also revised. In 2007, Myers and co-workers synthesized the revised mold of the chromophore and proposed the C10 epi compound (1) as the proper structure of chromophore. Herein, we report the successfull synthesis of the protected aglycon of kedarcidin chromophore 10. In our earlier work, we have succeeded in constructing the multicyclic diyne ansamacrolide, possessing the entire carbon skeleton of the kedarcidin chromophore aglycon. However, introduction of the C8-C9 epoxide was unsuccessful due to the transannular steric hindrance of the ansa-bridge. Therefore, an alternative synthetic strategy is required for the construction of the nine-membered epoxy diyne core. Our successful new strategy involves the C5-C6 cyclization of 11 which has the C8-C9 epoxide and a reductive olefination of the C4-C5 diol derivative 33. Synthesis was initiated by an etherification of newly prepared five-membered ring moiety 16, which possesses the proposed configuration at C10, with a pyridine moiety 4. Sonogashira coupling with an alkyne moiety 15 and subsequent macrolactonization gave 30 as a single atoropisomer. An amide formation with a naphtoate moiety 6 afforded 31. The nine-membered ring formation at C5-6 position via CeCl_3/LiHMDS mediated cydization protocol gave the nine-membered diyne 9 with the entire carbon skelton of the kedarcidin aglycon. To complete the synthesis of the protected aglycon 10, the reductive elimination of C4-5 dioxy functionalities was established. p-Trifluoromethylbenzoate and mesylate were chosen as the diol derivative. Selective deprotection of TES ether and mesylation followed by the formation of p-trifluoromethylbenzoate ester gave 33 as the suitable precoursor for C4-5 olefination. The Sml_2mediated reductive olefination smoothly proceeded to give the protected aglycon 10 as a sole product. Possible reduction of epoxide was not observed. As described, we achieved the total synthesis of the protected aglycon of kedarcidin chromophore 10 for the first time.

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Aspergillus fumigatus, a ubiquitous saprophytic fungus, is a potentially deadly pathogen that causes invasive aspergillosis in immunocompromised individuals. Some studies have indicated a relationship between the secondary metabolites of A. fumigatus and its virulence; therefore an understanding of mycotoxin biosynthesis will shed light on aspergillosis therapy as well as basic biology regarding its pathogenicity. Here, we present a comprehensive characterization of the ftm gene cluster of A. fumigatus with the aim of elucidating the secondary metabolites that arise from the gene cluster and their biosynthesis. Although it has been suggested that the ftm cluster is involved in the biosynthesis of diketopiperazine mycotoxins, such as fumitremorgins and tryprostatins, the biosynthesis that is based on the ftm cluster is mostly obscure. The main points of our study outcomes are as follows: (i) We elucidated all the secondary metabolites that arise from the ftm cluster, including a tremorgenic mycotoxin, verruculogen. (ii) We revealed that the ftm genes direct the biosynthesis of verruculogen through eight steps. Verruculogen was isolated as a tremorgenic mycotoxin in the 1970s and contains the unique epidioxy-bridge in the structure. (iii) We discovered the novel enzyme FtmF. This enzyme catalyzes the final step in the verruculogen biosynthesis, i.e., epidioxy formation of fumitremorgin B. (iv) We collected the natural products of the verruculogen biosynthesis and characterized its breast cancer resistance protein inhibitory activity.

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Alkylresorcinols and alkylpyrones, which have a polar aromatic ring and a hydrophobic alkyl chain, are phenolic lipids found in plants, fungi, and bacteria. In the Gram-negative bacterium Azotobacter vinelandii, phenolic lipids in the membrane of dormant cysts are essential for encystment. We report here an ars operon in A. vinelandii that is responsible for the biosynthesis of the alkylresorcinols in the cysts. The ars operon consisted of four genes, two of which encoded a type III polyketide synthase (PKS), ArsB and ArsC, and another two of which encoded a type I fatty acid synthase (FAS), ArsA and ArsD. In vitro experiments revealed that ArsB and ArsC were an alkylresorcinol synthase and an alkylpyrone synthase, respectively. In vivo and in vitro reconstitution of phenolic lipid synthesis systems with the Ars enzymes suggested that the C22-C26 fatty acids produced by ArsA and ArsD remained attached to the ACP domain of ArsA and were transferred hand-to-hand to the active-site cysteine residues of ArsB and ArsC. The phenolic lipids in A. vinelandii were thus found to be synthesized solely from malonyl-CoA by the four members of the ars operon. This is the first demonstration that a type I FAS interacts directly with a type III PKS through substrate transfer. The importance of the alkylresorcinols for encystment was confirmed by gene inactivation experiments; the lack of alkylresorcinols synthesis caused by ars mutations resulted in the formation of severely impaired cysts, as observed by electron microscopy.

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Benzalacetone synthase (BAS) is a plant-specific chalcone synthase (CHS) superfamily type III poyketide synthase (PKS) that catalyzes a one-step decarboxylative condensation of 4-coumaroyl-CoA with malonyl-CoA. The diketide forming activity of Rheum palmatum BAS is attributed to the characteristic substitution of the conserved active-site Phe215 with Leu (numbering in Medicago sativa CHS). To further understand the intimate three-dimensional structural details of the enzyme catalyzed processes, we carried out X-ray crystallographic analyses of R. palmatum BAS. The crystal structures solved at 1.6Å and 1.8Å resolution suggested that, unlike M. sativa CHS, R. palmatum BAS utilizes an alternative active-site pocket to lock the aromatic moiety of the coumaroyl starter for the diketide formation reaction. These results provided structural insights into the functional diversity of type III PKS superfamily enzymes, and suggest strategies for the engineered biosynthesis of plant polyketides.

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Only two different amino-acid "homopolymers" comprised of a single type of amino acid are known in nature: γ-poly-glutamic acid (γ-PGA) and ε-poly-L-lysine (ε-PL). ε-PL consisting of 25-35 L-lysine residues in isopeptide linkages has shown promise in medical and industrial applications. Elucidating the biosynthetic mechanism of ε-PL should open new avenues for creating novel classes of biopolymers. Here we report on the purification of an ε-PL synthase (Pls; 130kDa) and the cloning of its gene from an ε-PL-producing strain of Streptowyces albulus. Pls was found to be a membrane protein with six transmembrane domains. It was shown to be a single-module nonribosomal peptide synthase (NRPS) with traditional adenylation (A-domain) and thiolation domains (T-domain) in its amino (N)-terminal region. It had no traditional condensation or thioesterase domain; instead, it had three tandem carboxy (C)-terminal acyltransferase domains that catalysed L-lysine polymerization iteratively, yielding chains of diverse length (3-mer to 17-mer) in vitro. Furthermore, these domains catalysed the condensation of free L-lysine monomers (or ε-PL oligomers) as acceptors with enzyme-bound L-lysine as a donor; hence, L-lysine activated by the A-domain and T-domain was used only as an extender unit. A database search showed that Pls homologues were widely distributed among microorganisms and that amino-acid homopolymers, in addition to ε-PL and γ-PGA, might occur in nature.

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Our research has been focusing on the molecular mechanism of a luminous jellyfish, Aequorea aequorea. This jellyfish has a photoprotein (aequorin), which contains coelenterazine (CL) as a chromophore. The chromophore exists as an imidazopyrazinone peroxide as comfirmed by X-ray crystallography studies by Shimomura. We have tried to perform a dynamic analysis of CL, which is only observed on the surface of aequorin. Upon photolysis of the hydroperoxide in aequorin, we found several oxidation points. These modified sites were analyzed by using nano-LC-ESI-ion-trap (IT) -MSn using our pre-packed gradient (PPG) solvent elution program. Peptide fragnets resulting from trypsin digests of aequorin and photo-irradiated aequorin were compared. The nano-LC-ESI-IT-MSn analysis provided three modified peptides. We found that three peptide fragments (T20, T21 and T22) of aequorin disappeared in the mass spectra after photolysis. We also found that Cys residues in T20, T21 and T22 of aequorin were converted to oxidized dithiothreitol adducts. We concluded that photolysis of reconstituted aequorin generated a reactive oxygen specie, which modify only Cys residues locating nearby the peroxide moiety of the chromophore.

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The folding of glycoproteins is mediated by a quality control system (Figure 1) in the endoplasmic reticulum, where UDP-Glc: glycoprotein glucosyltransferase (UGGT) serves as a "folding sensor". In this study, we focused on the molecular basis for substrate-recognition by UGGT. To obtain precise understanding of the recognition mode of this folding sensor enzyme, we synthesized various systematic inhibitors (Figure 3) and high-mannose-type glycans modified with various types of aglycon (Figure 4 and Figure 5). Comparison of their inhibitory activity (Figure 3) or reactivity (Figure 5) provided support for the hypothesis that UGGT recognizes the innermost GlcNAc residue and the hydrophobic region of client glycoproteins. Through these experiments, we found efficient acceptor substrates for UGGT. In particular, introduction of Fmoc into the aglycon, made glycans highly reactive to UGGT (compound 8 in Figure 4). Derivatives carrying fluorescent functional group TAMRA or BODIPY are also excellent substrates (compound 9 and 10 in Figure 4) and expected to be valuable for detecting the activity of UGGT.

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Dysiherbaine (DH) and its congener neodysiherbaine A (NDH) are naturally occurring excitatory amino acids with high-affinity and subunit-selectivity for kainate type ionotropic glutamate receptors, especially GluR5 and GluR6 subunits. To elucidate why DH and NDH bind selectively to GluR5, we have determined the crystal structures of human GluR5 ligand-binding core in complexes with DH and NDH, in addition to the glutamate-complex. DH and NDH form unique hydrogen-bonding and hydrophobic interactions with the amino acid residues in the binding-cleft by excluding the water molecules, which mediate hydrogen bonding interactions with glutamate. The domain openings upon binding DH and NDH are much smaller than that found in the structures of partial-agonist- or antagonist-bound forms of GluR5. Here, we propose that the efficacy and ability of an agonist are not simply in inverse proportion to the extent of domain separation, but appear to be related to the stability of the "closed" conformation of the ligand-binding core. This structural information provides the basis for rational development of a series of novel synthetic analogues as useful probes for studying iGluRs in central nervous system and as potential therapeutic leads.

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Tumor promoters are chemicals that enhance tumor formation. Phorbol-type tumor promoters activate protein kinase C (PKC), a family of serine/threonine kinases that play a pivotal role in carcinogenesis. PKC isozymes are widely recognized as targets for anticancer therapy, and many PKC inhibitors have been identified as anticancer drugs. Contrary to these PKC-targeted drugs, bryostatin 1 (bryo-1), a natural product derived from marine bryozoan Bugula neritine, shows antineoplastic activity by activating PKC. The biological effects of bryo-1 are associated with unique activation mechanism of PKCδ, a member of the PKC family. Bryo-1 binds to two C1 domains (C1A, C1B) of PKCδ in a non-selective manner and then translocates the enzyme from the cytosol to the perinuclear region. On the other hand, tumor promoters show binding preference to the C1B and induce the translocation to the plasma membrane to activate PKCδ. We have found that polyacetate-type tumor promoter aplysiatoxin (ATX) and its analog (O-Me-ATX) displayed significant binding potency for the C1A of PKCδ. This result, coupled with the report that hydrophilic analogs (LogP<3) of phorbol esters translocated PKCδ to the perinuclear region like bryo-1, prompted us to develop three hydrophilic ATX analogs (Aplog-1, 2, 3) as possible candidates for functional equivalents of bryo-1. Each Aplog has been synthesized utilizing Keck's asymmetric allylation, Smith's iodocarbonate cyclization, and Yamaguchi's lactonization as key steps. Although Aplog-3 showed poor binding affinity to the C1A and C1B of PKCδ, Aplog-1 and 2 displayed nanomolar affinity for the C1B and significant binding potency for the C1A. PKCδ translocation assay in CHO-K1 cells overexpressing GFP-tagged PKCδ revealed that Aplog-1 and 2 translocated the enzyme to the perinuclear region, implying that these Aplogs could mimic the unique PKCδ activation mechanism of bryo-1. To examine whether these Aplogs could be anti-tumor promoters like bryo-1, we evaluated their inducing abilities of Epstein-Barr virus early antigen (EBV-EA). Aplog-2 strongly induced EBV-EA as observed in tumor promoters. In sharp contrast, Aplog-1 showed weak inducing ability by itself and significantly inhibited EBV-EA induction by potent tumor promoter TPA. These results suggest that Aplog-1 could be a promising candidate for a functional equivalent of bryo-1.

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Homoisocitrate dehydrogenase (HICDH) and 3-isopropylmalate dehydrogenase (IPMDH) belong to a unique family of bifunctional decarboxylating dehydrogenases. HICDH is involved in the α-aminoadipate pathway of L-lysine biosynthesis in higher fungi such as yeast and human pathogenic fungi. This enzyme catalyzes the oxidative decarboxylation of homoisocitrate into 2-ketoadipate using NAD^+ as a coenzyme. IPMDH is a key enzyme in L-leucine biosynthesis of microorganism and plants, and catalyzes the NAD^+ dependent oxidative decarboxylation of the substrate 3-isopropylmalate to 2-oxoisocaproate. Since L-lysine and L-leucine are essential amino acids for animal, these enzymes are considered to be a potential target for new antifungal and antimicrobial agents. Firstly, we designed and synthesized a series of aza-, oxa-, and thia-analogues of homoisocitrate as a potential inhibitor for HICDH as shown Fig. 3. Among them, thia-analogue showed a strong competitive inhibitory activity as K_i=97nM toward HICDH derived from Saccharomyces cerevisiae. Kinetic studies suggested that the formation of the keto-enolate intermediate played an important role in the inhibition. Based on this result, we also synthesized a series of aza-, oxa-, and thia-analogues as an inhibitor for IPMDH. As a result, thia-analogue was found to be a strong competitive inhibitor (K_i=64nM) toward IPMDH derived from Thermus thermophilus. In addition, T. thermophilus IPMDH-inhibitor-NAD^+ crystals were obtained and the structure showed that the product from thia-analogue inhibitor exited in the active site.

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Natural products have their important meanings in biological systems. Isolation of new natural products and synthesis of small molecules based on their scaffolds would be of chemical relevance to living cells and organs. In this study, we focused on the Wnt signaling, which pathways show aberrant activation in many cancer cells. We synthesized natural products melleumin A (1) and B (2), and trans-dihydroarcyriarubin C, which were isolated by us from myxomycetes (true slime molds), and also synthesized their isomers. Melleumin A (1) and B (2) are novel peptide compounds isolated from the myxomycete Physarum melleum. Their syntheses have been achieved by coupling procedures of their components, L-threonine, glycine, and an unusual amino acid, a tyrosine-attached acetic acid. We found cis-dihydroarcyriarubin C (12), and isomers of melleumin B are moderate inhibitors of Wnt signal transcription. We also developed a high-throughput system for analyzing of inhibition of making a complex between TCF and β-catenin by using microplates. The regulation of transcription by TCF/β-catenin complex is a final step of the Wnt signal pathway. In this assay, cis-dihydroarcyriarubin C (12) showed inhibition, which indicates that a cell-based Wnt signal inhibition would be caused by disruption of TCF/β-catenin complex. We also found several natural products from our natural products library that show a cell-based Wnt signal inhibition and a protein-based TCF/β-catenin complex inhibition.

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Glycinoeclepin A (1) and B (2) were isolated from dried roots of the kidney bean in 1982 as a hatch-stimulating agent of the soybean cyst nematode. Toward the total synthesis of 1 and 2, we designed a synthetic strategy based on the cyclopentene annulation method and an alkylation reaction using a bridgehead anion species. Stereoselective construction of the CD ring system was achieved through a conjugate addition reaction of optically active enone 9 with an anion species generated from nitrile 10 followed by a cyclization reaction promoted by acetic acid. After elongation of the side chain via a stereoselective hydroboration reaction followed by the Suzuki-Miyaura coupling, the configuration of the C12 hydroxyl group was inverted through stereoselective reduction of ketone 16 by using LiEt_3BH in toluene. Introduction of the A ring moiety was accomplished by the alkylation reaction of tosylate 5 with a bridgehead anion species generated from dimethylhydrazone derivative 26. Hydrolysis of 26 afforded the corresponding ketone, which was subjected to palladium-catalyzed carbonylation followed by selective oxidative cleavage of the terminal double bond to give aldehyde 3. Finally, Pinnick oxidation and subsequent hydrolysis of the ester groups provided Glycinoeclepin A (1). Glycinoeclepin B (2) was also synthesized from the common intermediate 3 by using the asymmetric Nozaki-Hiyama-Kishi coupling. Thus, Ni(II)/Cr(II)-mediated coupling reaction of aldehyde 3 and β-iodo-α,β-unsaturated ester in the presence of optically active sulfonamide 29 afforded adduct 30 as a major isomer. Although the dicarboxylic acid derived from 30 was not identical with Glycinoeclepin B, the corresponding C23-epimer 32 was converted to the natural product. These results suggested that the configuration of the C23 hydroxyl group would be β, which was further confirmed by using the modified Mosher's method.

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Pinnatoxins were isolated from the shellfish Pinna muricata and characterized by Uemura and co-workers in 1995. Owing to its unprecedented molecular architecture coupled with its biological profile as a Ca^<2+> channel activator, pinnatoxin A (1) has emerged as a highly attractive and important target for synthetic investigations. In 1998, Kishi and co-workers accomplished the first total synthesis of (-)-1, which also established the absolute stereochemistry of natural pinnatoxin A. In 2004, Hirama and co-workers reported a formal total synthesis, and very recently, Zakarian and Stivala have completed a total synthesis of 1. Herein we wish to report a total synthesis of pinnatoxin A (1). We have already reported a highly stereoselective construction of the 6,5,6-dispiro-ketal (BCD ring) system by exploiting the tandem hemiketal formation/hetero-Michael reaction sequence through the agency of LiOMe as a base. The assembly of the EF ring was effected upon exposure of the dispiroketal 8 to TsOH, completing the synthesis of the C10-C31 (BCDEF ring) portion of pinnatoxin A. With regard to the crucial Diels-Alder reaction to construct the G ring, the stereochemical arrangement requires an exo mode of cycloaddition. We found that the reaction of diene 5 with α-methylene lactone 6 exhibited complete regioselectivity and modest exo and diastereoface selectivity to give desired exo cycloadduct 4 as the major product, which was uneventfully transformed to enyne 3 in 8 steps. Upon employing a procedure reported by the Trost group, Ru-catalyzed cycloisomerization of enyne 3 proceeded with complete regioselectivity to provide 27-membered carbocycle 17 in 79% yield. The 1,2-disubstituted olefin thus formed was reduced with the Stryker reagent after oxidation of the allylic alcohol at C6. Functional group manipulations, oxidation state adjustment, self-catalyzed dehydration of the ketoamino acid to form cyclic imine (A ring) and global deprotection completed the total synthesis of pinnatoxin A.

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1,2-Cis glycosidic linkages are prevalent in natural glycans. Although key factors that control stereoselectivity of glycosylation have been largely understood, stereoselective synthesis of 1,2-cis glycosides is potentially problematic [1]. To achieve exclusive formation of desired isomer, approaches based on intramolecular aglycon delivery (IAD) are of special promise [2,3]. The methodology toward the strereoselective 1,2-cis-glycoside using naphthyl-methyl (NAP) ether-mediated IAD has been developed [4]. Namely, 2-O-NAP protected donor was cleanly converted to the mixed acetal upon oxidative activation with DDQ [5]. Subsequent activation of thioglycosidic linkage initiated the rearrangement of an aglycon from mixed acetal moiety to give a desired 1,2-cis-glycoside. Stereospecific constructions of various 1,2-cis linkages, such as β-mannopyrano-, β-arabinofurano-, and α-glucopyranosides were achieved through NAP-IAD. This methodology was successfully applied to the synthesis of various fragments of natural glycans. In particular the β-L-rhamnopyranosylation, which is one of the most challenging problems in the carbohydrate chemistry, has been successfully achieved through NAP-IAD [7]. It clearly suggests that this novel stereospecific IAD methodology is highly efficient, general and practical.

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We have previously reported that the restriction of a pyranose ring to an axial-rich conformation was an effective method for the stereoselective O-glucosidation. Here, we describe a highly β-selelctive O-glucosidation by means of the novel glucosyl fluoride whose ring was restricted to a twist-boat conformation with a 3,6-O-(o-xylylene) bridge. This glucosyl donor, 3,6-O-(o-xylylene)-D-glucopyranosyl fluoride (2), was synthesized through two routes from D-glucose derivatives 3 or 4. The glucosyl donor was activated by catalytic amount of SnCl_2-AgB(C_6H_5)_4 in benzotrifluoride to make β-glucosidic bond with various alcohols including primary, secondary, tertiary, and sterically hindered hydroxy groups on sugars in 83 to 97% yield. The all cases completed in 1〜4h with high stereoselectivity (α:β=1:>99). We also found that the reagent anomerized the produced glucosidic bond into thermodynamically more stable β-isomer, because the corresponding α-isomer was much less stable due to the restricted twist-boat conformation. This method is one of the most highly β-selective O-glucosidation without using the neighboring group participation of acyl groups at the 2-O-position. The o-xylylene group is a useful protecting group that is similar to benzyl group, which can be easily removed by hydrogenolysis. The new methodology of the restriction of the pyranose ring for the stereoselective O-glucosidation would be an alternative of the neighboring group participation, which has sustained the credibility more than a century.

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(-)-Standishinal having abeoabietane skeleton, which was isolated from Thuja standishii by Tanaka, is natural product possessing inhibitory activity against aromatase. Recently, many diterpenoids having the same skeleton have been isolated. These diterpenoids are also expected to have promising activities as medicines for female hormone-dependent cancers. Here, we will report the efficient synthetic method applicable to synthesis of the various abeoabietane type diterpenoids. In the beginning, we investigated asymmetric construction of the abeoabietane skeleton possessing quaternary stereogenic center. We anticipated that intramolecular asymmetric Heck reaction of diene 1 afford the desired tricyclic product. Because 1 was predicted to exist as two atropisomers, diene 1 was not suitable substrate for asymmetric reaction. However, we found that the asymmetric Heck reaction of 1 afforded the abeoabietane type intermediate in good yield with high enantiomeric excess by dynamic kinetic resolution through equilibration between two atropisomers. This is the first example of the asymmetric Heck reaction by dynamic kinetic resolution. Next, we applied this novel asymmetric Heck reaction to an asymmetric synthesis of natural products and succeeded in the first asymmetric synthesis of (-)-dichroanal B, (-)-dichroanone, (-)-taiwaniaquinone H.

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In a previous study, we found the significant acceleration of 6π-azaelectrocyclization by the obvious substituent effect due to the presence of a pair of C4-electron-withdrawing and C6-conjugating substituents in azatrienes to quantitatively produce the corresponding dihydropyridines. Based on this discovery, we have established the highly stereoselective asymmetric 6π-azaelectrocyclization using a chiral 7-isopropyl-cis-aminoindanol derivative 6, and moreover, we developed a one-pot asymmetric azaelectrocyclization protocol using vinylstannane, iodoolefin and aminoindanol as starting materials with high diastereoselectivity. Herein, to the further development of this one-pot reaction, we report novel synthetic methods for 2,4-disubstituted pyridines and 2,4,5-trisubstituted-2,5-chiral piperidines. At first we present a highly efficient and novel synthetic method of 2-arylpyridines using our own one-pot 6π-azaelectrocyclization followed by a base treatment. Thus, this method includes a sequence that consists of the coupling of three components (methanesulfonamide 1, vinylstannane 2, and iodoolefin 3) in the presence of a palladium catalyst, generation of dihydropyridine by smooth 6π-azaelectrocyclization, followed by aromatization by a treatment with DBU. This strategy has high generality to various vinylstannanes. In addition, more efficient pyridine synthesis on solid supports using sulfonamide resin 4 is also reported. Next, we present a useful method of the construction of the 2,4,5-trisubstituted-2,5-chiral piperidine skeleton via one-pot asymmetric 6π-azaelectrocyclization using a tetrasubstituted olefin 15, vinylstannanes 2 and aminoindanol 6. The new chiral center at C5 is completely controlled due to the thermodynamic stability of the resulting tetracyclic products containing the aminoindanol structure. This strategy also shows high generality to various vinylstannanes, which leads to successful applications to asymmetric total syntheses of two indole alkaloids: 20-epiuleine and (-)-corynantheidol.

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Gambierol (1) was isolated from cultures cells of ciguatera causative dinoflagellate, Gambierdiscus toxicus. The structure of 1 consists of trans-fused 6,6,6,6,7,6,6,7-membered octacyclic ether core (ABCDEFGH-ring) containing 18 chiral centers and a triene side chain including a conjugated (Z,Z)-diene systems. Gambierol (1) enhibits potent toxicity against mice at (LD_<50>=50μg/kg), and its symptoms resemble those caused by ciguatoxins, the principal toxin which is a very widespread seafood poisoning. The total syntheses of 1 have been accomplished based on the convergent strategy by the Sasaki, Kadota-Yamamoto, and Rainier groups, independently. We now report a formal total synthesis of gambierol (1) based on our developed two-directional strategy using SmI_2-induced cyclization and convergent strategy using an intramolecular Barbier reaction of iodo ester with t-BuLi. Synthesis of the AB-ring system: The synthesis of the AB-ring started with the B-ring 5, prepared from 2-deoxy-D-ribose. α-Hydroxy group of the A-ring was stereoselectively introduced by the SmI_2-induced intramolecular Reformatsky reaction of 6. Intramolecular hetero-Michael reaction of 8 constructed the A-ring to give 9, which was converted into the AB-ring iodo-alcohol 12 in a straightforward manner. Synthesis of the EFGH-ring system: The EFGH-ring 23 was stereoselectively synthesized by two synthetic routes through two-directional strategy. (1) The first synthesis of 23 started with the G-ring 13 prepared from 2-deoxy-D-ribose. The regioselective reactions of 14 led to diol 17 having the same functional groups at the left and right sides. The G-ring 17 was converted to the FGH-ring 19 based on double reaction at the left and right sides; i.e., hetero-Michael reaction, removal of thioacetal and SmI_2-induced double cyclization. The E-ring was then constructed by SmI_2-induced cyclization. (2) An alternative synthesis of the EFGH-ring 23 also started with the G-ring 13. The F-ring 25 having 1,3-diaxial Me groups was efficiently constructed by SmI_2-induced cyclization of 24, prepared from 13. The construction of the E- and H-rings was accomplished by using SmI_2-induced double cyclization to give trans-fused EFGH-ring 23. Treatment of 23 with disiamylborane chemoselectively reduced lactone to give ester-diol 32, which was hydrolyzed to the EFGH-ring carboxylic acid 33. Formal total synthesis of 1: The coupling of both segments 12 and 33 was performed by Shiina's procedure using MNBA and DMAP to give the iodo-ester 34. Upon treatment of 34 with t-BuLi at -78℃, the desired intramolecular Barbier reaction took place to give hemiacetal, which was dehydrated to give dihydropyran C-ring 35. Stereoselective hydroboration of 35 and protective group manipulation afforded the required α-alcohol 36, which was oxidized to cyclic ketone 37. Ring expansion of 37 by treatment with TMSCHN_2 gave the required oxepanone E-ring 38, corresponding to the key intermediate in Sasaki's total synthesis of gambierol (1). Thus, the formal total synthesis of gambierol (1) was accomplished.

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(-)-FR182877 was isolated from the fermentation broth of Streptomyces sp. No. 9885 by a research group of Fujisawa Pharmaceutical Company in 1998. This compound possesses an unprecedented hexacyclic ring system containing a bridgehead double bond as part of a vinylogous carbonate unit and 12 stereogenic centers, and exhibits microtubule stabilizing activity similar to that of Taxol. Hence, the complex structure and promising bioactivity of (-)-FR182877 have attracted considerable attention from synthetic chemists. Our synthetic approach began with the conversion of known aldehyde 8 to precursor 13 for the intramolecular Diels-Alder (IMDA) reaction. The IMDA reaction of the α,β-unsaturated aldehyde generated in situ from the corresponding acetal of 13 successfully provided the desired product 14 possessing the AB ring system as the single diastereomer. The CD ring system was constructed by the subsequent intramolecular hetero-Diels-Alder (IMHDA) reaction and the extensive studies suggested that the diastereoselectivity of the IMHDA reaction could be related to the E/Z geometry of alkene 16, which was generated in situ from 15. Consequently, we examined the IMHDA reaction of the substrate with the pure E-alkene to improve the diastereoselectivity. Horner-Wadsworth-Emmons reaction of 12 with 18E and subsequent reduction provide compound 19. Upon heating 19 in toluene, the stereoselective IMDA reaction occurred to afford the 20a as the major product. The cycloadduct was converted to the allyl alcohol 21, which was oxidized with MnO_2 to provide the corresponding aldehyde which in turn spontaneously underwent the intramolecular hetero-Diels-Alder (IMHDA) reaction in situ to afford the cycloadduct 23 as the sole product. Next, we examined the construction of the strained seven-membered ring moiety, and found that the intramolecular Heck reaction of compound 35 could only facilitate this transformation and afforded 36 incorporating the strained ABCDF ring system. The allylic alcohol 36 was successfully isomerized to the α-methyl ketone 38 with high diastereoselectivity, followed by the diastereoselective reduction, deprotection, and δ-lactone formation to accomplish the asymmetric total synthesis of (-)-FR182877. Starting from the C5 stereogenic center generated by the Evans aldol reaction, all other stereogenic centers have been successfully produced by asymmetric induction in this total synthesis.

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Dimeric pyrrole alkaloids, such as the axinellamines A and B (1), palau'amine (2) and the massadines (3) are marine derived natural products that represent a great opportunity to advance fundamental chemical synthesis. They bear as structural features a highly functionalized cyclopentane that is spiroannulated upon a guanidine unit, a second incorporation of a guanidine group within another ring framework, as well as one or two pyrrole moieties. To confirm the structures of this rare and peculiar group of natural products, many well-known synthetic organic chemists worldwide have embarked on their total synthesis. By considering a biosynthetic hypothesis, we reasoned that the syntheses of 1-3 should be feasible through an intermediate such as pre-axinellamines (4 and 5). As a result, we achieved the synthesis of an oxidative precursor to 4, namely 1,9-dideoxy-pre-axinellamine (9), via a tandem dehydration-chlorination event and direct incorporations of the two guanidine units as key steps in the 19-step synthetic accomplishment. Moreover, starting from an intermediate produced en route to 9, an oxidative ring-closing event of an intramolecular guanidine followed by a silver picolinate-catalyzed, novel C-H oxidation reaction that activated one of the guanidine units at the α position, allowed the achievement of the first total synthesis of axinellamines A and B.

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Norzoanthamine (1), isolated from zoanthid Zoanthus sp. by Uemura and co-workers in 1995, exhibits inhibition of IL-6 production. Further its hydrochloride was found to suppress the loss of bone weight and strength, and it is now considered to be a promising candidate for osteoporotic drug. In addition, 1 has attracted a great deal of attention from a synthetic point of view due to its unique structure including bisaminal moiety (DEFG rings) and congested C ring moiety. We have already developed an efficient methodology for the construction of the pentacyclic bisaminal structure from model precursor 7 under the mild conditions. On the basis of this methodology, we planned the synthetic strategy involving (1) construction of the stereochemically congested C ring at the first stage of the total synthesis, (2) intramolecular Diets-Alder reaction (IMDA) constructing the AB rings with proper stereochemistries, and (3) tandem cyclization on DEFG rings by our methodology at the last step. Now we reported here the total synthesis of Norzoanthamine (1). Starting from (-)-Hajos-Parrish ketone, we were able to obtain ketone 19 with requisite three quaternary chiral centers in the C ring by 1,4-addition to enone 18 as a key step. After introducing diene moiety to 20, AB rings were stereoselectively constructed by IMDA reaction/Ito-Saegusa oxidation sequence. Unprecedented oxidative cleavage of hydroxyketone 27 was fortunate and crucial step, affording formylester 28 possessing essential formyl group for Horner-Emmons reaction. Finally, crucial bisaminal formation from 38 was found to proceed smoothly, affording Norzoanthamine (1) in excellent yield.

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