OM197-MP-AC induces the maturation of human dendritic cells and promotes a primary T cell response.

Dendritic cell (DC) maturation is critical for the induction of antigen-specific T lymphocyte responses and may be essential for the development of human vaccines relying on T cell immunity. We investigated the effects on human DC of OM-197, a synthetic pseudodipeptide derived from amino acids, linked to three fatty acid chains and devoid of endotoxin properties. OM-197 upregulated the expression of HLA-DR, CD80, CD86, CD83, CD40 and CD54 at the surface of myeloid DC naturally present in blood as well as of DC generated in vitro from monocytes using IL-4 and GM-CSF. OM-197 also induced the release of IL-12 and TNF-alpha from DC. Finally, DC incubated with OM-197 after pulsing with hepatitis B surface antigen (HBs Ag) induced in vitro expansion of IFN-gamma-secreting HBs Ag-specific CD4(+) T lymphocytes from naive individuals. Taken together, these data identify OM-197 as a potential vaccine adjuvant for the induction of Th1-type responses.

Carbohydrate mimetics-based glycosyltransferase inhibitors.

The search for carbohydrate mimetics-based glycosyltransferase inhibitors is a dynamic field that emerged 10 years ago. This review presents a description of the different types of glycosyltransferase inhibitors containing a carbohydrate mimetic (primarily an iminosugar, a carbasugar or a C-glycoside) and data on their biological activity whenever such data are available. The purpose of this account is to foster a synergy between the two expanding research areas of glycomimetics and glycosyltransferases.

D-Fructose-L-sorbose interconversions. Access to 5-thio-D-fructose and interaction with the D-fructose transporter, GLUT5.

Epimerisation and subsequent functionalization at C-5 of D-fructopyranose derivatives under Mitsunobu and Garegg's conditions provided efficient access to 5-thio-D-fructose (2) as well as to 5-azido-5-deoxy-1,2-O-isopropylidene-beta-D-fructopyranose (19), a known precursor to 2,5-deoxy-2,5-imino-D-mannitol (3). The interaction of 2 with the D-fructose transporter GLUT5, was found to be weaker than that of D-fructose, a result that suggests involvement of the ring oxygen atom in the recognition of D-fructose by GLUT5.

alpha- and beta-homogalactonojirimycins (alpha- and beta-homogalactostatins): synthesis and further biological evaluation.

The homoiminosugars alpha- and beta-homogalactonojirimycins were prepared from a common intermediate, tetra-O-benzyl-D-galacto-heptenitol 6, by way of highly stereoselective reaction sequences involving, as the key steps, an internal amidomercuration (alpha-epimer) and a double reductive amination (beta-epimer). alpha-Homogalactonojirimycin retains a large part of the potent activity of the parent galactonojirimycin and 1-deoxygalactonojirimycin as an inhibitor of alpha-galactosidases. However, by contrast with the parent iminosugars, it does not inhibit beta-galactosidases, with the exception of the Jack beans enzyme. beta-Homogalactonojirimycin is a weak alpha-galactosidase inhibitor and is completely devoid of activity towards beta-galactosidases. Thus, a marked selectivity toward one family of enzymes has been achieved by the addition of an alpha-CH(2)OH group in the structure of the parent iminosugars.

A new, stereocontrolled approach to iminosugar C-glycosides from L-sorbose.

The efficient synthesis of the iminoalditols derivatives 1 and 2 (nojirimycin alpha-C-glycosides) has been achieved in 10 steps from commercially available 2,3;4,6-di-O-isopropylidene-alpha-L-sorbofuranose in an overall yield of 23-27%.

In vitro inhibition and intracellular enhancement of lysosomal alpha-galactosidase A activity in Fabry lymphoblasts by 1-deoxygalactonojirimycin and its derivatives.

Fabry disease is a lysosomal storage disorder caused by deficient lysosomal alpha-galactosidase A (alpha-Gal A) activity. Deficiency of the enzyme activity results in progressive deposition of neutral glycosphingolipids with terminal alpha-galactosyl residue in vascular endothelial cells. We recently proposed a chemical chaperone therapy for this disease by administration of 1-deoxygalactonojirimycin, a potent inhibitor of the enzyme, at subinhibitory intracellular concentrations [Fan, J.-Q., Ishii, S., Asano, N. and Suzuki, Y. (1999) Nat. Med. 5, 112-115]. 1-Deoxygalactonojirimycin served as a specific chaperone for those mutant enzymes that failed to maintain their proper conformation to avoid excessive degradation. In order to establish a correlation between in vitro inhibitory activity and intracellular enhancement activity of the specific chemical chaperone, a series of 1-deoxygalactonojirimycin derivatives were tested for activity with both alpha-Gal A and Fabry lymphoblasts. 1-Deoxygalactonojirimycin was the most potent inhibitor of alpha-Gal A with an IC50 value of 0.04 microM. alpha-Galacto-homonojirimycin, alpha-allo-homonojirimycin and beta-1-C-butyl-deoxygalactonojirimycin were effective inhibitors with IC50 values of 0.21, 4.3 and 16 microM, respectively. N-Alkylation, deoxygenation at C-2 and epimerization at C-3 of 1-deoxygalactonojirimycin markedly lowered or abolished its inhibition toward alpha-Gal A. Inclusion of 1-deoxygalactonojirimycin, alpha-galacto-homonojirimycin, alpha-allo-homonojirimycin and beta-1-C-butyl-deoxygalactonojirimycin at 100 microM in culture medium of Fabry lymphoblasts increased the intracellular alpha-Gal A activity by 14-fold, 5.2-fold, 2.4-fold and 2.3-fold, respectively. Weaker inhibitors showed only a minimum enhancement effect. These results suggest that more potent inhibitors act as more effective specific chemical chaperones for the mutant enzyme, and the potent competitive inhibitors of alpha-Gal A are effective specific chemical chaperones for Fabry disease.

Revised structure of a homonojirimycin isomer from Aglaonema treubii: first example of a naturally occurring alpha-homoallonojirimycin.

The structure of a homonojirimycin isomer isolated from Aglaonema treublii and originally proposed as alpha-3,4-di-epi-homonojirimycin was revised to alpha-4-epi-homonojirimycin 3 ("alpha-homoallonojirimycin") on the basis of NMR analysis and synthetic studies. Its activity as a glycosidase inhibitor is compared to that of other homonojirimycin isomers.

Sucrose analogues modified at position 3: chemoenzymatic synthesis and inhibition studies of dextransucrases.

Conditions for the large-scale (molar) oxidation of sucrose by Agrobacterium tumefaciens were improved, thus leading to homogeneous solutions of 3-ketosucrose in 40% yield. Treatment of this solution with hydroxylamine or methoxylamine afforded the corresponding oximes 3a and 3b (isolated as acetates) in excellent yield. Dissolving-metal reduction of these oximes gave mixtures of amino disaccharides in which the gluco epimer (3-amino-3-deoxysucrose) was predominant. A more efficient approach to this amino sucrose was provided by the highly stereoselective hydrogenation of 3-ketosucrose peracetate (7), which gave exclusively the allo isomer 8 (2,4,6-tri-O-acetyl-alpha-D-allopyranosyl 1,3,4,6-tetra-O-acetyl-beta-D-fructofuranoside). Upon reaction with lithium azide, the triflate derived from 8, compound 9, afforded 3-azido-3-deoxysucrose peracetate (10) which was converted into 3-amino-3-deoxysucrose (12). The reaction of triflate 9 with potassium ethylxanthate led to a mixture of products (the expected 3-S-ethoxythiocarbonyl-3-thiosucrose derivative and the peracetates of 3-thiosucrose and of 3-thiosucrose disulfide), which could be all converted into 3-thiosucrose (17). Sucrose analogues 12 and 17 were not substrates of dextransucrases from various strains of L. mesenteroides, nor did they participate in glycosyl transfer reactions to an acceptor (maltose). Compounds 3a and 12 were found to be strong competitive inhibitors of the dextran synthesis process (dextransucrase from strain B-1397). These results indicate that 3a and 12 compete effectively with sucrose for the sucrose binding site but are unable to participate as glycosyl donors in the polymerization or glycosyl-transfer processes.

Multiple and long-range participation of benzyl groups in intramolecular C-arylation reactions of benzylated glycosides.

The intrinsic reactivity of furanosides bearing activated O-benzyl substituents (3-methoxybenzyl), in the presence of bidentate Lewis acids such as tin(IV) chloride, was explored. These glycosides were found to exhibit extremely interesting chemical properties. Thus, with three reactive substituents (at O-2,3,5), the corresponding glycosides (1 and 7) underwent a novel internal bis-C-arylation process, which involved successive alkylations of the benzyl groups at O-2 and O-3 ("multiple participation"), leading to the formal replacement of the two C-O bonds at the anomeric center of the glycoside by two C-C bonds. The bis-C-arylated constitution of the resulting polycyclic compounds 4 and 8, and the cis configuration of their fused ring system (a tetrahydro-[2]benzopyrano[3,4-d][2]benzoxepin derivative), were determined on the basis of their n.m.r.-spectral parameters. With two 3-methoxybenzyl substituents (at O-3 and O-5, compound 6), intramolecular alkylation of the benzyl group at O-3 or O-5 occurred when glycoside 6 was reacted with titanium(IV) chloride or tin(IV) chloride, respectively, thereby leading to novel bicyclic internal aryl C-glycosides (9 and 12) as major products ("long-range participation"). The constitution of compounds 9 and 12 was unambiguously established by the reactions of analogs of 6 bearing only one 3-methoxybenzyl substituent at a specific position (at O-3: 15; at O-5: 20). The unexpected divergent behavior of 6 in the presence of titanium(IV) and tin(IV) chloride remains to be explained. The availability of compound 9 made it possible to independently prepare the bis-C-arylated derivative 8 (by way of the reverse sequence of internal C-arylation reactions) and thereby to definitively demonstrate its constitution. These unprecedented reactions extend the scope of the intramolecular C-glycosidation of substituted sugars and provide novel methodologies in synthetic carbohydrate chemistry.

Synthesis of C-glycosylarenes by way of internal reactions of benzylated and benzoylated carbohydrate derivatives.

"Internal" C-glycosylarenes [e.g., (2R,3S,3aS,9bR)-3,3a,5,9b-tetrahydro-3-methoxy- and -3,7-dimethoxy-2-methoxymethyl-2H-furo[3,2-c][2]benzopyran] were prepared by intramolecular reactions of 2-O-benzyl derivatives of methyl 3,5-di-O-methyl-D-xylofuranoside (2) and their conversion into authentic C-glycosylated aromatic systems was investigated. The auxiliary benzylic linkage could not be cleaved by hydrogenolysis; isochroman derivatives (e.g., (3S)-3,4-dihydro-3-[(1R,2R)-2-hydroxy-1,3-dimethoxypropyl]-5-metho xy-1H- 2-benzopyran) were obtained under these conditions. However, oxidation of the primary benzylic position with ruthenium tetraoxide gave the corresponding lactone (a dihydroisocoumarin derivative, e.g., (2R,3S,3aS,9bR)-2,3,3a,9b-tetrahydro-3,7-dimethoxy-2-methoxy - methyl-5H-furo[3,2-c][2]benzopyran-5-one) which could be opened by saponification, thereby leading to a stereochemically unique C-glycosylbenzoic acid derivative. The same type of lactone was obtained directly from a derivative of 2 bearing a sufficiently reactive benzoyl group at O-2 (3,5-dimethoxybenzoyl); this process provides a useful approach to a heterocyclic system present in a variety of natural products. In related studies, the 2-O-phenyl substituent was found to be much less reactive than the 2-O-benzyl group in intramolecular Friedel-Crafts reactions of 2-O-substituted glycofuranosides. The first examples of successful internal C-arylation in the pyranoid series were achieved from 2-O-(3-methoxybenzyl)-D- mannopyranosides; the resulting "internal C-glycosides" [( 2R,3S,4S,4aS,10bS)-2,3,4,4a,6,10b- hexahydro-3,4,8-trimethoxy-2-methoxymethylpyrano[3,2-c][2]benzopyr an and 3,4-bis(benzyloxy)-2-benzyloxymethyl-8-methoxy analog] contain a heterocyclic skeleton closely related to that of the natural product bergenin.

Intramolecular C-arylation of 2,3,5-tri-O-benzyl- and 2,3,5-tri-O-(3-methylbenzyl)-pentofuranose derivatives.

Upon treatment with tin(IV) chloride, 1-O-acetyl-2,3,5-tri-O-benzyl- and 1-O-acetyl-2,3,5-tri-O-(3-methylbenzyl)pentofuranose (D-ribo, L-arabino) undergo intramolecular Friedel-Crafts alkylation of the aromatic substituent at O-2 to give unusual internal C-glycosyl compounds (isochroman derivatives) in high yield. The final products are also partially debenzylated at O-3 or O-5 (up to 25%) under these conditions. By contrast, the corresponding methyl glycosides are poor substrates for the intramolecular C-arylation reaction, as methyl 2,3,5-tri-O-(3-methylbenzyl)-beta-D-ribofuranoside was found to give preponderantly methyl 3,5-di-O-(3-methylbenzyl)-alpha-D-ribofuranoside (11) (49%), and the C-arylation product in 30% yield only in the presence of the same Lewis acid. The competitive formation of 11 is thought to be due to the anomerization of the substrate leading to a tin(IV) complex coordinated with O-1 and O-2, which promoted the cleavage of the benzyl group at O-2. These reactions provide a novel and efficient C-arylation method and suggest a new approach to selectively protected D-ribofuranose derivatives. Evidence for the uncommon C-arylated structure of the new products was gained from their 1H- and 13C(APT)-n.m.r. spectra.

Syntheses of 1-(5-deoxy-beta-D-arabino-hexofuranosyl)cytosine.

1-(5-Deoxy-beta-D-arabino-hexofuranosyl)cytosine (4'-homoara-C) (11), a higher homolog of the antileukemic agent ara-C (1-beta-D-arabinofuranosylcytosine), was prepared by two independent routes. The first one involved the inversion of configuration at C-2' of the D-ribo epimer (1-(5-deoxy-beta-D-ribo-hexofuranosyl)cytosine, 4'-homocytidine) by the diphenylcarbonate technique; the 5-deoxy-D-ribo-hexofuranosyl moiety of 4'-homocytidine was obtained by way of an anti-Markovnikov addition of iodine trifluoroacetate to the double bond of 5,6-dideoxy-1, 2-O-isopropylidene-3-O-p-tolylsulfonyl-alpha-D-ribo-hex-5-enofu ranose and reduction of the resulting iodide(s). In the second approach, 5-deoxy-1,2-O-isopropylidene-3-O-p-tolylsulfonyl-beta-D-xylo-he xofuranose was acetolyzed and condensed with 4-acetyl-N-bis(trimethylsilyl)cytosine, and alkaline treatment gave 11 by way of a 2',3'-anhydro intermediate. The structure of 11, in particular the configuration at C-2', was confirmed by its 1H- and 13C-n.m.r. spectra.