Erich Zbiral's research while affiliated with University of Hull and other places

Efficient stereospecific N-formylation of ribosylamine has been achieved, affording the α-anomer directly (by reaction with formic-acetic anhydride) and the β-anomer via the corresponding formamidine derivative (by reaction with dimethylformamide dimethyl acetal). Dehydration of the α-anomer gave the corresponding isocyanide without compromising the anomeric purity. The amidine route was extended to give the N-formyl derivatives of α-xylosylamine and α-arabinosylamine.

Starting from 3-O-mesyl-1,2-O-isopropylidene-α-D-allofuranose (9) the anomeric mixtures of the requisite carbohydrates 1,2-di-O-acetyl-6-O-benzoyl-5-deoxy-3-O-mesyl-D-allofuranose 17Aα/β, 1,2-di-O-acetyl-5,6-di-O-benzoyl-3-O-mesyl-D-allofuranoses 17Bα/β, and 1,2-di-O-acetyl-5,6-di-O-benzoyl-3-O-mesyl-L-talofuranoses 17Cα/β were synthesized. 1,2-Di-O-acetyl-5-O-benzoyl-6-deoxy-3-O-mesyl-D-allofuranoses 17Dα/β and the corresponding L-talofuranoses 17Eα/β were obtained from 6-deoxy-3,5-di-O-benzoyl-1,2-O-isopropylidene-α-D-allofuranose (12) and the corresponding β-L-talofuranose 13. Coupling of these sugar derivatives with thymine gave the β-nucleoside derivatives 18A-E. Treatment of compounds 18A-E with DBU produced the corresponding 2,3′-anhydro nucleosides 19A-E with a free 2′-OH group. After deoxygenation of 2′-O-[[(4-methylphenyl)oxy]thiocarbonyl] compounds 20A-E with tributyltin hydride the 2,3′-anhydro bridge of the 2′-deoxynucleosides 21A-E was opened with LiN3 to produce the protected 3′-azido-2,3′-dideoxynucleoside derivatives 22A-G. Saponification with NaOCH3 gave 1-(3′-azido-2′,3′,5′-trideoxy-β-D-allofuranosyl) thymine (2; homo-AZT), the 5′-C-(hydroxymethyl) derivatives of AZT 1-(3′-azido-2′,3′-dideoxy-β-D-allofuranosyl)thymine (3) and 1-(3′-azido-2′,3′-dideoxy-α-L-talofuranosyl)thymine (4), and the 5′-C-methyl derivatives of AZT 1-(3′-azido-2′,3′,6′-trideoxy-β-D-allofuranosyl) thymine (5) and 1-(3′-azido-2′,3′,6′-trideoxy-α-L-talofuranosyl) thymine (6). Compounds 2-6 were evaluated for their inhibitory effect on human immunodeficiency virus type 1 (HIV-1) and type 2 (HIV-2) replication in MT-4 cells and found inactive at subtoxic concentrations. Compounds 2-4 and 6 are not effective against herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2), vaccinia virus (VV), and vesicular stomatitis virus (VSV) at 400 μg/mL. 5 is slightly active against HSV-1, HSV-2, and VV at 150, 300, and 300 μg/mL, respectively.

The initial step of influenza infection is binding of the virus particles via their hemagglutinin to cell-surface sialic acids. This study was initiated to elucidate the functional groups of the nine-carbon sialic acid molecule which interact with the hemagglutinin and contribute to the affinity of this sugar to the protein. In order to address this question, synthetic sialic acid analogues were tested in a virus adsorption inhibition assay for their inhibitory potency. Modifications in three regions of the sialic acid molecule were evaluated: the glycerol side chain (C7-C9), the N-acetyl group at C5, and the carboxy group (C1). In the glycerol side chain, the hydroxy groups at C7 and C8 appear to be important for binding through hydrogen bonds, whereas the hydroxyl at C9 does not appear to be involved. The N-acetyl group is critical for the interaction of sialic acid with the hemagglutinin. The results suggest that its contribution is mediated through hydrophobic interactions of the methyl group. Finally, the orientation of the carboxy group is essential for the binding of sialic acid to the hemagglutinin. The information gained in this study will be useful in developing novel compounds which bind more avidly to the influenza virus hemagglutinin. Such a strategy may contribute to the design of new anti-influenza drugs.

A series of neuraminic acid derivatives modified in the side chain or at C-3, C-4 or C-5 were tested as substrates of inhibitors of N-acetylneuraminate lyase (EC 4.1.3.3) from Clostridium perfringens. The results, together with Km and Ki values reported previously, indicate that the region most important for the binding of sialic acids is an equatorial zone reaching from C-8 via the ring oxygen atom to C-4 of the sugar molecule, whereas the substituents at C-9 and C-5 may be varied to a higher extent without significantly disturbing enzyme action. It is shown that stereo-electronic factors are responsible for the immediate heterolytic fragmentation of the cyclic sialic acid into pyruvic acid and 2-acetamidomannose or a related C-6 sugar.

The α-methylketoside of N-acetylneuraminic acid methylester (4) is transformed via the deacetylated compound5 into the 9,8-O-isopropylidenderivative6 which could be oxidized regioselectively by RuO4 to the corresponding 4-oxo-sialic acid analogue7. Reduction with the boraneammonia complex produces a 1:1 mixture of6 and the desired α-methylketoside of 9,8-O-isopropyliden-4-epi-N-acetyl-neuraminic acid methylester (8). Removing of the isopropylidene group gives the α-methylketoside of 4-epi-N-acetylneuraminic acid methylester (9), which was further transformed to the ammonium salt of 4-epi-N-acetylneuraminic acid α-methylketoside (10). On the other hand compound5 was turned into the 4,8,9-tri-O-t-butyldimethylsilylderivative11 a from which the corresponding 7-oxo-compound12 by oxidation with RuO4 derives. The reduction of12 with BH3 - NH3 yielded a 1:1 mixture of the starting material11 a and the desired 7-epi-derivative13 a which gives either via the purified peracetylated α-methylketosid of 7-epi-N-acetylneuraminic acid methylester (14) or a direct saponification the sodium salt of 7-epi-N-acetylneuraminic acid-α-methylketoside (15). Applying the Königs-Knorr procedure to the peracetylated 8-epi-N-acetylneuraminic acid methylester (16) gives rise to the formation of a 1:1 mixture of the corresponding α- and β-methylketosides17 and18 besides traces of the corresponding 2,3-dideoxy-2,3-dideohydro-sialic acid derivative19. After chromatographic separation of17 further saponification leads to the sodium salt of 8-epi-N-acetylneuraminic acid-α-methylketoside (20). In an analogous procedure the sodium salt of 7,8-di-epi-N-acetylneuraminic acid-α-methylketoside (25) was prepared starting from the peracetylated 7,8-di-epi-N-acetylneuraminic acid methylester (21), whereby a mixture of the α- and β-methylketosides22 and23 was formed in a ratio 95:5 besides traces of the peracetylated 2,3-dideoxy-2,3-didehydrosialic acid methylester (24).

The [heteroaryl(hydroxy)methyl]phosphonates 2-5 (indolizines 2, imidazo[1,2-a]pyridines 3, imidazo[1,2-a]pyrimidines 4, and 2-phenylthiazoles 5, are converted readily to the corresponding O,O-thiocarbonates 6-8 and 18 on treatment with p-tolyloxythiocarbonyl chloride in acetonitrile/4-dimethylaminopyridine, while reaction in pyridine/dichloromethane affords the isomeric O,S-thiocarbonates 9-11 and 19, respectively. Homolytic cleavage of both series of compounds proceeds smoothly, using tributyltin hydride/azobisisobutyronitrile (AIBN) in warm toluene to give the new heteroarylmethylphosphonates 12-14 and 20. Conversion of the O,S-thiocarbonates 9-11 and 19 into the synthetically attractive α-methylthio-substituted derivatives 15-17 and 21 is effected efficiently upon saponification with sodium methoxide in methanol and subsequent alkylation with methyl iodide.

Regioselective 1,2-addition of dimethylphosphite, diethyl (lithiomethyl)phosphonate, or (lithiomethyl)diphenylphosphine oxide to 2-cyclopenten-1-one (1a) or 2-cyclohexen-1-one (1b) affords the corresponding phosphorus containing, tertiary allylic alcohols 2-4. These, upon oxidation with chromium trioxide reagents are rearranged to the new dialkyl (3-oxo-1-cycloalkenyl)phosphonates, -methylphosphonates or [(3-oxo-1-cyclo-alkenyl)methyl]diphenylphosphine oxides 5-7, which are easily transformed to the corresponding epoxy derivatives 8-10.

The synthesis of the sodium 5-acetamido-2,3,5-trideoxy-2,3 didehydro-D-galacto-2,8-nondiulopyranosidonate 8,8-dimethyl acetal (8) and of the methyl-5-acetamido-3,5-dideoxy-a-D-galacto-2,8-nondiulopyranosidonate 8,8 dimethyl acetal (11) as well as of the methyl-5-acetamido-3,5-dideoxy-a-D-galacto-2,8-nondiulopyranosidonic acid (12) is reported involving the easily accessible 8-oxoderivative5. Compounds11 and12, respectively, showed to have glycosidic bond with remarkable stability to acidic conditions.

2,3-Didehydro-2-deoxy-N-trifluoroacetylneuraminic acid (5-trifluoroacetyl-Neu2en) (3) has been synthesised from Neu5Ac2en (1) by hydrazinolysis, to give Neu2en (2), followed by N-trifluoroacetylation. 2,3-Didehydro-2,3-dideoxy-D-glycero-D-galacto-2-nonulopyranoson ic acid (Kdn2en, 8) and 5-azido-2,3-didehydro-2,3,5-trideoxy-D-glycero-D-galacto-2-nonu lopyranosonic acid (5-azido-5-deoxy-Kdn2en, 9) have been prepared from the acetylated methyl esters of Kdn (4) and 5-azido-5-deoxy-Kdn (5) via Zemplén saponification. The behaviour of the above 2,3-didehydro-2-deoxysialic acids towards Vibrio cholerae sialidase has been investigated.

2-Deoxy-2-Heq-N-acetylneuraminic acid (1a) has been shown to be a versatile mimic for sialosyl 2-α-glycosides to study the hemagglutinin-sialic acid interaction. Starting with a 4-oxo derivative of 2-deoxy-2-Heq-sialic acid, we obtained the 4-C-methylax and 4-C-methyleq derivatives 4 and 5 as a separable mixture. The derivative 4 was formed as a single product by using tributoxymethylzirconium. The 4-C-methylene derivative 10 was formed by treatment of the ketone with Cp2ZrCl2/Zn/CH2I2. Catalytic hydrogenation of the exo-methylene compound 10 yielded the two 4-deoxy-4-C-methyl diastereomers 13 and 14. All above-mentioned derivatives could be transformed into the unprotected 4-branched 2-deoxy-2-Heq-sialic acids 7, 9, 12, 16, and 18. 2-Deoxy-2Heq-4-oxosialic acid (21) was synthesized by a different pathway.