Biosynthesis of aristeromycln: Evidence for the intermediacy of a 4β-hydroxymethyl-1α, 2α, 3α-trihydroxycyclopentanetriol.

Evidence for the intermediacy of a 4β-hydroxymethyl-1α, 2α, 3α-trihydroxycyclopentanetdol (5 or6) in the biosynthesis of the nucleoside antibiotic aristeromycin (1) has been obtained by administration of doubly-labeled forms of D-glucose to the fermentation broth ofStreptomyces citricolor followed by trapping of the tetrol5 using isotope dilution methods.     

First and efficient synthesis of phosphonodifluoromethylene analogues of nucleoside 3'-phosphates: crucial role played by sulfur in construction of the target molecules

Phosphoric esters of secondary alcohols are ubiquitous in biological systems. However, despite the obvious interest of the corresponding difluoromethylene phosphonates as isopolar mimics, a single example of such an analogue featuring this particular substitution pattern has so far been reported in the literature, due to synthetic problems associated with their preparation. The lithium salt of diethyl difluoromethylphosphonothioate 28d provides a solution to this problem, as demonstrated by an 8-step synthesis of all five fully protected analogues of nucleoside 3'-phosphates in 9-18% overall yield, from readily available ketones. Sulfur is shown to play a crucial role in the introduction of the phosphorus-substituted difluoromethylene unit onto the furanose ring. Complete diastereoselectivity is observed in the three steps of the process requiring stereocontrol. The key conversion of the P=S bond into its oxygenated analogue is simply achieved by use of m-chloroperoxybenzoic acid. It is noteworthy that the synthesis can be carried out on large scale: a 31-g batch of compound 26b has been prepared. The deprotected nucleoside 3'-phosphate analogues can be liberated from their precursors as exemplified by the conversion of 7b, 8b, and 9b into the corresponding difluorophosphonic acids, isolated in the form of their disodium salts.     

Carbocyclic analogs of nucleosides. Part 2.. Synthesis of 2′,3′-dideoxy-5′-homonucleoside analogs

A set of derivatives of cyclopentaneacetic acid cis-substituted at position 3 by nucleoside bases (both purines and pyrimidines) were prepared and characterized (see 11, 14 , and 23a, b; Schemes 2–4). These molecules are carbocyclic analogs of 2′,3′-dideoxy-5′-homonucleosides. In this synthesis, the skeleton was constructed from norbornanone and a novel method based on Mitsunobu chemistry used for the introduction of nucleoside-base substituents. The scope of this method was further explored via the preparation of a cyclobutyl analog of dideoxyguanosine (see 17 , Scheme 3).     

New synthesis of 2',3'-didehydro-2',3'-dideoxynucleosides.

The 3'-iodonucleoside 4 and the 3'-O-methylsulfonylthymidine 9 have been synthesized by condensation of silylated uracils 2 with methyl 5-O-tert-butyldiphensilyl-2,3-dideoxy-3-iodo-D-threo-pentofuran oside (3) and methyl 5-O-tert-butyldiphenylsilyl-2-deoxy-3-methylsulfonyl-D-erythro- pentofuranoside (8), respectively. The nucleoside 4 and 9 produced the corresponding 2',3'-didehydro-2',3'-dideoxynucleosides 5 in an elimination reaction on treatment with sodium methoxide. The compounds 5b showed no antiviral activity against HIV-1.     

Synthesis Of 2'-Amino-3'-Methoxyflavone (Pd 98059)

2'-Amino-3'-methoxyflavone selectively blocks the activity of MAP kinase kinase (MEK) by inhibiting the activation of MAP kinase. This paper reports a straightforward synthesis of PD 98059 and 2-fert-butyl-8-methoxy-l,4-dihydro-quinolin-4-one.     

Thio effects on the departure of the 3'-linked ribonucleoside from diribonucleoside 3',3'-phosphorodithioate diesters and triribonucleoside 3',3',5'-phosphoromonothioate triesters: implications for ribozyme catalysis

To provide a solid chemical basis for the mechanistic interpretations of the thio effects observed for large ribozymes, the cleavage of triribonucleoside 3',3',5'-phosphoromonothioate triesters and diribonucleoside 3',3'-phosphorodithioate diesters has been studied. To elucidate the role of the neighboring hydroxy group of the departing 3'-linked nucleoside, hydrolysis of 2',3'-O-methyleneadenosin-5'-yl bis[5'-O-methyluridin-3'-yl] phosphoromonothioate (1 a) has been compared to the hydrolysis of 2',3'-O-methyleneadenosin-5'-yl 5'-O-methyluridin-3'-yl 2',5'-di-O-methyluridin-3'-yl phosphoromonothioate (1 b) and the hydrolysis of bis[uridin-3'-yl] phosphorodithioate (2 a) to the hydrolysis of uridin-3'-yl 2',5'-di-O-methyluridin-3'-yl phosphorodithioate (2 b). The reactions have been followed by RP HPLC over a wide pH range. The phosphoromonothioate triesters 1 a,b undergo two competing reactions: the starting material is cleaved to a mixture of 3',3'- and 3',5'-diesters, and isomerized to 2',3',5'- and 2',2',5'-triesters. With phosphorodithioate diesters 2 a,b, hydroxide-ion-catalyzed cleavage of the P--O3' bond is the only reaction detected at pH >6, but under more acidic conditions desulfurization starts to compete with the cleavage. The 3',3'-diesters do not undergo isomerization. The hydroxide-ion-catalyzed cleavage reaction with both 1 a and 2 a is 27 times as fast as that compared with their 2'-O-methylated counterparts 1 b and 2 b. The hydroxide-ion-catalyzed isomerization of the 3',3',5'-triester to 2',3',5'- and 2',2',5'-triesters with 1 a is 11 times as fast as that compared with 1 b. These accelerations have been accounted for by stabilization of the anionic phosphorane intermediate by hydrogen bonding with the 2'-hydroxy function. Thio substitution of the nonbridging oxygens has an almost negligible influence on the cleavage of 3',3'-diesters 2 a,b, but the hydrolysis of phosphoromonothioate triesters 1 a,b exhibits a sizable thio effect, k(PO)/k(PS)=19. The effects of metal ions on the rate of the cleavage of diesters and triesters have been studied and discussed in terms of the suggested hydrogen-bond stabilization of the thiophosphorane intermediates derived from 1 a and 2 a.     

Oligonucleotides incorporating 8-aza-7-deazapurines: synthesis and base pairing of nucleosides with nitrogen-8 as a glycosylation position

Oligonucleotides incorporating the unusually linked 8-aza-7-deazapurine N8-(2'-deoxyribonucleosides) 3a,b (purine and 6-amino-2-chloropurine analogues) were used as chemical probes to investigate the base pairing motifs of the universal nucleoside 8-aza-7-deazapurin-6-amine N8-(2'-deoxyribofuranoside) 2 (adenine analogue) and that of the 2,6-diamino compound 1. Owing to the absence of an amino group on the nucleoside 3a the low stability of oligonucleotide duplexes incorporating this compound opposite to the four canonical DNA-constituents indicate hydrogen bonding and base pairing for the universal nucleosides 1 and 2 which form much more stable duplexes. When the 6-amino-2-chloro-8-aza-7-deazapurine nucleoside 3b replaces 1 and is located at the same positions, two sets of duplexes are formed (i) high Tm duplexes with 3b located opposite to dA or dC and (ii) low Tm duplexes with 3b located opposite to dG or dT. These results are due to the steric clash of the 2-chloro substituent of 3b with the 2-oxo group of dT or the 6-oxo group of dG while the 2-halogeno substituents are well accommodated in the base pairs formed with dA or dC. For comparison duplexes incorporating the regularly linked nucleosides 4-6a,b containing the same nucleobases as those of 1-3a,b were studied.     

A new strategy for the stereoselective synthesis of 2,3-dideoxy-3-fluoro-D-ribofuranose derivatives

The important intermediate for syntheses of various anti-HIV nucleoside analogues, methyl 2,3-dideoxy-3-fluoro-6-O-(p-methylbenzoyl)-a-D-ribofuranoside, was synthesized starting from 2-deoxy-D-ribose in 5 steps with an overall yield of 24%. This strategy can be also used for synthesizing other analogues with various substitutions at 3-C position.     

Molecular orbital studies on nucleoside antibiotics. IV. conformation of 3'-deoxyadenosine (cordycepin) and 3'-amino-3'-deoxyadenosine

PCILO computations have been carried out on the conformation of 3'-deoxyadenosine (cordycepin) and 3'-amino-3'-deoxyadenosine. These nucleoside antibiotics results as a consequence of modification in the sugar part of adenosine. Both C(2')-endo and C(3')-endo sugar puckerings have been considered in the computations and the results obtained indicate that these nucleoside antibiotics have very similar conformational preferences as compared to those of the parent nucleoside adenosine. This similarity which is quite marked in aqueous solution has important biological significance.     

A ring-closing metathesis based synthesis of bicyclic nucleosides locked in S-type conformations by hydroxyl functionalised 3′,4′-trans linkages

A [4.3.0]bicyclic nucleoside that contains an unsaturated hydroxylated 3′,4′-trans linkage has been efficiently synthesised. Thus, from diacetone-d-glucose as the starting material, stereoselective Grignard reactions for the introduction of allyl groups, a nucleobase coupling and, subsequently, a ring-closing metathesis (RCM)-reaction were applied as the key reactions. The cyclohexene moiety introduced in this nucleoside reveals a large potential for further derivatisation, and as the first example, a stereoselective dihydroxylation followed by deprotection afforded a multihydroxylated bicyclic nucleoside. The configuration and conformational behaviour was determined by NMR spectroscopy and ab initio calculations, and both this bicyclic nucleoside and its unsaturated analogue were found to be strongly restricted in S-type conformations.     

An EPR study of γ-irradiated single crystals of cholesta-4, 6-diene-3-one

Cholesta-4, 6-diene-3-one single crystals irradiated with γ-rays at room temperature have been investigated by electron spin resonance. EPR spectra at room temperature exhibit a characteristics triplet which splits into two doublets. The main triplet has been interpreted as being caused by the addition of a hydrogen atom to the 7-position of the molecule, leaving an unpaired electron in the 2p₂ orbital of the carbon atoms in position 6 and 4. The hyperfine spectrum is generated by interaction of the unpaired electron with two equivalent -protons in position 4 and 6 and with tow non-equivalent β-protons in position 7. The principal values of the hydrogen hyperfine tensors are determined together with the g tensor of this radical.     

Nucleoside-boron cluster conjugates - Beyond pyrimidine nucleosides and carboranes

General methods for the synthesis of new purine and pyrimidine nucleosides modified with borane clusters and metallacarborane complexes are presented. They include: (1) attachment of carborane modification at 2′ position of nucleoside via formacetal linkage formation, (2) tethering of the metallacarborane group at nucleobase part of the nucleoside via dioxane ring opening in oxonium metallacarborane, carborane or dodecaborate derivatives, and (3) ‘‘click” chemistry approach based on Huisgen 1,3-dipolar cycloaddition. Proposed methodologies extend the range of nucleoside-boron cluster conjugates available and open new areas for their applications.     

Effets des substituants sur la conformation du cycle γ-lactonique : I. Dérivés de la γ-butyrolactone

The structural study of some γ-butyrolactones substituted (i) in position 2 (position ): C₄H₄O₂Br₂ (II) and C₄H₅O₂R [R = Oφ (III); R = OCOφ (IV); R = OH (V); R = Br (VI); R = Cl (VII)] or (ii) in position 3 or 4 (β or β′): C₄H₅O₂Cl (VIII and IX) has been carried out by using different techniques of physical chemistry. Crystallographic data analysis demonstrates that in the solid state, 2,2-dibromo-γ-butyrolactone, unlike the 2,2-diphenyl-γ-butyrolactone, adopts an “envelope” structure which is comparable to those of compounds (III) and (IV). Spectroscopic data relative to the methylene bending mode δ(CH₂) are interpreted for the dissolved state in terms of rigid (III, IV, V, IX) or exchanging (VI, VII, VIII) “envelope” forms. For and β halogenated derivatives (VI, VII, VIII), quantitative analysis of infrared spectra shows a pseudo-axial predominance in apolar solvents, as found by application of the PCILO method. Interpretation of NMR spectra recorded at 250 MHz (III, IV, V, VI) confirms the data obtained by vibrational spectroscopy.     

Synthesis of gem-difluorinated nucleoside analogues of the liposidomycins and evaluation as MraY inhibitors

Two gem-difluoromethylenated nucleoside moieties of liposidomycins, 3 and 4, were designed and synthesized. Compound 3 was assembled from lactol 5 and gem-difluoromethylenated nucleoside 6. In the synthesis of target molecule 4, the coupling of the trichloroacetimidate derivative of gem-difluoromethylenated furanose 7 with nucleoside 8 in the presence of TMSOTf gave the unexpected compound 16 when CH3CN was used as solvent. This results from acetonitrile acting as a nucleophile and participating in the glycosylation reaction. This unusual process may be correlated with the presence of the electron-withdrawing gem-difluoro substituents at the C-2 position of furanose. Compound 3 demonstrated 29% inhibition of MraY at 11.4 mM.     

Enantioselective hydrogenation of α,β-unsaturated carboxylic acids over cinchonidine-modified Pd catalysts: effect of substrate structure on the adsorption mode

Systematic variation of the substrate structure in the enantioselective hydrogenation of α,β-unsaturated carboxylic acids over cinchonidine-modified Pd catalysts indicated that aryl substituents in β position changed the adsorption mode of the substrates on the metal surface and eventually the configuration of the products formed in excess as compared to aliphatic substrates. The configuration of the products formed in excess by hydrogenation of eight α,β-unsaturated carboxylic acids, of which two have not been described yet, indicated that the substrates bearing aromatic ring in β position were adsorbed on the opposite face of the CC group compared to acids having aliphatic group in β position.     

Synthesis and Anti-HIV Activity of Triazolo-fused 3',4'-Cyclic and 4'-Spiro Nucleoside Analogues

Triazolo-fused 3',4'-cyclic nucleoside 4'-spiro nucleoside analogues were synthesized by an intramolecular 1,3-dipolar cycloaddition of 4'-azido nucleoside derived azido-alkynes in a regio- and stereo-specific manner. The thymine nucleoside base in these target compounds was transformed into the corresponding 5-methyl cytosine component. The synthesized com­pounds were examined in an MAGI(multinuclear-activation galactosidase indicator) assay for exploring the anti-HIV activity and in a H9 T(human T lymphocytes H9) assay for measuring the cell toxicity.     

Synthesis of a novel uridine analogue and its use in attempts to form new cyclonucleosides using ring-closing metathesis

One novel nucleoside analogue having a hex-5-enyl group and an allyl group in the 5′-C and 3-N position was synthesized regio- and diastereoselectively from d-glucose in twelve steps. In order to reach a particular conformation of nucleosides the nucleoside formation of restricted cyclonucleoside analogues was studied via Ring-Closing Metathesis.     

Zinc-catalyzed cycloisomerizations. Synthesis of substituted furans and furopyrimidine nucleosides

5-Endo-dig cycloisomerization of 1,4- and 1,2,4- mostly aryl-substituted but-3-yn-1-ones in the presence of a catalytic amount of zinc chloride etherate (10 mol %) in dichloromethane at room temperature gave 2,5-di- and 2,3,5-trisubstituted furans in high yields (85-97%). DSC studies confirmed that a solely thermal process does not take place. A relevant catalytic process, employing mu-oxo-tetranuclear zinc cluster Zn4(OCOCF3)6O, yielded bicyclic furopyrimidine nucleosides, when starting from acetyl-protected 5-alkynyl-2'-deoxyuridines (85-86%). Furopyrimidine was deprotected or simultaneously converted into pyrrolopyrimidine nucleoside. The time/concentration dependence for the reaction of 1-phenyl-4-(4-methylphenyl)butynone to 2-(4-methylphenyl)-5-phenylfuran displayed first-order kinetics with the rate dependent on catalyst concentration. The plot of ln k(obs) versus ln[ZnCl2] indicated first-order cycloisomerization, as referred to ZnCl2 concentration, using both NMR and UV-vis reaction monitoring. The crystal structure of propyl furopyrimidine nucleoside (orthorhombic, P2(1)2(1)2(1), a/b/c = 5.684(2)/6.682(2)/36.02(2) A, Z = 4) shows C2'- endo deoxyribose puckering, and the base is found in the anti position in crystalline form.     

Simple quantification of complement factors C3 and C3b using separation by isotachophoresis

The separation of complement factors C3 and C3b by isotachophoresis in 1% agarose gel followed by immunoprecipitation and quantification is presented. Glycine was used as spacer in a nonequilibrium isotachophoresis (Acevedo, F., J. Chromatogr. 1991, 545, 391-396). Tricine, beta-alanine and Tris were the leading ion, terminating ion and counter ion, respectively. After electrophoresis the gel was incubated in rabbit anti-human complement factor C3c. The amounts of C3 and C3b in the sample were measured by optical densitometry of the Coomassie Brilliant blue-stained immunoprecipitates in the agarose gel. The correlation coefficient obtained for the logarithm of the integrated densitometric measurement vs. the logarithm of the amount of applied C3 was higher than 0.98 in calibration experiments. The extent of complement factor C3 activation is calculated as the ratio between the amount of C3b and the amount of C3b plus C3 and expressed as percent. The progress of complement activation from human blood plasma samples induced by Mg2+ and zymosan are presented as examples.