Highly Diastereoselective Synthesis of (2′S)-[2′-2H]-2′-Deoxyribonucleosides from the Corresponding Ribonucleosides

Abstract

The four (2′S)-[2′-²H]-2′-deoxynucleosides (>90 atom % ²H), were synthesized from the corresponding ribonucleosides involving six steps of reactions, i.e., oxidation of their 2′-hydroxyl group, stereoselective reductive deuteration of the resulting 2′-ketonucleoside intermediates with NaB²H₄ in EtOH-H₂O or EtOH, triflation, bromination with LiBr, highly stereoselective Bu₃SnH-Et₃B reduction of the resulting bromide, and, finally, unmasking.     

Synthesis of RNA Fragments Using the H-Phosphonate Method and 2′-(2′-Chlorobenzoyl) Protection

Abstract

The performance of 2′-(2-chlorobenzoyl) protected ribonucleoside H-phosphonates in the synthesis of oligoribonucleotides has been studied.     

2′-(E)-O-p-coumaroylgalactaric acid and 2′-(E)-O-feruloylgalactaric acid in citrus

2′-(E)-O-p-Coumaroyl- and 2′-(E)-O-feruloylgalactaric acids, hitherto unknown in nature, have been isolated and identified from orange peel.     

The Weak Cytokinins N,N′-bis-(1-naphthyl)urea and N,N′-bis-(2-naphthyl)urea may Enhance Rooting in Apple and Mung Bean

The present research investigates the possibility that 2 weak urea-type cytokinins, the N,N′-bis-(1-naphthyl)urea and the N,N′-bis-(2-naphthyl)urea, enhance adventitious root formation. The rooting activity was assessed using the stem slice test, the mung bean rooting test and the rooting of apple microcuttings. The two compounds influenced the adventitious rooting process differently as regards the bioassay used. In the stem slice test, in the presence of exogenous auxin, both compounds enhanced the rooted slice percentage. In mung bean shoots, the N,N′-bis-(1-naphthyl)urea enhanced the root formation at the lowest concentration used (0.01 μM) while the N,N′-bis-(2-naphthyl)urea enhanced rooting at higher concentrations. In the rooting test of apple microcuttings the N,N′-bis-(1-naphthyl)urea and the N,N′-bis-(2-naphthyl)urea slightly enhanced only the mean root number per microcutting.     

Mechanism of killing of HeLa cells by the antitumor sulfonylurea,N-(4-methylphenylsulfonyl)-N′-(4-chlorophenyl)urea (LY 181984)

Summary The antitumor sulfonylureas appear to inhibit both mitochondrial activity in susceptible human colon lines and to inhibit the oxidation of NADH by isolated plasma membrane vesicles from HeLa cells. The results reported here describe the morphological appearance of HeLa cells treated with the antitumor sulfonylurea N-(4-methylphenylsulfonyl)-N-(4-chlorophenyl)urea (LY181984). The cells remain viable for several days although the rate of increase in cell number is slowed especially at high concentrations of the drug. Cells become smaller with normal nuclei or maintain a normal size but contain multiple or enlarged nuclei. The morphological observations suggest that the drug may somehow interfere with the ability of the cells to enlarge following cytokinesis. Between 72 and 96 h, the cells begin to die. Cell death is accompanied by a condensed and fragmented appearance of the nuclear DNA as revealed by fluorescence microscopy with 4,6-diamidino-2-phenylindole suggestive of apoptosis. Early transients in loss of pH control (4 min after sulfonylurea addition) and an increase in cytoplasmic calcium (4 h after sulfonylurea addition) were observed but were small and perhaps secondary to the mechanism responsible for the failure of the cells to grow and ensuing cell death.     

Synthesis of 2′-Deoxynucleoside 5′-Methylenebis-(phosphonate)s Using 2-(4-Nitrophenyl)ethyl Methylenebis(phosphonate) as the Phosphonylating Agent.

Abstract

2-(4-Nitrophenylethyl) methylenebis(phosphonate) (1) has been prepared by reaction of 2-(4-nitrophenyl)ethyl alcohol with methylenebis(phosphonyl) tetrachloride. Compound 1 was treated with diisopropylcarbodiimide (DIC) to give bicyclic intermediate 2, which in reaction with suitably protected 2′-deoxynucleosides 3 gave P¹,P²-disubstituted methylenebis(phosphonate)s 4. Removal of the nitrophenylethyl group by β-elimination with DBU afforded the corresponding 2′-deoxynucleoside 5′-methylenebis(phosphonate) analogues 5.

     

5′-O-Dephosphorylated 2′,5′-oligoadenylate (2-5A) with 8-methyladenosine at the 2′-terminus activates human RNase L

Human ribonuclease L (RNase L), an interferon-induced endoribonuclease, becomes enzymatically active after binding to 2-5A. The 5′-phosphoryl group of 2-5A is reportedly necessary for the conformational change leading to RNase L activation. However, we found that 5′-O-dephosphorylated 2-5A tetramer analogs with 8-methyladenosine at the 2′-terminus were more effective as an activator of RNase L than the parent 2-5A tetramer. Introduction of 8-methyladenosine is thought to induce a dramatic shift of 2-5A in the binding site of RNase L.     

A differential effect of 3-(3′4′ dichlorophenyl)-1,1 dimethyl urea and atrazine on fluorescence kinetics in chloroplasts

It was found that DCMU had a differential effect at two concentration ranges on variable fluorescence kinetics in isolated chloroplasts. The increase in fluorescence rate at low concentrations of DCMU was abolished by preincubation of chloroplasts with ferricyanide or formate, treatments which were shown to convert Fe in the PS II reaction center (i.e., the FeQ_A complex) into a non-oxidizable form, but it was not affected by Tris treatment. Increase in fluorescence kinetics (at the initial linear rate) at high concentrations of DCMU was found to be abolished by Tris treatment but it was only marginally affected by ferricyanide or formate treatments. The effect of Tris could be abolished by addition of hydroquinone-ascorbate, which restored electron flow to the pool of secondary acceptors.Contrary to the effect of DCMU, no such differential concentration dependence of the variable fluorescence kinetics was found for atrazine.The increase in fluorescence kinetics (at the initial linear rate) at a low concentration rate of DCMU is presumably restricted to units which contain an oxidizable Fe in the FeQ_A complex. Increase in fluorescence kinetics (at the initial linear rate) at high DCMU concentration is probably related to the effect of DCMU at the Q_B site.Abbreviations DCMU 3-(34 dichlorophenyl)-1,1 dimethyl urea - PS II Photosystem II - Tris tris (hydroxymethyl) aminomethane     

Effect of 2-Amino Substitution on the Antiviral Effects of 5-Ethyl-2′-Deoxyuridine and (E)-5-(2-bromovinyl)-2′-Deoxyuridine and Their Incorporation into DNA

Abstract

The 2-amino derivatives of 5-ethyl-2′-deoxyuridine (EDU) and (E)-5-(2-bromovinyl)-2′-deoxyuridine (BVDU) have been synthesized and evaluated for anti-herpesvirus activity. They were at least 1000-fold less effective against herpes simplex virus replication than the parent compounds EDU and BVDU. The 5′-triphosphates of the 2-amino substituted EDU, BVDU and thymidine derivatives were also synthesized and examined on their substrate/inhibitor properties against different DNA polymerases. None of the compounds proved markedly inhibitory to HSV-1 DNA polymerase or cellular DNA polymerase a. Nor were they incorporated into the growing DNA chain.     

Synthesis of (E)-5-(2-cIOdovinyl)-3′-0-(1-Methyl-1,4-Dihydropyridyl-3 -Carbonyl)-2′-Fluoro-2′-Deoxyuridine (Ivfru-Cds) for Brain Targetted Delivery of Ivfru,an Antiviral Nucleoside

Abstract

(E)-5-(2-lodovinyl)-2′-fluoro-3′-0-(1-methyl-1,4-dihydropyridyl-3-carbonyl)-2′-deoxyuridine (11) was synthesized for future evaluation as a lipophilic, brain-selective, pyrimidine phosphorylase-resistant, antiviral agent for the treatment of Herpes simplex encephalitis (HSE). Treatment of (E)-5-(2-iodovinyl)-2′-fluoro-2′-deoxyuridine (6) with TBDMSCI in the presence of imidazole in DMF yielded the protected 5′-O-t-butyldimethylsilyl derivative (7). Subsequent reaction with nicotinoyl chloride hydrochloride in pyridine afforded (E)-5-(-2-iodovinyl)-2′-fluoro-3′-O-(3-pyridylcarbonyl)-5′-O-t-butyldimethylsily-2′-deoxyuridine (8). Deprotection of the silyl ether moiety of 8 with n-Bu₄N⁺F^? and quaternization of the resulting 3′-O-(3-pyridylcarbonyl) derivative 9 using iodomethane afforded the corresponding 1-methylpyridinium salt 10. The latter was reduced with sodium dithionite to yield (E)-5-(2-iodovinyl)-2′-fluoro-3′-O-(1-methyl-1,4-dihydropyridyl-3-carbonyl)-2′-deoxyuridine (11).     

Microbiological synthesis of 5-ethyl- and (E)-5-(2-bromovinyl)-2′-deoxyuridine

Summary The title compounds were prepared by an enzymatic transdeoxyribosylation from 2 dGuo or 2 dThd to the respective heterocyclic bases, 5-ethyluracil and (E)-5-(2-bromovinyl)uracil, using the whole bacterial cells ofEscherichia coli as a biocatalyst.     

Reaction of the 2′-Silyl and 2′-Stannyl Derivatives of 6-(Bromomethyl)Dimethylsilyl-1′,2′-Unsaturated Uridine Under Radical Conditions

The mode of cyclization (5-exo versus 6-endo) of 2-sila-5-hexen-1-yl radicals generated from 2′-tributylstannyl- and 2′-trimethylsilyl-6-(bromomethyl)dimethylsilyl-1′,2′-unsaturated uridines (8 and 9) was investigated. Although the actual structure of the reaction products differ from each other, reflecting the ease of elimination of the 2′-substituent, it was found that both substrates prefer the 5-exo-cyclization pathway.     

Nucleosides and Nucleotides. 139. Stereoselective Synthesis of (2′S)-2′-C-Alkyl-2′-deoxyuridines

Abstract

A synthetic method for (2′S)-2′-C-alkyl-2′-deoxyuridines (9) has been described. Catalytic hydrogenation of 1-[2-C-alkynyl-2-O-methoxalyl-3,5-O-TIPDS-β-D-arabino-pentofuranosyl]uracils (5) gave 1-[2-C-(2-alkyl)-2-O-methoxalyl-3,5-O-TIPDS-β-D-arabino-pentofuranosyl]uracils (4) as a major product, which were then subjected to the radical deoxygenation, affording (2′S)-2′-alkyl-2′-deoxy-3′,5′-O-TIPDS-uridines (7) along with a small amount of their 2′R epimers.

     

Degradation of the diarylpropane lignin model compound 1-(3′,4′-diethoxyphenyl)-1,3-dihydroxy-2-(4′'-methoxyphenyl)-propane and derivatives by the basidiomycete Phanerochaete chrysosporium

The white rot basidiomycete Phanerochaete chrysosporium metabolized 1-(3,4-diethoxyphenyl)-1,3(dihydroxy)-2-(4'-methoxyphenyl)-propane (XII) in low nitrogen stationary cultures, conditions under which the ligninolytic enzyme system is expressed. 3,4-Diethoxybenzyl alcohol (IV), 1,2(dihydroxy)-1-(4-methoxyphenyl)ethane (XX) and anisyl alcohol were isolated as metabolic products indicating an initial , bond cleavage of this dimer. Exogenously added XX was rapidly converted to anisyl alcohol, indicating that XX is an intermediate in the metabolism of XII. Fungal cleavage of the , bond of 1-(3-4-diethoxyphenyl)-1-(hydroxy)-2-(4'-methoxyphenyl)ethane (XI) also occurred, indicating that a hydroxymethyl group is not a prerequisite for this reaction. P. chrysosporium also metabolized 1-(4-ethoxy-3-methoxyphenyl)-2,2(dihydroxy)-2-(4'-methoxyphenyl)propane-1-ol (XIII). The major products of the degradation of this triol included 4-ethoxy-3-methoxybenzyl alcohol (III) and 2-hydroxy-1-(4-methoxyphenyl)-1-oxoethane (XXI). The nature of the products formed indicates that this triol is also cleaved directly at the , bond. The significant difference in the nature of the products formed from the diaryl propane (XII) and the triol (XIII), however, suggests that XIII is not an intermediate in the major pathway for the degradation of XII. Metabolites were identified after comparison with chemically synthesized standards by GLC-mass spectrometry.Abbreviations GLC Gas liquid chromatography - TMSi trimethylsilyl - TLC thin layer chromatography - MS mass spectrometry     

Synthesis of (±)-Methyl 5-(1′,2′-Epoxy-2′,6′-dimethylcyclohexyl)-3-methyl-(2Z,4E)-2,4-pentadienoate

The effect of tripropyltin chloride (TPT) on transport systems in E. coli was investigated. The inhibition on uptakes of ¹⁴C-l-leucine, l-proline, adenine and methyl-(α-d-gluco)pyrano-side (α-methylglucoside) by TPT was examined. The active uptake of l-leucine which utilized ATP molecule as an energy source was 100% inhibited at the concentration of 10 µg/ml TPT. On the other hand, the uptake of l-proline which was generated by an “energied” membrane state of the cells was inhibited only 40% at the same concentration of TPT. α-Methylglucoside uptake was scarcely inhibited. Adenine uptake was intensely inhibited at 20 µg/ml TPT. The effect of the delayed addition of TPT on transport systems was also examined. l-Leucine incorporated into cells was completely released from cells by TPT. Leucine binding protein (LBP) was prepared from E. coli cells and the effect of TPT on LBP activity was examined. TPT scarcely inhibited LBP activity.     

Mechanism of Action of the Fungicide Thiabendazole, 2-(4′-Thiazolyl) Benzimidazole

Thiabendazole, 2-(4'-thiazolyl) benzimidazole (TBZ) inhibited the growth of Penicillium atrovenetum at 8 to 10 mug/ml. Oxygen consumption with exogenous glucose was inhibited at 20 mug/ml, but endogenous respiration required more than 100 mug/ml. TBZ inhibited completely the following systems of isolated heart or fungus mitochondria: reduced nicotinamide adenine dinucleotide oxidase, succinic oxidase, reduced nicotinamide adenine dinucleotide-cytochrome c reductase, and succinic-cytochrome c reductase at concentrations of 10, 167, 10, and 0.5 mug/ml, respectively. Cytochrome c oxidase was not inhibited. Antimycin A and sodium azide caused the usual inhibition patterns for both fungus and heart terminal electron transport systems. In the presence of antimycin, the fungicide inhibited completely succinate-dichloro-phenolindophenol reductase and succinate-2, 2-di-p-nitrophenyl-(3, 3-dimethoxy-4, 4-biphenylene-5, 5-diphenylditetrazolium)-reductase at 2 and 4 mug of TBZ per ml, respectively. Coenzyme Q reductase required 15 mug/ml. TBZ reduced the uptake by P. atrovenetum of glucose and amino acids and decreased the synthesis of various cell components. At 120 mug/ml, the incorporation of labeled carbon from amino acids-U-(14)C was decreased: lipid, 73%; nucleic acids, 80%; protein, 80%; and a residual fraction, 89%. TBZ did not inhibit peptide synthesis in a cell-free protein-synthesizing system from Rhizoctonia solani. Probably the primary site of inhibition is the terminal electron transport system and other effects are secondary.     

Selective synthesis of cerebrosides: (2S, 3R, 4E)-1-O-β-d-galactopyranosyl-N-(2′R and 2′S)-2′-hydroxytetracosanoyl-sphingenine

The cerebrosides were first isolated by Thudicum in 1874 and the structures were established by Carteret al. in 1950 (for review, see [2]). In 1961 Shapiro and Flowers [3] reported the first total synthesis of a cerebroside1 (Fig. 1) which was identified with the natural sample, only through comparison of their i.r. data. In order to confirm the absolute configuration at C-2 of natural cerebroside1, we describe here an unambiguous synthesis of two stereoisomeric cerebrosides1 and2, and found that the¹H-NMR spectra of the synthetic1 (Fig. 2) was completely identical with that of the natural cerebroside reported recently by Dabrowskiet al. [4].In planning the synthetic route, the target structures1 and2 were disconnected at the dotted lines to give three key synthetic intermediates3, 4 and5 or6 (Fig. 1).Abbreviations Bu butyl - Ph phenyl - t-BuPh₂SiCl t-butyldiphenylsilyl chloride - MTPA -methoxy--trifluoromethylphenylacetic acid - THF tetrahydrofuran Part 36 in the series Synthetic Studies on Cell-surface Glycans, for part 35, see [1]     

Synthesis of the 2′-,3′-Didehydro-2′-,3′-dideoxy and 2′-,3′-Dideoxy Derivatives of 6-Azauridine and a New Route to 2′-,3′-Didehydro-2′-,3′-dideoxy-5-chlorouridine

Abstract

The 5′-O-(4,4′-dimethoxytrityl) and 5′-O-(tert-butyldimethylsilyl) derivatives of 2′-,3′-O-thiocarbonyl-6-azauridine and 2′,3′-O-thiocarbonyl-5-chlorouridine were synthesized from the parent nucleosides by reaction with 4, 4′-dimethoxytrityl chloride and tert-butyldimethylsilyl chloride, respectively, followed by treatment with 1,1′-thiocarbonyldiimidazole. Introduction of a 2′-,3′-double bond into the sugar ring by reaction of the 5′-protected 2′-,3′-O-thionocarbonates with 1, 3-dimethyl-2-phenyl-1, 3, 2-diazaphospholidiine was unsuccessful, but could be accomplished satisfactorily with trimethyl phosphite. Reactions were generally more successful with the 5′-silylated than with the 5′-tritylated nucleosides. Formation of 2′-,3′-O-thiocarbonyl derivatives proceeded in higher yield with 5′-protected 6-azauridines than with the corresponding 5-chlorouridines because of the propensity of the latter to form 2,2′-anhydro derivatives. In the reaction of 5′-O-(tert-butyldimethylsilyl)-2′-,3′-O-thiocarbonyl-6-azauridine with trimethyl phosphite, introduction of the double bond was accompanied by N³-methylation. However this side reaction was not a problem with 5′-O-(tert-butyldimethylsilyl)-2′-, 3′-O-thioarbonyl-5-chlorouridine. Treatment of 5′-O-(tert-butyldimethylsilyl)-2′-, 3′-didehydro-2′-,3′-dideoxy-6-azauridine with tetrabutylammonium fluoride followed by hydrogenation afforded 2′-,3′-dideoxy-6-azauridine. Deprotection of 5′-O-(tert-butyldimethylsilyl)-2′-, 3′-didehydro-2′-,3′-dideoxy-5-chlorouridine yielded 2′-,3′-didehydro-2′-,3′-dide-oxy-5-chlorouridine.     

Total synthesis of cerebrosides: (2S, 3R, 4E)-1-O-β-d-galactopyranosyl-N-(2′R and 2′S)-2′-hydroxytetracosanoylsphingenine

Total syntheses of both (2S, 3R, 4E)-1-O-β-d-galactopyranosyl-N-(2′R)-2′-hydroxytetracosanoylsphingenine 23 and the (2′S) stereoisomer were performed in an unambiguous way by employing either (2S, 3R, 4E)-N-(2′R)-2′-(tert-butyl-diphenylsilyloxy)tetracosanoylsphingenine or its (2′S) stereoisomer as the key glycosyl acceptors. The synthetic cerebroside 23 was shown to be identical with the natural product through comparison of their 400-MHz, ¹H-n.m.r. spectra, thus providing synthetic evidence for the 2′R configuration of the natural cerebroside.     

Response of NADH oxidation and growth in K-562 cells to the antitumor sulfonylurea,N-(4-methylphenylsulfonyl)-N′-(4-chlorophenyl)urea (LY181984)

Summary Growth of K-562 cells in culture is inhibited by the antitumor sulfonylureaLY181984 (N-(4-methylphenylsulfonyl)-N-(4-chlorophenyl)urea) with an ED₅₀ of about 30 M. LY181984 was shown previously to inhibit NADH oxidation by plasma membranes from HeLa cells and other sources and to influence mitochondrial oxidative phosphorylation. With K-562 cells, NADH oxidation by plasma membranes was transiently stimulated and then inhibited by LY181984. NADH oxidation by whole cells was transiently stimulated and then inhibited by 0.1 to 100 M LY181984 as well. Both the stimulations and inhibitions of activity were time-dependent. NADH oxidation by lower phase membranes depleted of plasma membranes by aqueous two-phase partition also was inhibited by micromolar and submicromolar concentrations of LY181984. Inhibition did not correlate with mitochondrial presence but rather with membranes that appeared to be fragments of the Golgi apparatus. The oxidation of NADH by whole cells and of plasma membranes that was inhibited by LY181984 was distinguished from mitochondrial NADH oxidation by resistance to inhibition by cyanide and by proceeding under oxygen-depleted conditions or an argon atmosphere. In contrast to the active antitumor agent LY181984, the inactive but chemically-related analog, LY181985 (N-(4-methylphenyl-sulfonyl)-N-(4-phenylurea), inhibited neither growth nor NADH oxidation with K-562 cells or cell fractions.