Stereocontrolled Facile Synthesis and Biological Evaluation of (3′S) and (3′R)-3′-Amino (and Azido)-3′-Deoxy Pyranonucleosides

This article describes the synthesis of (3 ′S) and (3 ′R)-3 ′-amino-3 ′-deoxy pyranonucleosides and their precursors (3 ′S) and (3 ′R)-3 ′-azido-3 ′-deoxy pyranonucleosides. Azidation of 1,2:5,6-di-O-isopropylidene-3-O-toluenesulfonyl-α-D-allofuranose followed by hydrolysis and subsequent acetylation afforded 3-azido-3-deoxy-1,2,4,6-tetra-O-acetyl-D-glucopyranose, which upon coupling with the proper silylated bases, deacetylation, and catalytic hydrogenation, obtained the target 3 ′-amino-3 ′-deoxy-β-D-glucopyranonucleosides. The desired 1-(3 ′-amino-3 ′-deoxy-β-D-allopyranosyl)5-fluorouracil was readily prepared from the suitable imidazylate sugar after azidation followed by a protection/deprotection sequence and reduction of the unprotected azido precursor. No antiviral activity was observed for the novel nucleosides. Moderate cytostatic activity was recorded for the 5-fluorouracil derivatives.     

Antibacterial 5′-O-(N-dipeptidyl)-sulfamoyladenosines

The aminoacyl-tRNA synthetase (aaRS) class of enzymes is a validated target for antimicrobial development. Aminoacyl analogues of 5′-O-(N-l-aminoacyl)-sulfamoyladenosines are known to be potent inhibitors of aaRS, but whole cell antibacterial activity of these compounds is very limited, and poor penetration into bacteria has been proposed as the main reason for this. Aiming to find derivatives that better penetrate bacteria, we developed a simple and short method to prepare dipeptidyl-derivatives of 5′-O-(N-l-aminoacyl)-sulfamoyladenosines, and used this method to prepare 18 5′-O-(N-dipeptidyl)-sulfamoyladenosines. The antibacterial activity of these derivatives and a number of reference compounds against S. aureus, E. faecalis and E. coli was determined. Several of the new derivatives showed improved antibacterial activity and an altered spectrum of antibacterial activity.     

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.     

Oligomerization of 3′-amino-3′-deoxyguanosine-5′-phosphorimidazolidate on a d(CpCpCpCpC) template

Summary 3-Amino-3-deoxyguanosine-5-phosphorimidazolidate (ImpGnh ₂) oligomerizes more rapidly and regiospecifically than related nucleotide derivatives on a d(CpCpCpCpC) template. The greater nucleophilicity of the amino group leads to efficient oligomerization even when the structure of the double-helical complex formed by the template and the substrate is not optimal for reaction. The use of amine-containing analogues should permit us to develop models of potentially prebiotic polymerization reactions that cannot be studied easily using natural nucleotides.     

Anti-AIDS agents 84. Synthesis and anti-human immunodeficiency virus (HIV) activity of 2′-monomethyl-4-methyl- and 1′-thia-4-methyl-(3′R,4′R)-3′,4′-di-O-(S)-camphanoyl-(+)-cis-khellactone (DCK) analogs

In a continuing investigation into the pharmacophores and structure–activity relationship (SAR) of (3′R,4′R)-3′,4′-di-O-(S)-camphanoyl-(+)-cis-khellactone (DCK) as a potent anti-HIV agent, 2′-monomethyl substituted 1′-oxa, 1′-thia, 1′-sulfoxide, and 1′-sulfone analogs were synthesized and evaluated for inhibition of HIV-1 replication in H9 lymphocytes. Among them, 2′S-monomethyl-4-methyl DCK (5a)³ and 2′S-monomethyl-1′-thia-4-methyl DCK (7a) exhibited potent anti-HIV activity with EC₅₀ values of 40.2 and 39.1 nM and remarkable therapeutic indexes of 705 and 1000, respectively, which were better than those of the lead compound DCK in the same assay. In contrast, the corresponding isomeric 2′R-monomethyl-4-methyl DCK (6) and 2′R-monomethyl-1′-thia-4-methyl DCK (8) showed much weaker inhibitory activity against HIV-1 replication. Therefore, the bioassay results suggest that the spatial orientation of the 2′-methyl group in DCK analogs can have important effects on anti-HIV activity of this compound class.     

阴离子交换树脂分离制备腺嘌呤核糖核苷(3′，5′-)环磷酸(cAMP)

腺嘌呤核苷3′,5′-环磷酸对癌细胞在特定条件下有抑制作用,对冠心病和牛皮癣等有缓解作用,临床上已证明。本文所介绍的合成、分离、纯化、鉴定等方法,简易而又可靠,比目前国外报道的方法有很大改进,特别是采用国产离子交换树脂,分离简便、容量大、效果好,为大量生产创造了有利条件。——编者     

Resistance Profiles of (+) 2′-Deoxy-3′-oxa-4′-thiocytidine and (-) 2′-Deoxy-3′-oxa-4′-thio-5-fluorocytidine

Abstract

Resistant variants were selected in vitro against two novel nucleoside analogues, (+) dOTC and (-) dOTFC using the HIV-1 molecular clone HXB2D. The variants obtained displayed 6.5-fold and 10-fold resistance to these compounds, respectively. Cloning and sequencing of the RT genes of the selected viruses identified two mutations, M184I for (+) dOTC and M184V for (-) dOTFC. Results with mutated recombinant clones of HXB2D confirmed the importance of these mutations in MT-4 cells. The resistance profiles of clinical samples with wild-type or 3TC-resistant phenotypes were also studied; low to moderate levels of cross-resistance were observed against the novel compounds.     

The influence of CpG (5′-d(CpG)-3′ dinucleotides) methylation on ultrasonic DNA fragmentation

Abstract

DNA methylation is an important way of gene regulation. The variety of methods for DNA methylation analysis based on chemical modification or enzyme digestion has been proposed. However, DNA is able to undergo transformations under physical power. Here, we report that the cytosine methylation in CpG dinucleotides determines the difference in fragmentation rate of methylated and unmethylated DNA under sonication. We found that at the beginning of sonication, methylated DNAs are degraded faster than unmethylated one, and the difference in fragmentation degree can be evaluated with high reliability by quantitative polymerase chain reaction (qPCR). The optimal parameters that provide the greatest difference in amount of amplifiable DNA targets corresponding to fragmentation degree are the following: moderate amplicon size (about 150–250?bp), medium CpG sparseness (one CpG dinucleotide per ～12–14 nucleotides of the chain), and short sonication time (less than 5?min). Along with CpG, the CpA and CpT contents of amplified regions should be taken into account for proper DNA fragmentation by ultrasound as well. The obtained data could be used for elaboration of a method for comparative methylation testing, when there is no need to detect methylation of certain CpG dinucleotides. This method will be simple (can be used by any technician familiar with PCR), low cost (no need to use an expensive reagents), and fast (only brief DNA sonication and conventional qPCR are carried out).

Communicated by Ramaswamy H. Sarma     

Electrochemical Analysis of 3′-Azidothymidine (AZT)

Abstract

3′-Azido-3′-deoxythymidine (AZT) exhibits a two-electron diffusion—controlled polarographic reduction wave, with conversion to 3^′?amino-3′-deoxythymidine. The mechanism of reduction, analytical and clinical applications, and its use for one-step synthesis of amino from azido nucleosides, are described.     

Synthesis of (±)-4′-Ethynyl and 4′-Cyano Carbocyclic Analogues of Stavudine (d4T)

The synthesis of (±)-4′-ethynyl (8) and 4′-cyano (9) carbocyclic analogues of the anti-HIV agent stavudine (5, d4T) is reported. The carbocyclic unit (16) was constructed from readily available β-keto ester 10. The ethynyl or cyano group of 8 and 9 were prepared, after the introduction of thymine base to 16, by manipulation of the ester function. Evaluation of the anti-HIV activity of 8 and 9 was also carried out.     

Phosphonoformate Esters of Anti-HIV Nucleosides: 3′-Azido-3′-deoxythymidine and 2′,3′-Dideoxycytidine Derivatives Containing a Small 5′-(0-Alkoxycarbon-ylphosphinyl) or 5′-(O-Cholesterylcarbonylphosphinyl) Substituent

Abstract

A convenient general method of synthesis of 5′-O-(alkoxycarbonyl)phosphonate esters of 2′,3′-dideoxyribonucleosides is presented, using the 5′-O-(methoxycarbonyl)phosphinyl, 5′-0-(ethoxycarbonyl)phosphinyl, and 5′-O-(cholesterylcarbonyl)phosphinyl derivatives of 3′-azido-3′-deoxythymidine (AZT) and the 5′-0-(ethoxycarbonyl)phosphinyl derivative of 2′,3′-dideoxycytidine (ddC) as examples. Reaction of trimethyl phosphonoformate, methyl phosphonoformate, or dimethyl cholesterylcarbonylphosphonate with phosphorus pentachloride in carbon tetrachloride, followed by direct condensation of the resulting phosphonyl chloride with the nucleoside, gave the fully esterified phosphonoformate derivatives, which on treatment with sodium iodide in tetrahydrofuran underwent selective cleavage of the P-OMe or P-OEt groups, leaving the carboxylate esters intact. The resulting products were converted from sodium salts to ammonium salts by ion-exchange chromatography.     

癌变过程中酪氨酸转氨酶(TAT)和5′-核苷磷酸二酯酶(5′-NPDase)的变化

本文测定了大鼠及人体胚胎发育和癌变过程中TAT和5′-NPDase活力变化,所得结果表明: (一)5′-NPDase快速同工酶区带在甲胎蛋白(AFP)阴性肝癌患者血清中阳性率较高。(二)TAT,5′-NPDase和AFP也许可作为研究癌、胚以及肝癌癌变过程中基因表达的三个互补生化指标。     

利用~1H-NMR模拟谱研究三核苷酸A3′P(CH_3)5′A3′P(CH_3)5′A的构象性质

根据Altona等人的方法.利用~1H—NMR模拟谱分析了A3′P(CH_3)5A3′P(CH_3)5′A的三种非对映异构体:a、b和C的构象状态.发现:在室温下三个核糖环都是S型构象占优势(X_N≤0.44),而且随温度升高S型构象成分增多;两个二核苷酸片段-PAPA和APAP-的磷酸骨架扭角分别与A3′P(CH_3)5′A两种异构体a或b数值接近.差值约5°左右;二核苷酸片段中核糖环的重叠程度与手性磷原子的构型有关、R构型比S构型的核糖环重叠程度小;从而推测,A3′P(CH_3)5′A3′P(CH_3)5′A各种非对映异构体的稳定性为RR>RS.SR>SS,与化学合成中所见相符合.     

Synthesis and Biological Effects of 5′-Deoxy-5′-(Cyclo-Propylmethylthio)Adenosine

Abstract

A novel synthesis of the nucleoside analog, 5′-deoxy-5′-(cyclopropylmethylthio)adenosine (CPMTA, 1) has been developed. CPMTA is a closely related structural analog of 5′-deoxy-5′-(isobutylthio)-adenosine (SIBA, 2), which has been widely studied and shown to exert a multitude of biological effects. The in vitro and in vivo antitumor (L1210 leukemia) activity of CPMTA has been found to be comparable to that of SIBA, whereas its in vitro antiviral (HSV and VSV) activity is diminished. These agents are being developed as inhibitors of methylation and/or polyamine synthesis.     

Complexes of Co(II), Ni(II), Cu(II) and Zn(II) with 3′AMP and 2′AMP

Derivatives of Co(II), Ni(II), Cu(II) and Zn(II) with 3′AMP and 2′AMP were synthesized and characterized by IR UV-Vis and fluorescence spectroscopy. There seems to be bonding of the metal ion to the base in all cases. The activation test, using the complexes as allosteric labels, was carried out with rabbit muscle glycogen phosphorylase b, but the enzyme was not activated, confirming that the phosphate group must necessarily be bonded to position 5′ of the ribose in order to activate this enzyme.     

Cobalt-requiring 5′-deoxy-5′-methylthioadenosine nucleosidase from soybean (Glycine max) cotyledon

5??-Deoxy-5??-methylthioadenosine nucleosidase (MTA nucleosidase, EC 3.2.2.9) was purified from soybean (Glycine max) cotyledon. The nucleosidase was a trimer consisting of three identical subunits with a molecular mass of 59.5?kDa. The nucleosidase was a cobalt-requiring enzyme for its catalytic function. The enzymatic activity increased in a dose-dependent manner in the presence of cobalt. Cobalt was bound to the nucleosidase with a stoichiometry of 1 equivalent of cobalt/subunit. A thiol group-specific reagent reduced the enzymatic activity. Four cysteinyl residues of each subunit are considered to play an important role in binding cobalt.     

The Structure of 2′ (R)-Mercapto-2′-deoxyneplanocin A (Nucleosides and Nucleotides 48.)

Abstract

The molecular and crystal structure of 2′(R)-mercapto-2′-deoxyneplanocin A, C₁₁H₁₃N₅O₂S M.W.=279.32, has been determined by X-ray analysis. The space group is P2₁2₁2₁ with a=10.322(1), b=22.870(2), c=5.273(1)Å and z=4. The structure was solved by direct method, and least-squares refinement using 1806 reflections with |Fo| > 30(F) led to the final R value of 0.045. The sugar C(2′) atom is displaced by 0.35Å opposite to the base N(9), i.e., C(2′)-exo conformation and the torsion angle about the N(9)-C(1) bond is 26.3(4)° (anti conformation).     

Synthesis and Biological Activity of BIS(Pivaloyloxymethyl) Ester of 2′-Azido-2′-Deoxyuridine 5′-Monophosphate

Abstract

Bis(pivaloyloxymethyl) ester of 2′-azido-2′-deoxyuridine 5′-monophosphate was prepared as a prodrug to generate 2′-azido-2′-deoxyuridine 5′-diphosphate inside the cell. A synthetic route utilizing stannyl phosphate was adopted in the preparation. The prodrug was evaluated for cell growth inhibition against a variety of tumor cell lines along with 2′-azido-2′-deoxyuridine and 2′-azido-2′-deoxycytidine.     

The oxidation of N,N′-bis(4-aminophenyl)-N,N′-dimethylethylenediamine (BED) by rat liver

We have examined the oxidation of N,N′-bis(4-aminophenyl)-N,N′-dimethylethylenediamine (BED) by tissue homogenates and fractions of liver homogenates. We find that this agent both gives osmiophilic deposits in tissue blocks and readily increases the uptake of oxygen by hepatic homogenates. The highest activity was in the mitochondrial and, next, in the microsomal fractions. Kinetic evidence indicates that the former represents two enzymatic activities while the latter is only a single site. The activity was greatest in the outer membrane of the mitochondria, in agreement with electron micrographic studies and in the rough microsomal fraction. Further, it was very sensitive to both formaldehyde and detergents. The activity was not well associated with either monamine oxidase (benzylamine substrate) or xanthine oxidase activities. Activity was observed in a large number of tissues.