Synthesis of 6-Alkyl- and Arylamino-9-(tetrahydro-2-pyranyl)purines via 6-Methylsulfonylpurine

Abstract

A series of six potential plant hormones, 6-alky- and arylamino-9-(tetrahydro-2-pyranyl)purines were prepared in three steps in 35 to 45% overall yield from 6-methylthiopurine via 6-methylsulfonylpurine.     

Synthesis of 2-methylthio-cis- and trans-ribosylzeatin and their isolation from Pisum tRNA

The cis isomer of 6-(4-hydroxy-3-methyl-2-butenylamino)-2-methylthiopurine and its 9-β- and 9-α-d-ribofuranosyl derivatives have been synthesized and their physical and spectroscopic properties are described. The biological activities of these compounds have been determined in the tobacco bioassay and are compared with those of 6-(4-hydroxy-3-methyl-trans-2-butenylamino)-2-methylthiopurine and its β-ribofuranoside. The 6-(4-hydroxy-3-methyl-2-butenylamino)-2-methylthio-9-β-d-ribofuranosylpurine (ms-ribosylzeatin) isolated from a Pisum tRNA preparation was shown to consist of both isomers, which were separated by TLC and identified by comparisons of UV and MS with those of the synthetic compounds.     

Cytokinins in Corynebacterium fascians Cultures: Isolation and Identification of 6-(4-Hydroxy-3-methyl-cis-2-butenylamino)-2-methylthiopurine

In addition to the four cytokinins, 6-(3-methyl-2-butenylamino)purine, 6-methylaminopurine and the cis and trans isomers of 6-(4-hydroxy-3-methyl-2-butenylamino)purine, reported earlier from our laboratories, three cytokinin-active fractions have been obtained from the aqueous medium of 6-day-old Corynebacterium fascians cultures. One of these has been identified as 6-(4-hydroxy-3-methyl-cis-2-butenylamino)-2-methylthiopurine (2-methylthio-cis-zeatin, c-ms²io⁶ Ade).     

Effects of coumarin, thiopurines, and pyronin Y on amplification of phleomycin-induced death and deoxyribonucleic acid breakdown in Escherichia coli

Phleomycin (/=10 mug of phleomycin per ml were observed among 10(11)E. coli B cells screened, such mutants occurred with a frequency of 10(-6) to 10(-7) among cultures resistant to 1 to 2 mug of phleomycin per ml. These double mutants were cross-resistant to phleomycin plus caffeine. The amplifying compounds, though structurally dissimilar, shared the common characteristic of binding selectively to denatured DNA as measured by equilibrium dialysis methods. The implications of these observations in supporting a model of phleomycin amplification proposed previously (6) and their utility in providing a logic for developing a new class of antibiotics are discussed.     

Recognition of Double-Stranded Dna by Peptide Nucleic Acid

Abstract

Peptide nucleic acid (PNA) is an oligonucleotide mimic in which the backbone of DNA has been replaced by a pseudopeptide. We here show that there are distinct variations as to how PNA oligomers interact with double-stranded DNA depending on choice of nucleobases. Thymine-rich homopyrimidine PNA oligomers recognise double-stranded polynucleotides by forming PNA₂-DNA triplexes with the DNA purine strand. By contrast, cytosine-rich homopyrimidine PNAs add to double-stranded polynucleotides as Hoogsteen strands, forming PNA-DNA₂ triplexes, while homopurine, or alternating thymine-guanine, PNA oligomers invade DNA to form PNA-DNA duplexes.     

Isopoly-L-ornithine Derivatives of Thymine and Thymidine†

Abstract

L-Ornithine derivatives of thymine, and thymidine gave oligomers by solid phase elongation reactions. These oligomers 2 and 3, however, hardly interact with the complementary polynucleotide. Conformational studies of the oligomers with CD and NMR revealed that stable intramolecular hydrogen bonding was formed between thymine base and ornithine unit.

     

Synthesis of Protected 8-Substituted Deoxyribonucleosides and Its Helix Stability in Oligodeoxyribonucleotides Containing the Eco RI Recognition Site

Abstract

Octadeoxyribonucleotides with the sequences d(GGA?ATTCC), d(GGAA?TTCC), and d(GG?AATTCC) have been prepared by solid phase synthesis using the H-phosphonate units containing modified base moieties. These oligomers which have an isosterically altered recognition sequence of the restriction endodeoxyribonuclease Eco RI. The oligomers, with replacement to deoxy-7,8-dihyroadenosine-8-one (dA^OH), 8-methoxydeoxyadenosine (dA_OMe) and 8-methoxydeoxyguanosine (dG^OMe) from deoxyadenosine or deoxyguanosine were used for studying recognition phenomena at the functional group level. From thermodyamic data of these alternating octamers it was shown that the oligomer containing 8-methoxydeoxyadenosine in the center of the recognition sequence destabilizes such duplexes less strongly than the oligomers containing other 8-substituted nucleosides in the 5′-side of the recognition sequences. Further, the hydrolysis by Eco RI of the modified oligomers perfectly resisted compared to d(GGAATTCC).     

Natural and Phosphorothioate-Modified Oligodeoxyribonucleotides Exhibit a Non-Random Cellular Distribution

Abstract

Addition of specific antisense oligomers to LTK- cells infected with HSV-1 has been shown to decrease viral production (1,8). We have investigated the cellular components that contain these oligomers and a phosphorothioate derivative by Normarski light microscopy and gel analysis of sucrose gradient cell fractions. Fractionation analysis suggests that these oligomers are distributed throughout the cell in a non-random manner and gel analysis suggest that intact oligomers are not equally distributed in the cytosol, nuclear or membrane component. Information about the cellular location of antisense oligomers should aid in the understanding of their antiviral effect and in the design of more effective oligonucleotide derivatives as potential antiviral agents.     

New Building Blocks for the Preparation of Branched Oligonucleotides

Abstract

New building blocks 2 and 3 were prepared and successfully employed for the synthesis of branched oligonucleotides 5 and 6. The structure of oligomers obtained was confirmed by electrospray ionisation mass spectrometry.     

Xylose-DNA: Sequence-Dependent Duplex Stability as Function of Backbone Configuration

Abstract

The base pairing of a series of homooligomeric or telomeric oligonucleotides built-up from 2′-deoxy-β]-D-xyloadenosine and/or -thymidine were synthesized and studied with respect to their thermodynamics of duplex formation. Oligo (2′-deoxyxylonucleotides) are more stable than the corresponding regular oligomers the more d(xA-xT) elements they contain.     

Dinucleoside Monophosphate Analogues Containing Disulfide Linkages

Abstract

Two dinucleoside monophosphate analogues containing disulfide linkages (1 and 2) have been prepared for incorporation into oligonucleotides. The modified oligomers will be tested for their potential as antisense agents.     

Novel DNA Damage Mediated by Oxidation of Various Modified Nucleotide Residues

Abstract

Permanganate reaction of DNA oligomers containing an 8-oxoadenine or 5hydroxyuracil residue was studied, and the results were compared with those for an 8-oxoguanine-containing oligomers. We obtained similar results and found that the nucleotide residues neighboring the modified base were damaged and that the novel damage was induced by the oxidation of the modified base.     

Synthesis of Oligodeoxyribonucleoside Methylphosphonates Containing 2-Aminopurine

Abstract

Oligodeoxyribonucleoside methylphosphonates containing multiple 2-aminopurine bases were synthesized by solid-phase phosphonamidite chemistry for investigating their triple helix formation with natural DNA or other duplex targets and for cellular uptake studies of the methylphosphonate oligomers. The base-labile phenoxyacetyl group was used as the N ²-amino protecting group for 2-aminopurine, allowing the final deprotection of the oligomers to be performed under the standard ethylenediamine condition.     

Synthesis of 6-alkyl- and arylamino-9-(tetrahydro-2-pyranyl)purines via 6-methylsulfonylpurine

A series of six potential plant hormones, 6-alky- and arylamino-9-(tetrahydro-2-pyranyl)purines were prepared in three steps in 35 to 45% overall yield from 6-methylthiopurine via 6-methylsulfonylpurine.     

Synthetic Approaches to Oligodeoxyribonucleoside Phosphorodithioates Using Tervalent Phosphorus Monomers

Abstract

Two routes to the title compounds using nucleoside phosphorodiamidites or thiophosphoramidites have been explored. A suitably substituted thymidine thiophosphoramidite could be used for tetrazole catalysed solid support syntheses of oligomers (2 to 10 mers).     

Enzymatic Hydrolysis and Biological Activity of Oligonucleotides Containing 5-Substituted Pyrrmidine Bases

Abstract

Two series of alternating ODNs containing 5-n.alkyl-, alkenyl- and alkynyl-dU and -dC units have been prepared in order to study the kinetics of their hydrolysis by SV PDE and human serum, respectively. Both in (r⁵dUpdA)₁₀ and (r⁵dCpdG)₆ series the rate of hydrolysis decreased with increasing length of side-chain. Replacement of thymidines by 5-hexynyl-dU in different antisense oligomers resulted in considerably higher biological activity relative to that of the thymidine-containing counterparts.     

Linear and cyclic ester Oligomers of succinic acid and 1,4-butanediol: Biocatalytic synthesis and characterization

Abstract

The lipase-catalyzed synthesis of cyclic ester oligomers from non-activated succinic acid (A) and 1,4-butanediol (B) in the presence of immobilized Candida antarctica lipase B was investigated. Batch and pulse fed-batch systems were implemented to increase the formation of cyclic ester products. The substrate conversions after 24 h were 86% and 95% under batch and fed-batch operation, respectively and the product of the reaction was, for both systems, a mixture of cyclic (CEOs) and linear (LEOs) ester oligomers. Fed-batch operation afforded a product containing 71% cyclic ester oligomers (CEOs) as compared with only 52% CEOs in batch operation. Cyclic ester oligomers accumulated as the reaction progressed, with the dimer CEO₁ the most predominant product (i.e. 50% of the total products formed in fed-batch operation). The pulse fed-batch operation system was superior to the batch operation not only because higher substrate conversion and more CEOs were obtained, but also because it resulted in products with a higher degree of polymerization (DP up to 7). Cyclic ester oligomers are produced from the early stage of the reaction simultaneously with the linear ester oligomers by a ring-closure reaction on the active site of the enzyme, and not as a result of ring-chain equilibria.     

Synthesis and Some Properties of Modified Oligonucleotides. II. Oligonucleotides Containing 2′-Deoxy-2′-fluoro-β-D-arabinofuranosyl Pyrimidine Nucleosides

Abstract

In order to find the effects of unnatural nucleosides on the stability of duplex, several oligonucleotides containing 1-(2-deoxy-2-fluoro-β-D-arabinofuranosyl)-uracil(FAU),-cytosine (FAC) and -thymine (FMAU) were synthesized by two alternative approaches: phosphoramidite method on an ABI 392 synthesizer and H-phosphonate procedure on our GeneSyn I universal module synthesizer. It was shown from the melting profiles that the presence of FMAU has a large stabilizing effect on the duplex. Replacement of thymidine with FAU, or deoxycytidine with FAC resulted in the formation of less stable duplexes. Temperature-dependent CD spectroscopy demonstrated that the structures of the fluorine containing oligomers are very similar to those of unmodified oligomers.     

Phosphoramidate,Phosphorothioate, and Methylphosphonate Analogs of Oligodeoxynucleotide : Inhibitors of Replication of Human Immunodeficiency Virus

Abstract

Modified oligodeoxynucleotides complementary to RNA of human immunodeficiency virus (HIV-1) were tested for their ability to inhibit virally induced syncytium formation and expression of viral p24 protein. The modification of oligomers include replacement of phophodiester backbone with phosphorothioate, methylphosphonate and various phosphoramidates. Cells infected for four days, then treated with the antisense oligomers also showed inhibition of viral expression.     

Synthesis,Biophysical and Biological Evaluations of Novel Antisense Oligonucleosides Containing Dephosphono-Internucleosidic Linkages

Abstract

Efficient large-scale syntheses of methylene(methylimino) (MMI) linked mixed base nucleosidic dimers have been accomplished. These dimers were successfully incorporated into deoxyoligonucleotides by automated solid-support synthesis. The hybridization properties, nuclease stability, RNase H mediated cleavage and in vitro biological activity of novel chimeric oligomers have been studied. The biophysical and biological evaluation of these chimeric oligomers containing MMI linkages suggests that MMI is a promising chemical modification of the backbone linkage for the construction of antisense molecules useful as therapeutics.