1,N-Etheno-2′-deoxytubercidin and pyrrolo-C: synthesis, base pairing, and fluorescence properties of 7-deazapurine nucleosides and oligonucleotides

The synthesis of 1,N⁶-etheno-7-deaza-2′-deoxyadenosine (12b) which was prepared from 7-deaza-2′-deoxyadenosine (5a) with chloroacetaldehyde is described. Also the regioselective glycosylation of the 7-deazapurine-2-one at nitrogen-1 (19) furnishing the pyrrolo-C nucleoside 7a is reported and a side chain derivative with a terminal triple bond (7d) is prepared. The fluorescence properties of these nucleosides and related compounds were determined. The etheno nucleoside 12b is strongly fluorescent showing a Stokes shift of 134 nm and a quantum yield of Φ=0.53. It proved to be stable, both in acidic and in alkaline medium while the parent purine compound 10b is labile under both conditions. Compound 12b was converted into its phosphoramidite 14 and was incorporated into oligonucleotides. Compound 12b destabilizes oligonucleotide duplexes when it is located in the center of the molecule; it stabilizes when it is incorporated in the terminal base pair or acts as an overhanging nucleoside. Temperature-dependent fluorescent measurements yielded sigmoidal melting profiles when compound 12b is stacked to the terminal base pair while a linear decrease of the fluorescence is observed when the molecule is located opposite to the four canonical nucleosides in the center of the duplex.     

Synthesis and ‘double click’ density functionalization of 8-aza-7-deazaguanine DNA bearing branched side chains with terminal triple bonds

The 7-[di(prop-2-ynl)amino]prop-1-ynyl derivative of 8-aza-7-deaza-2′-deoxyguanosine (1) was synthesized from 7-iodo-8-aza-7-deaza-2′-deoxyguanosine (7) by Sonogashira cross-coupling and converted into the phosphoramidite building block 10. Oligonucleotides bearing branched side chains with terminal triple bonds were prepared by solid-phase synthesis containing single or multiple residues of 1 as 2′-deoxyguanosine surrogates. T_m measurements demonstrate that compound 1 has a positive effect on duplex stability, which is comparable to the stabilizing effect of the octa-1,7-diynylated non-branched nucleoside 2. Nucleoside 1 and corresponding oligonucleotides were functionalized by the Cu(I)-mediated 1,3-dipolar cycloaddition ‘double click’ reaction with diverse ligands (AZT 3, benzyl azide 4, 11-azidoundecanol 5 and m-dPEG™₄-azide 6). The conjugation reactions were carried out in solution and on solid support. Nucleoside 1 allowed ‘double’ functionalization of a single residue with two reporter groups. The ‘double click’ reaction proceeded smoothly even when two residues of nucleoside 1 were arranged in proximal positions. Hybridization with complementary strands led to a stable oligonucleotide duplex. Molecular modeling indicates that inspite of the crowded steric situation with four AZT ligands within closest proximal positions, all ligands are well accommodated in the major groove not disturbing the DNA helix.     

Oligonucleotides containing 7-propynyl-7-deazaguanine: synthesis and base pair stability

Oligonucleotides incorporating the propynyl derivative of 7-deaza-2′-deoxyguanosine (1) were synthesized by solid-phase oligonucleotide synthesis. As building blocks the phosphoramidites 7a,b were prepared. The incorporation of 1 into oligonucleotides exerts a positive effect on the DNA duplex stability. The duplex stabilization by 1 was higher than that of 7-iodo-7-deaza-2′-deoxyguanosine (2b). The stabilizing effect of the 7-propynyl group introduced in the 7-deazapurines is similar to that reported for 8-aza-7-deazapurines. From CD spectra it was deduced that the B-DNA structure is not significantly altered by compound 1.     

Template-directed photoreversible ligation of DNA via 7-carboxyvinyl-7-deaza-2′-deoxyadenosine

An efficient template-directed photoligation of oligodeoxynucleotide (ODN) using 7-deaza-2′-deoxyadenosine derivative ^VZA is described. When ODN containing ^VZA at the 5′ end was photoirradiated with ODNs containing a pyrimidine base at the 3′ end in the presence of template ODN, rapid and efficient ligation (cycloaddition reaction) was observed without any byproduct formation. ODNs containing ^VZA showed an extremely high reactivity as compared with those reported in previous photoligations.     

Hydrogelation and spontaneous fiber formation of 8-aza-7-deazaadenine nucleoside ‘click’ conjugates

Nucleoside hydrogels based on benzyl azide ‘click’ conjugates of 8-aza-7-deaza-2′-deoxyadenosine bearing 7-ethynyl, 7-octa-(1,7-diynyl), and 7-tri-prop-2-ynyl-amine side chains were synthesized (1, 3, 4). The cycloaddition adduct with the shortest linker (1) yields the most powerful hydrogelator forming stable gels at a concentration of 0.3 wt % of 1 in water. One molecule of 1 catches 7500 water molecules. Cycloaddition of the 8-aza-7-deaza-7-azido-2′-deoxyadenosine (9) and 3-phenyl-1-propyne (10) leads to the isomeric conjugate 2, with a C-N connectivity between the nucleobase and triazole moiety. This gel is less stable than that of the adduct 1. Both gels show a similar stability over a wide pH range (4.0-10.0). Xerogels of 1 and 2 studied by scanning electron microscopy (SEM) reveal that both click adducts (1 and 2) form long fibers spontaneously.     

Efficient synthesis of brominated tetrathiafulvalene (TTF) derivatives: solid-state structure and electrochemical behaviour

An efficient synthesis is reported for 4,5-dibromo-[1,3]dithiole-2-thione (1) and 4-bromo-1,3-dithiole-2-thione (7) by bromination of lithiated vinylene trithiocarbonate. Compound 1 acts as a convenient precursor to a number of asymmetric electron donors. This is exemplified by the formation of 4,5-dibromo-4′,5′-bis(2′-cyanoethylsulfanyl)TTF (3) by cross-coupling methodology and subsequent conversion into 4,5-dibromo-4′,5′-ethylenedithioTTF (4) by reaction with caesium hydroxide and 1,2-dibromoethane. The new donor 4,5-dibromo-4′,5′-ethylenedithiodiselenadithiafulvalene (5) was prepared by cross-coupling of 1 and 4,5-ethylenedithio-1,3-diselenol-2-one (6). The X-ray structures of 3 and 5 are reported.     

Nucleoside and oligonucleotide pyrene conjugates with 1,2,3-triazolyl or ethynyl linkers: synthesis,duplex stability,and fluorescence changes generated by the DNA-dye connector

Fluorescent nucleosides and oligonucleotides functionalized with pyrene were synthesized using ‘click’ chemistry or the Sonogashira cross-coupling reaction. The dye was connected to position-7 of 7-deaza-2′-deoxyguanosine or to the 2′-deoxyribofuranose moiety. Four different DNA-dye connectors with 1,2,3-triazolyl residues or triple bonds were constructed. Phosphoramidites of the pyrene conjugates (9, 14, 25) were prepared and used in solid-phase synthesis. Short linkers (2, 4) destabilize DNA, while long linkers (1) increased duplex stability. Nucleosides and oligonucleotides with single dye incorporations show linker dependent fluorescence. Linker dependent excimer emission with pyrenes in proximal positions was also observed. A ‘superchromophore’ formed by the 7-deaza-2′-deoxyguanosine ethynylpyrene conjugate shows strong red shifted fluorescence emission at 495 nm.     

Non-C2-symmetrical antimony-phosphorus ligand, (R/S)-2-diphenylphosphano-2′-di(p-tolyl)stibano-1,1′-binaphthyl (BINAPSb): preparation and its use for asymmetric reactions as a chiral auxiliary

A new non-C₂-symmetrical antimony-phosphorous ligand, (±)-2-diphenyl-phosphano-2′-di(p-tolyl)stibano-1,1′-binaphthyl (BINAPSb) 3, has been prepared from 2-bromo-2′-diphenylphosphano-1,1′-naphthyl 4 via its borane complex 6, and could be resolved by the separation of a mixture of the diastereomeric palladium complexes 8A and 8B derived from the reaction of (±)-3 with optically active palladium reagent (S)-7. The enantiomerically pure BINAPSb 3 has proved to be highly effective in the palladium-catalyzed asymmetric hydrosilylation of styrene as a chiral auxiliary.     

6′-Chloro-7- or 9-(2,3-dihydro-5H-4,1-benzoxathiepin-3-yl)-7H- or 9H-purines and their corresponding sulfones as a new family of cytotoxic drugs

A series of 1-(2,3-dihydro-5H-4,1-benzoxathiepin-3-yl)pyrimidine derivatives were synthesized and two of them (8 and 9) showed a modest antiproliferative activity against the MCF-7 breast cancer cell line. We then decided to change the pyrimidine base for the more lipophilic 6′-chloropurine, and the N-9′ purine (15) and N-7′ purine (17) were obtained. The sulfone N-7′-alkylated-6-chloropurine 18 was the most active derivative. Compound 17 was found to be slightly more active than its regioisomer 15, with an activity similar to that of 5-fluorouracil as a reference drug. Encouraged by these values, we tested these compounds against both the HT-29 human colon cancer cell line and the IEC-6 normal rat intestinal epithelial cell line, and 15 was found to be 12.7-fold more active against HT-29 than versus IEC-6.     

A simple synthesis of 4-(2-aminoethyl)-5-hydroxy-1H-pyrazoles

A simple four-step synthesis of 4-(2-aminoethyl)-5-hydroxy-1H-pyrazoles 8 (or their 1H-pyrazol-3(2H)-one tautomers 8′) as the pyrazole analogues of histamine was developed. First, enamino lactam 3 was prepared as the key intermediate in two steps from 2-pyrrolidinone (1). Next, acid-catalysed ‘ring switching’ transformations of 3 with monosubstituted hydrazines 4 gave N-[(1-substituted 5-hydroxy-1H-pyrazol-4-yl)ethyl]benzamides 7a-k and N-[2-(2-heteroaryl-3-oxo-2,3-dihydro-1H-pyrazol-4-yl)ethyl]benzamides 7′l-o. Benzamides 7a-k and 7′l-o were finally hydrolysed by heating in 6 M hydrochloric acid to furnish 1-substituted 4-(2-aminoethyl)-5-hydroxy-1H-pyrazoles 8a-k and 4-(2-aminoethyl)-2-heteroaryl-1H-pyrazol-3(2H)-ones 8′l-o in good overall yields.     

Simple preparation of 7-alkylamino-2-methylquinoline-5,8-diones: regiochemistry in nucleophilic substitution reactions of the 6- or 7-bromo-2-methylquinoline-5,8-dione with amines

7-Alkylamino-2-methylquinoline-5,8-diones (7) were prepared from 6-bromo-2-methylquinoline-5,8-dione (2) not from 7-bromo-2-methylquinoline-5,8-dione (1). The chemistry of the transformation of 6-bromo-2-methylquinoline-5,8-dione (2) and various alkylamines, such as piperidine, 2-methylaziridine, benzylamine, n-butylamine, cyclohexylamine, t-butylamine, and ammonia, to 7-alkylamino compounds 7 as well as the transformation of 7-bromo compound 1 and the alkylamines to 6-alkylamino-2-methylquinoline-5,8-diones 11 was studied. The efficient and simple synthetic routes of the key intermediates, 6- and 7-bromo-2-methylquinoline-5,8-diones (2 and 1), from 5,8-dihydroxy-2-methylquinoline (15) and 5,7-dibromo-8-hydroxy-2-methylquinoline (9), respectively, were developed. We also proposed the mechanism for the unusual regioselectivity on the nucleophilic amination of 6- and 7-bromo-2-methylquinoline-5,8-diones (2 and 1).     

The conversion of isothiazoles into pyrazoles using hydrazine

The conversion of isothiazoles into pyrazoles on treatment with hydrazine is investigated. The influence of various C-3, C-4 and C-5 isothiazole substituents and some limitations of this ring transformation are examined. When the isothiazole C-3 substituent is a good nucleofuge, 3-aminopyrazoles are obtained. However, when the 3-substituent is not a leaving group it is retained in the pyrazole product. Treatment of 4-bromo-3-chloro-5-phenylisothiazole 56 or 3-chloro-4,5-diphenylisothiazole 57 with anhydrous hydrazine at ca. 200 °C for a few minutes gives the corresponding 3-hydrazinoisothiazoles 61 and 64 respectively in high yields; the stability of these new hydrazines is investigated. 5,5′-Diphenyl-3,3′-biisothiazole-4,4′-dicarbonitrile 78 reacts with hydrazine to give 5,5′-diphenyl-3,3′-bi(1H-pyrazole)-4,4′-dicarbonitrile 79. Methylhydrazine reacts with 3-chloro-5-phenylisothiazole-4-carbonitrile 1 to give 3-(1-methylhydrazino)-5-phenylisothiazole-4-carbonitrile 83 and 3-amino-1-methyl-5-phenylpyrazole-4-carbonitrile 84. All products are fully characterised and rational mechanisms for the isothiazole into pyrazole transformation are proposed.     

Nitrenic reactivity of diazirines

Butyl 3-bromo-3H-diazirine-3-carboxylate (7) and 3-bromo-3-phenyl-3H-diazirine (17) exhibit nitrenic reactivity with phenylmagnesium bromide or tetrabutylammonium cyanide. The formation of several N,N′-disubstituted amidines is attributed to the intermediacy of 1-phenyl or 1-cyano-1H-diazirines possessing a singlet imidoylnitrene character at the N2 atom. Most notably, the reaction of 7 with PhMgBr in diethyl ether affords 2-hydroxy-2,2,N-triphenylacetamidine (9) and 2-methyl-5,5-diphenyl-4-phenylamino-2,5-dihydrooxazole (10) as products derived from nitrene insertion to the ether α-C–H bond.     

Cytotoxic and anti-HIV-1 constituents from leaves and twigs of Gardenia tubifera

Two new natural cycloartanes, tubiferolide methyl ester (1) and tubiferaoctanolide (2), together with the known coronalolide (3) and coronalolide methyl ester (4) have been isolated from leaves and twigs of Gardenia tubifera. In addition, a new flavone 5,3′,5′-trihydroxy-7,4′-dimethoxyflavone (5), five known flavones 6-10 and hexacosyl 4′-hydroxy-trans-cinnamate (11) were also obtained from the same source. The structures were assigned on the basis of spectroscopic methods. Compounds 3, 7, 9, and 10 showed significant cytotoxic activities only in P-388 cell line. Compound 1 was cytotoxic against P-388, KB, Col-2 and Lu-1, while 4 was active in P-388 and BCA-1. Compounds 3 and 4 displayed significant anti-HIV activities in the HIV-1RT assay; compound 7 showed moderate activity in this assay. Compounds 5-10 were also found to be active in the ^ΔTat/RevMC 99 syncytium assay.     

Enantioselective synthesis of axially chiral 1-(1-naphthyl)isoquinolines and 2-(1-naphthyl)pyridines through sulfoxide ligand coupling reactions

Racemic 1-(1′-isoquinolinyl)-2-naphthalenemethanol rac-12 was prepared through a ligand coupling reaction of racemic 1-(tert-butylsulfinyl)isoquinoline rac-7 with the 1-naphthyl Grignard reagent 10. Resolution of rac-12 was achieved through chromatographic separation of the Noe-lactol derivatives 14 and 15, providing (R)-(−)-12 of >99% ee and (S)-(+)-12 of 90% ee. The ligand coupling reaction of optically enriched sulfoxide (S)-(−)-7 (62% ee) with Grignard reagent 10 furnished rac-12, with the absence of stereoinduction resulting from competing rapid racemisation of the sulfoxide 7. Reaction of optically enriched (S)-(−)-7 with 2-methoxy-1-naphthylmagnesium bromide was also accompanied by racemisation of the sulfoxide 7, and furnished optically active (+)-1-(2′-methoxy-1′-naphthyl)isoquinoline (+)-3b in low enantiomeric purity (14% ee). The absolute configuration of (+)-3b was assigned as R using circular dichroism spectroscopy, correcting an earlier assignment based on the Bijvoet method, but in the absence of heavy atoms. Optically active 2-pyridyl sulfoxides were found not to undergo racemisation analogous to the 1-isoquinolinyl sulfoxide 7, with the ligand coupling reactions of (R)-(+)- and (S)-(−)-2-[(4′-methylphenyl)sulfinyl]-3-methylpyridines, (R)-(+)-17 and (S)-(−)-17, with 2-methoxy-1-naphthylmagnesium bromide providing (−)- and (+)-2-(2′-methoxy-1′-naphthyl)-3-methylpyridines, (−)-18 and (+)-18, in 53 and 60% ee, respectively. The free energy barriers to internal rotation in 3b and 18 have been determined, and the isoquinoline (R)-(−)-12 examined as a ligand in the enantioselectively catalysed addition of diethylzinc to benzaldehyde; (R)-(−)-12 was also converted to (R)-(−)-N,N-dimethyl-1-(1′-isoquinolinyl)-2-naphthalenemethanamine (R)-(−)-19, and this examined as a ligand in the enantioselective Pd-catalysed allylic substitution of 1,3-diphenylprop-2-enyl acetate with dimethyl malonate.     

Preparation and structures of 6- and 7-coordinate salen-type zirconium complexes and their catalytic properties for oligomerization of ethylene

A series of salen-type zirconium complexes of the general formula LZrCl₂ (L = N,N′-ethylenebis(salicylideneiminate), 3a; N,N′-ethylenebis(3,5-di-tert-butylsalicylideneiminate), 3b; N,N′-ethylenebis(5-methoxysalicylideneiminate), 3c; N,N′-ethylenebis(5-chlorosalicylideneiminate), 3d; N,N′-ethylenebis(5-nitrosalicylideneiminate), 3e; N,N′-o-phenylenebis(salicylideneiminate), 4a; N,N′-o-phenylenebis(3,5-di-tert-butylsalicylideneiminate), 4b; N,N′-o-phenylenebis(5-methoxysalicylideneiminate), 4c; N,N′-o-phenylenebis(5-chloro-salicylideneiminate), 4d) were prepared. The crystal structures of 6- and 7-coordinate zirconium complexes 4b and [4b · OCMe₂] were determined by X-ray crystallography, which reveals that a salen-type zirconium complex possesses a labile coordination site on the Zr center with a relatively stable framework and that the coordination and the dissociation of O-donor molecules occur readily at this site. The catalytic properties of 3(a-e) and 4(a-d) were studied for ethylene oligomerization in combination with Et₂AlCl as co-catalyst. Complex 3c featuring a methoxy-substituted salen ligand displayed higher activity than its analogous precursors having chloro and nitro groups as substituents. The catalytic reactions by 3(a-e) and 4(a-d) gave C₄-C₁₀ olefins and low-carbon linear α-olefins in good selectivity.     

Synthesis and ring opening reactions of 2-glyco-1,4-dimethyl-3-nitro-7-oxabicyclo[2.2.1]hept-5-enes

The high-pressure asymmetric Diels-Alder reactions of d-galacto- (1a) and d-manno-3,4,5,6,7-penta-O-acetyl-1,2-dideoxy-1-nitrohept-1-enitol (1b) with 2,5-dimethylfuran (2) afforded mixtures of cycloadducts, from which the (2S,3R)-3-exo-nitro (3a and 3b), (2R,3S)-3-exo-nitro (4a and 4b), and (2R,3S)-1′,2′,3′,4′,5′-penta-O-acetyl-1′-C-(1,4-dimethyl-3-endo-nitro-7-oxabicyclo[2.2.1]hept-5-en-2-exo-yl)-d-galacto-pentitol (5b) were isolated pure. Deacetylation of these compounds led to new chiral mono-, bi-, and tricyclic ethers, being their asymmetric centers arising from the chiral inductor used in the cycloaddition reaction. A ring opening mechanism through a 1-nitro-1,3-cyclohexadiene intermediate has been proposed.     

Selective radical cyclisation of propargyl bromoethers to tetrahydrofuran derivatives via electrogenerated nickel(I) tetramethylcyclam

The controlled-potential reduction of [1-bromo-2-methoxy-2-(prop-2′-ynyloxy)ethyl]benzene (1a), 1-[2-bromo-2-phenyl-1-(prop-2′-ynyloxy)ethyl]-4-methoxybenzene (1b) and 2-bromo-3-(3′,4′-dimethoxyphenyl)-3-propargyloxypropanamide (1c) catalysed by (1,4,8,11-tetramethyl-1,4,8,11-tetraazacyclotetradecane)nickel(I), [Ni(tmc)]⁺, at a vitreous carbon cathode in DMF/Et₄NBF₄ leads to 2-methoxy-4-methylene-3-phenyl-tetrahydrofuran (2a), 2-(4′-methoxyphenyl)-4-methylene-3-phenyl-tetrahydrofuran (2b) and 2-(3′,4′-dimethoxyphenyl)-3-carbamoyl-4-methylenetetrahydrofuran (2c), respectively, in very high yields.     

A simple and stereodivergent strategy for the synthesis of 3′-C-branched 2′,3′-dideoxynucleosides exploiting (Z)-but-2-en-1,4-diol and (R)-2,3-cyclohexylideneglyceraldehyde

Barbier type additions of allylic bromide 4, derived from (Z)-but-2-en-1,4-diol 2 to (R)-2,3-cyclohexylideneglyceraldehyde 1 were performed through mediation with Zn employing Luche’s procedure and also with low valent Cu, Co, and Fe which were produced via bimetal redox strategy in THF to afford 5c,d as the major products. From these, 5a,b were prepared following an oxidation-reduction protocol. Compound 5c was exploited as a representative starting material to develop a simple and inexpensive strategy toward the synthesis of 3′-C-branched 2′,3′-dideoxynucleosides having stereodiversity at 3′- and 4′-positions.     

Synthesis and crystal structure of 2′-deoxy-2′-fluoro-4′-thioribonucleosides: substrates for the synthesis of novel modified RNAs

We report herein the synthesis of appropriately protected 2′-deoxy-2′-fluoro-4′-thiouridine (5), -thiocytidine (7), and -thioadenosine (35) derivatives, substrates for the synthesis of novel modified RNAs. The synthesis of 5 and 7 was achieved via the reaction of 2,2′-O-anhydro-4′-thiouridine (3) with HF/pyridine in a manner similar to that of its 4′-O-congener whereas the synthesis of 35 from 4′-thioadenosine derivatives was unsuccessful. Accordingly, 35 was synthesized via the glycosylation of the fluorinated 4-thiosugar 25 with 6-chloropurine. The X-ray crystal structural analysis revealed that 2′-deoxy-2′-fluoro-4′-thiocytidine (8) adopted predominately the same C3′-endo conformation as 2′-deoxy-2′-fluorocytidine.