CA2135498A1 - Synthesis of nucleotide monomers - Google Patents
Synthesis of nucleotide monomersInfo
- Publication number
- CA2135498A1 CA2135498A1 CA 2135498 CA2135498A CA2135498A1 CA 2135498 A1 CA2135498 A1 CA 2135498A1 CA 2135498 CA2135498 CA 2135498 CA 2135498 A CA2135498 A CA 2135498A CA 2135498 A1 CA2135498 A1 CA 2135498A1
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- Prior art keywords
- synthesis
- deoxyadenosine
- contacting
- benzoyl
- yield
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D473/00—Heterocyclic compounds containing purine ring systems
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H19/00—Compounds containing a hetero ring sharing one ring hetero atom with a saccharide radical; Nucleosides; Mononucleotides; Anhydro-derivatives thereof
- C07H19/02—Compounds containing a hetero ring sharing one ring hetero atom with a saccharide radical; Nucleosides; Mononucleotides; Anhydro-derivatives thereof sharing nitrogen
- C07H19/04—Heterocyclic radicals containing only nitrogen atoms as ring hetero atom
- C07H19/06—Pyrimidine radicals
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H19/00—Compounds containing a hetero ring sharing one ring hetero atom with a saccharide radical; Nucleosides; Mononucleotides; Anhydro-derivatives thereof
- C07H19/02—Compounds containing a hetero ring sharing one ring hetero atom with a saccharide radical; Nucleosides; Mononucleotides; Anhydro-derivatives thereof sharing nitrogen
- C07H19/04—Heterocyclic radicals containing only nitrogen atoms as ring hetero atom
- C07H19/16—Purine radicals
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- Saccharide Compounds (AREA)
Abstract
Methods for synthesis of 2',3'-dideoxycytidine, 2',3'-dideoxyguanosine, 2',3'-dideoxyadenosine, 2',3'-dideoxyinosine and 2',3'-dideoxyguanosine, and 3'-O-benzoyl-2'-deoxyribonucleoside.
Description
A ~ ~3 ~93/23413 PCT/U~93/04239 DESCRIPTION
SYNTHESIS OF NUCLEOTIDE MONOMERS
Backqround of the Invention This invention relates to the chemical synthesisof 2',3'-dideoxycytidine, 2~l3'-dideoxyguanosine,
SYNTHESIS OF NUCLEOTIDE MONOMERS
Backqround of the Invention This invention relates to the chemical synthesisof 2',3'-dideoxycytidine, 2~l3'-dideoxyguanosine,
2',3'-dideoxypurine-nucleosides (i.e., adenosine, guanosine and inosine nucleosides~, and 3'-0-benzoyl-2'-deoxyadenosine.
Haoqiang and Chu, 20 Synth. Commun. 1039 t 1990, describe a method for synthesis of 2'-deoxyuridine using acetyl bromide and HBr and in situ generated tributyltin hydride. Prisbe and Martin, 15 Svnth. Comm. 401, 1g85, describe a method for synthesis of ddC from 2'-deoxycytidine using pivaloyl chloride. Starret et al., g Nucleosides ~ N~ucleotides 885, 1990, describe a method for synthesis of 2',3'-didehydro-2',3'-dideoxycytidine, and a method for prep~ration of 2'-bromo-2'-deoxy-3',5'-di-O-acetyl uridine. Bhat et al., 9 Nucleosides & Nucleotides 1061, 1990, describe a method for synthesis of 2',3'-dideoxycytidine from 2'-deoxycytidine. Xaskar and Markovac, 26 J. HeterocYcl. Chem. 1531, 1989, describe a method for synthesis of 2',3'-dideoxycytidine from 2'-deoxycytidine. Chu et al.~ 54 _Ora. Chem. 2217, 1989, describe the synthesis of ddC from cytidine.
Chemical synthesis of 2',3'-unsaturated purine nucleosides i5 described by McCarthy et al., 88:7 J.A.C.S.
1549, 1966. Jain et al., 39 J. Ora. Chem. 30, 1974, describe synthesis of 2',3'-unsaturated nucleosides as intermediates in the prep~ration of 2'3'-dideoxynucleosides using acetoxyisobutyril bromide and a reductiv~ elimination step, but were unable to isolate a 2',3'-unsaturated derivative of guanosine. Chu, 54 J.
Ora. Chem. 2217, 1989, des~ribes the general synthesis o 2',3'-dideoxynucleosides starting from ribonucleosides and using tribytiltin hydride. Robins, 25 Tetrahedron Lett.
$
Haoqiang and Chu, 20 Synth. Commun. 1039 t 1990, describe a method for synthesis of 2'-deoxyuridine using acetyl bromide and HBr and in situ generated tributyltin hydride. Prisbe and Martin, 15 Svnth. Comm. 401, 1g85, describe a method for synthesis of ddC from 2'-deoxycytidine using pivaloyl chloride. Starret et al., g Nucleosides ~ N~ucleotides 885, 1990, describe a method for synthesis of 2',3'-didehydro-2',3'-dideoxycytidine, and a method for prep~ration of 2'-bromo-2'-deoxy-3',5'-di-O-acetyl uridine. Bhat et al., 9 Nucleosides & Nucleotides 1061, 1990, describe a method for synthesis of 2',3'-dideoxycytidine from 2'-deoxycytidine. Xaskar and Markovac, 26 J. HeterocYcl. Chem. 1531, 1989, describe a method for synthesis of 2',3'-dideoxycytidine from 2'-deoxycytidine. Chu et al.~ 54 _Ora. Chem. 2217, 1989, describe the synthesis of ddC from cytidine.
Chemical synthesis of 2',3'-unsaturated purine nucleosides i5 described by McCarthy et al., 88:7 J.A.C.S.
1549, 1966. Jain et al., 39 J. Ora. Chem. 30, 1974, describe synthesis of 2',3'-unsaturated nucleosides as intermediates in the prep~ration of 2'3'-dideoxynucleosides using acetoxyisobutyril bromide and a reductiv~ elimination step, but were unable to isolate a 2',3'-unsaturated derivative of guanosine. Chu, 54 J.
Ora. Chem. 2217, 1989, des~ribes the general synthesis o 2',3'-dideoxynucleosides starting from ribonucleosides and using tribytiltin hydride. Robins, 25 Tetrahedron Lett.
$
3 PCT/US93~04235 367, 1984, and Mansuri et al., 54 J. Orq. Chem. 4780, 1989, used acetoxyisobutyril bromide and a reductive elimination step for synthesis of 2',3'-unsaturated adenosine. Sanger et al., 74 Proc. Nat. Acad. Sci. USA
5463, 1977, describe the synthesis of dideoxynucleosides and use of their 5'-triphosphates for DNA sequencing.
Herdewijn 31 J. Med. Chem. 2040, 1988 describes synthesis of 2',3'-dideoxynucleosides and their anti-HIV activity.
Herdewijn, 31 J. Med. Chem. 2040, 1988, describes synthesis o~ 2',3'-dideoxynucleosides and their anti-HIV activity. McCarthy et al., 88:7 J.A.C.S. 1549, lg66, describe synthesis of adenine nucleosides. Jain et al., 39 J. Orq. Chem. 30-, -1974, describe synthesis of 2',3'-dideoxy nucleosides. Chu, 54 J. Orq. Chem. 2217, 1989, describes the general synthesis of 2',3'-dideoxynucleosides. Sanger et al., 74 Proc. Natl. Acad.
Sci. USA 5463, 1977, describe the synthesis of dideoxyguanosine.
Holy, 1 Synthetic Procedures in Nucleic Acid ChemistrY 172, 1968, describes a method for synthesis of 3'-O-acetyl-2'-deoxyadenosine from 2'-deoxyadenosine in four steps and 45% overall yield; Ogilvie, 51 Can.~_J.
Chem. 3799, '973, describes a method for synthesis o~ 3i-O-benzoyl-2'-deoxyadenosine from 2'-deoxyadenosine in four steps and 71~ overall yield, and alternatively in 3- steps- -and 38~ overall yield~ Applicant was told that it is-~
preferred to use benzoyl cyanide for selective benzoylation of deoxynucleosides.
Summary of the Invention -In a first aspect, this invention features an improved economical sy~thetic method for the preparation of dideoxyuridine (ddU) which is then converted- lnto dideoxycytidine (ddC) via a 4-triazolyl intermediate. The method is not only cost efficient, but can be scaled up to several hundred gram quantities. The method generally utilizes inexpensive uridine as a starting material which 93/2~13 ~ ;) 8 PCT/US93/04239 is converted in a 7 step reaction sequence to ddC with a yield of about 30~.
The ddC can be used for chemical synthesis of sugar-modified nucleotides; chemical synthesis of DNA
chain terminators; and chemical synthesis of anti-HIV
2',3'-dideoxynucleosides.
Specifically, in this aspect, the invention features a method for chemical synthesis of 2',3'-dideoxycytidine in which 5'-silylated-2',3'-deoxyuridine is used to form ddC, e.q., by reaction with l,2,4-triazole, a phosphoro~s oxychloride, followed by treatment with ammonia, and the silyl group of the resultins product cleaved by use of an ion-exchange resin (specifically, that referred to as the Amberlyst A-26 (F )) in toluene.
In a second aspectl the invention features a method for synthesis of dideoxyguanosine by directly converting N2-isobutyrylguanosine to a 2',3'-unsaturated derivative by action of acetoxyisobutyryl bromide in moist acetonitrile, with subsequent reductive elimination by Zn/Cu/DMF and deacylation by ~aOMe/methanol. Catalytic hydrogenation and ammonolysis of the N2-isobutyryl-2',3'-didehydroguanosine provides 2',3'-dideoxyguanosine in 30%
yield.
The method of this invention utiliæes N2-isobutyrylguanosine as a starting compound, and thusovercomes problems with isolation of unsaturated compounds. The described method is cost efficient and can be scaled up to multi-gram ~uantity.
-Thus, in this aspect, the invention features a method for chemical synthesis of 2',3'-dideoxyguanosine, including contacting N2-isobutyrylguanosine with ace~oxyisobutyryl bromide in acetonitrile with subsequent reductive elimination, deacylation, hydrogenation and ammonolysis.
In a third aspect, the invention features a convenient "one-pot" (i.e., without purification of intermediates~ synthesis of key intermediates: 5'-O-tert-`
G1~3~
W093/234]3 PCT/US~3/04239 butyldiphenylsilyl-3'-O~mesyl(tosyl)-2'-deoxyadenosine, 5'-O-tert-butyldiphenylsilyl-3'-O-mesyl(tosyl)-2'-deoxyinosine, and N2 isobutyryl~O6[(p-nitrophenyl)ethyl]-5'-O-tert-butyldiphenylsilyl-3'-O-mesyl(tosyl)-2'-deoxyguanosine, and their conversion to 2',3'-unsaturated derivatives by simultaneous elimination-deblocking procedures, and subsequent hydrogenation to 2',3'-dideoxynucleosides.
The method of this invention utilizes selective protection of the 5'-OH group of 2'-deoxyadenosine (or 2'-deoxyinosine) and N2-isobutyryl-2'-deoxyguanosine by introducing a tert-butyldiphenylsilyl protecting group.
The high selectivity of this step allows the preparation of the above key compounds in a "one-pot" procedure without isolation of intermediates. Simultaneous removal of the 5'-Q-tert-butyldiphenylsilyl group during the elimination step reduces the total number of steps and gives a high yield of 2',3'-unsaturated nucleosides. The ,-described methods are cost efficient, and can be scaled up to multi-gram quantity.
Thus, in this aspect, the invention features a method for chemical synthesis of 2',3'-dideoxyadenosine, 2',3'-dideoxyinosine, and 2',3'-dideoxyguanosine wlthout purification of intermediate compounds by sequentially treating: (a) N2-isobutyryl 2'-deoxyguanosine with a t- -- -butyl-diphenylsilylchloride, triphenylphosphine, diethyl--- -azodicarboxylate, p-nitrophenylethanol and methanesulfonyl (or p-toluenesulfonyl) chloride; or (b) by treating 2'-deoxyadenosine or 2'-deoxyinosine sequentially witX -t-butyldiphenylsilylchloridet and methanesulfonyl (or p-toluenesulfonyl) chloride.
In a fourth aspect, the invention features a method for synthesis of 3'-O-benzoyl-2'-deoxyadenosine based on the efficient protection of the 5'-hydroxyl group of 2'-deoxyadenosine, using stable t butyldimethylsilyl ether (TBDMS) protection. Acylation with benzoyl cyanide occurs exclusively at the remaining free sugar hydroxyl 7 d~ ~ 8 ~93/2~13 P~T/U~93/~4239 group. Other benzoylating agents (e.q., benzoic anhydride, benzoyl chloride) are not as selective as the chosen benzoylating agent, and yield N-6-benzoylated derivatives, as well as the desired derivative.
The three step procedure of this invention provides high yields of 5'-0 and 3'-O-protection reactions. The procedures used are simple and require no chromatographic purific~tions. Thus, this three step synthesis is a useful improvement over existing methods and provides a high overall yield of about 59%. In addition, the method can be scaled up for synthesis of kilogram quantities of the desired co~pounds~
Thus, in this aspect, the invention features a method for rapid and selective synthesis of 3'-O-~enzoyl-2'-deoxyadenosine, a protected nucleoside intermediate suitable for modification at C-5' and/or N-~ position.
Specifically, as discussed above, the nucleoside is treated with a chemical which provides a protecting group at the 5'-OH group (e.q., TBDMS), and the remaining 3'-OH
g oup is benzoylated using benzoyl cyanide. The protecting group is then removed by treatment with a reagent such as tetrabutylammonium fluoride (TBAF) or its equivalent.
Other features and advantages of the invention will be apparent from the following description of the .
preferred embodiments thereof, and from the claims.
Descri~tion of the Preferred Embodiments The drawings will first briefly be described.
Drawinas:
Fig. 1 is a diagrammatic representation of the chemical synthesis of ddC from uridine by a method of this invention.
Fig. 2 is a diagrammatic representation of the synthesis of 2',3'-dideoxyguanosine.
Fig. 3 is a diagrammatic representation of a method of this invention for synthesis of 2',3'-dideoxyguanosine.
W093/2~13 ~1 3 '~J ~ ~ ~ PCr/~Sg3/Q423~
Fig. 4 is a diagrammatic representation of a method of this invention for synthesis of 2',3'-dideoxyadenosine and 2',3'-dideoxyinosine.
Fig. 5 is a diagrammatic representation of a method for synthesis of 3'-0-Benzoyl-2'-deoxyadenosine.
Synthesis:
The following are specific examples of various aspects of this invention. Those in the art will recognize that equivalent methods within the claims can be readily devised and that the examples are not limiting in the invention.
Exam~le 1: 2' 3'-DideoxYcytidine Referring to Fig. 1, in general, 2'-deoxyuridine is formed from uridine by a method similar to that of Haoqiang and Chu, su pra, and then converted to ddC by the steps shown.
Referring to Fig. 1, uridine (60 g) was treated with acetyl bromide (with or without HBr) in acetonitrile to give 3',5'-di-O-acetyl-2'-bromo-2'-deoxyuridine. The crude material was deoxygenated using tributyltin hydride in the presence of benzoyl peroxide (or 2,2'-azobis-(2-methyl)propionitrile, or azobisisobutyronitrile) to give, after deacetylation with methanolic ammonia, 2'-deoxyuridine in 70% yield. Selective silylation with t-butyldimethylsilyl (TBDMS; triisopropylsilyl chloride ort-butyldiphenylsilyl chloride can be used in place of- - ~~
TBDMS) chloride in pyridine yielded 5'-silylated-2'-deoxyuridine as a crystalline solid in 77% yield. This compound was treated with phenylchloro~hionoformat~ (or 1,1'-thiocarbonyldiimidazole) to give 5'-0-TBDMS-3'-O-phenoxythiocarbonyl-2'-deoxyuridine in 86~ yield. Burto~
d~oxygenation of the above derivative with tributyltin hydride (or tributyltin chloride and sodium borohydride) yielded 5'-silylated-2',3'-dideoxyuridine in 77% yield.
Reaction of this compound with 1,2,4-triazole and phosphorusoxychloride followed by ammonia treatment afforded 5'-silylated-2'3'-dideoxycytidine in 88% yield.
l93/2~13 ? 1 ~ - 4 (') '~ PCT/US93/04239 Cleavage of the 5'-silyl group using ion-exchange resin Amberlyst A-26 (F ) (a macroreticular quaternary ammonium resin, 20-60 mesh particle size) in refluxing toluene gave 2',3'-dideoxycytidine (ddC) in 99% yield. The overall yield (from uridine) was 31%.
Example 2: 2' 3'-DideoxY~uanosine Referring to Fig. 2, the following is an example of a method for synthesis of 2',3'-dideoxyguanosine.
The synthesis of N~-isobutyryl-2',3'-dideoxy-2',3'-didehydroguanosine was formed as follows.
N2-isobutyrylguanosine 6,74 g (20 mM) was suspended in 80 ml of dry acetonitrile. 0.4 ml water was added followed by acetoxyisobutyryl bromide and the mixture stirred -by room temperature (about 24C) for 1 hour. The reaction mix~ure was filtered, dilu~ed with 300 ml of ethylacetate and added to cold saturated NaHCO3 solution. The org~nic layer was separated and washed by water (70 ml x 2). The combined water extracts were washed by ethylacetate (2 x 250 ml). The organic extracts were combined, dried (MgSO4), evaporated to 50 ml, diluted with dry methanol (50 ml), and evaporated to 50 ml; this process was xepeated twice. The remaining solution (50 ml) was added to freshly prepared Zn/Cu couple (from 4.4 g of Cu(OAc)2 and 26.14 of Zn powder according to Mansuri,- ~ ~ 25 54 J. Ora. Chem. 4780, 1989) under vigorous stirring.
After 5 minutes, the reaction mixture was filtered, the filter cake washed with methanol (2 x 75 ml), the filtrate evaporated to dryness, and the residue partitioned between 250 ml of ethylacetate and 70 ml of water. The organic = 30 layer was separated, dried, evaporated, dissolved in t00 ml of dry methanol and treated by 20 mM MeONa in methanol.
After 10 minutes the reaction was quenched by AcOH, evaporated and purified by flash chromatography in CHCl3-CHCl3-methanol (9:1). 2.4 g of N2-isobutyryl-2',3'-dideoxy-2',3'-didehydroguanosine (40%) was isolated as a solid.
2',3'-dideoxyguanosine was formed as follows:
W O 93/23413 ~ P ~ /VS93/04239 2.6 g of N2-isobutyryl-2',3'-dideoxy-2',3'-didehydroguanosine (8.125 mM) in methanol (250 mml) was hydrogenated at 32 psi in the presence of 10% Pd/C (1.3 g) for five hours. The catalyst was filtered and the solvent evaporated. The purification of the crude product by flash chromatography using CHCl3 methanol (10:1~ and subsequent ammonolysis yielded 2.0 g (77%) of 2',3'-dideoxyguanosine as a solid.
Example 3: 2',3'-DideoxYquanosine ~;
Referring to Fig. 3, the synthesis of N~-isobutyryl-6-O-[tp-nitrophenyl~ethyl3-3'-O-mesyl-5'-O-tert-butyldiphenylsilyl-2'-deoxyguanosine was as follows.
N2-isobutyryl-2'-deoxyguanosine 12.9 g (40 mM) dried by coevaporation with pyridine (2 x 200 ml~ was dissolved in dry pyridine ~200 ml) and tert-butyl diphenylsilylchloride 12.00 ml (45.46 mM, 1.1~ equivalents (eq)) was added by use of a syringe over a 10 minute period. The reaction mixture was kept at room temperature (about 24C) for 24 hours, with exclusion of moisture, and 10 ml of methanol added. After an additional 30 minutes the mixture was evaporated to dryness. The residue was coevaporated with toluene (2 x 75 ml), dissolved in chloroform (400 ml), washed with water (2 x 100 ml), dried (MgSO4), evaporated, and coevaporated with dry dioxane (100 ml). The remaining oil was suspended in dry dioxane - -- -(250 ml) and triphenylphosphine 15.73 g (60 mM, 1.5 eq), 2-(4-nitrophenyl~ethanol 10.03 g (60 mM, 1.5 eq), followed by diethyl azodicarboxylate (DEAD) 9.44 ml (60 mM, 1.5 eq) which was added under argon over 20 minutes (slightly exothermic). The reaction mixture was left at room temperature for 4.5 hours, evaporated to about 100 ml, and dry pyridine (100 ml) and methanesulfonyl chloride 9.35 ml (120 mM, 3 e~ added under argon. The reaction mixture was left at the room temperature overnight. Methanol (30 ml) was added and after 1 hour the reaction mixture was evaporated to dryness, and coevaporated with toluene. The residue was dissolved in chloroform (500 ml), washed with 93/23413 ~'~ 3 S ~ 9 ~ PCT/US93/04239 water (3 x loO ml), dried (MgSO4~, evaporated ~nd purified by flash chromatography on silica gel (800 g) in CHCl3-2%
MeOH. N2-isobutyryl-6-O-[(p-nitrophenyl)ethyl]-3'-O-mesyl-5'-0-tert-butyldiphenylsilyl-2'-deoxyguanosine was obtained as a solid in 80% yield.
N2- isobutyryl-2',3'-dideoxy-2',3'-didehydroguanosine was prepared as follows.
N2-isobutyryl-6-O-t(p-nitrophenyl)ethyl)-3'-O-mesyl-5'-0-tert-butyldiphenylsilyl-2'-deoxyguanosine 4.77 g (6 mM) was dissolved in 60 ml of dry N,N-dimethylformamide and 1.62 g (30 mM, 5 eq) sodium methanolate was added. After 1 hour 50 ml methanol was added, and the reaction mixture neutralized by Amberlite IRC 50 (H'3. The resin was filtered, washed by methanol (2 x 25 ml), the filtrate evaporated and partitioned between water (250 ml) and ether (200 ml). The water layer was separated, concentrated to 50 ml and applied on XAD -2 column (200 ml) which was washed with water (800 ml) and eluted with 55% methanol. N2-isobutyryl-2',3'-dideoxy-2',3'-didehydroguanosine 1.26 g (65.6%) was isolated as a solid.
- N2-isobutyryl-2',3'-dideoxyguanosine was prepared as follows.
_ 2.6 g (8.125 mM) N2-isobutyryl-2',3'-dideoxy-~- - 25 2',3'-didehydroguanosine in methanol (250 mml) was hydrogenated at 32 psi in the presence of 10% Pd/C(1.3 g) for 5 hours. The catalyst was filtered off and the - -- solvent evaporated. The purification of the crude product by ~lash chromatography using CHCl3tmethanol (10:1) yielded _ 30 2.0 g (77%) of the title compound as a solid.
Subsequent ammonolysis (NH3/MeOH) of this material led to 2',3'-dideoxyguanosine in 90% yield.
Exam~le 4: DideoxYadenosine and Dideoxyinosine Referring to Fig. 4, dideoxyadenosine and dideoxyinosine can be formed in a similar manner with similar yields to that described in Example 3. In contrast to Example 3, however, there is no need to treat W093/2~13 ~ ~ PCT/US93/0423~
the starting material with triphenylphosphine 2-(4-nitrophenyl)ethanol and diethyl azodicarboxylate after the silylchloride treatment.
The synthesis of 3',0-tosyl-5'-0-t-S butyldiphenylsilyl-2'-deoxyadenosine was performed as follows: ~-10.4 g (40 mM) 2'-deoxyadenosine, dried by coevaporation with pyridine (2 x 100 ml), was dissolved in dry pyridine (100 ml) and 12.66 ml (48 mM, 1,2 eq) -~
t-butyldiphenylsilylchloride added. The mixture was stirred at room temperature (about 24C) for 24 hours.
The reaction mixture was cooled to 0-5C, and 10.0 g (52 mM, 1.3 eq) p-toluenesulfonyl chloride added with ;
additional pyridine (50 ml). The reaction mixture was i kept for 2 hours at 0C, then 24 hours at room temperature (about 24C), with exclusion of moisture, and 30 ml methanol added. After an additional 30 minutes, the ¦ -reaction mixture was evaporated to dryness and coevaporated with toluene. The residue was dissolved in ! -chlorofo~m (500 ml), washed with water (3 x 100 ml), dried (MgSO4), evaporated and purified by flash chromatography on silica gel (800 g~ in CHCl3- 2% MeOH. 3',0-tosyl-5'-0-t-butyldiphenylsilyl-2'-deoxyadenosine was obtained as a- ¦
solid in 85% yield.
2~,3'-dideoxy-2',3~-didehydroadenosine -was~
prepared as follows: - -9.3 g (1~.37 mM) 3',0-tosyl-5'-0-t-butyldiphenylsilyl-2'-deoxyadenosine was dissolved in 150 ml dry N,N-dimethylformamide, and 3.78 g (70 mM, 5 èq) 30 sodium methanolate added. After 1 hour 30 ml methanol was --~
added, and the reaction mixture neutralized by Amber~ite~~-~
IRC 50 (H~). The resin was filtered, washed with methanol ~2 x 50 ml), the filtrate evaporated and partitioned betw~en water (300 ml) and CHC13 (100 ml). The water layer was separated, concentrated to 50 ml and applied on XAD-2 column (300 ml~, which was washed with water (lL) and ~93/2~13 ~1 3 ~ 3 PCT/US93/04239 eluted with 30% methanol. 2.5 g (73.4%) of 2',3'-dideoxy-2',3'-didehydroadenosine was isolated as a solid.
Subsequent hydrogenation of 2',3'-dideoxy-2',3'-didehydroadenosine in methanol at 35 psi of H2 in the presence of 10% Pd/C for 6 hours led to 2',3'-dideoxyadenosine in 80% yield.
The preparation of 2',3'-dideoxyinosine is similar to the above procedure for 2',3'-dideoxyadenosine, with comparable yields.
Example 5: 3'-0-Benzoyl-2'-deoxyadenosine As discussed above, the invention generally features a three step procedure which can be performed without chromatographic purification steps.
2'-Deoxyadenosine (10 g) was dissolved in dry pyridine under reflux, the solution cooled to room temperature (about 24CC) and t-butyldimethylsilylchloride (TBDMSCl; 1.1 equivalents (eq.)) added (t-butyldiphenyl silyl-or triisopropylsilyl chloride could also be used).
Precipitate was formed, and after 1 hour of stirring at room temperature, water and chloroform were added and the mixture filtered. The precipitate was washed successively - with chloroform and water and dried to give 5'-0-TBDMS-2'-deoxyadenosine in 85% yield.
The above product was suspended in acetonitrile- 25 (or dimethylformamide) and benzoyl cyanide (1.1 eq.) added. Triethylamine (5 ml) was added in 5 portions to dissolve all the material. ~fter 20 minutes of stirring a heavy precipitate formed which was filtered off, and washed with acetonitrile. Recrystallization from acetonitrile yielded 3'-0-benzoyl-5'-0-TBDMS-2'-deoxyadenosine in 75% yield.
The above material was suspended in dry tetrahydrofuran (TKF) and 1 M tetrabutylammonium fluoride (TBAF~ in THF added (~mberlyst A-26 (F-) (an ion-exchange resin, speci~ically a mazroreticular quaternary ammonium resin, 2~-60 mesh particle size) in toluene could also be used). The solution was stirred for 1 hour at room WO93/2~13 ~ PCT~US93/04234 temperature and concentrated in vacuo to a syrup.
Methanol was added and the precipitate collected and washed with methanol to give 3'-O-benzoyl-2'-deoxyadenosine in 93% yield. Thus, the overall yield in ~ .
this 3 step reaction was S9%.
Other em~odiments are within the followingclaims.
5463, 1977, describe the synthesis of dideoxynucleosides and use of their 5'-triphosphates for DNA sequencing.
Herdewijn 31 J. Med. Chem. 2040, 1988 describes synthesis of 2',3'-dideoxynucleosides and their anti-HIV activity.
Herdewijn, 31 J. Med. Chem. 2040, 1988, describes synthesis o~ 2',3'-dideoxynucleosides and their anti-HIV activity. McCarthy et al., 88:7 J.A.C.S. 1549, lg66, describe synthesis of adenine nucleosides. Jain et al., 39 J. Orq. Chem. 30-, -1974, describe synthesis of 2',3'-dideoxy nucleosides. Chu, 54 J. Orq. Chem. 2217, 1989, describes the general synthesis of 2',3'-dideoxynucleosides. Sanger et al., 74 Proc. Natl. Acad.
Sci. USA 5463, 1977, describe the synthesis of dideoxyguanosine.
Holy, 1 Synthetic Procedures in Nucleic Acid ChemistrY 172, 1968, describes a method for synthesis of 3'-O-acetyl-2'-deoxyadenosine from 2'-deoxyadenosine in four steps and 45% overall yield; Ogilvie, 51 Can.~_J.
Chem. 3799, '973, describes a method for synthesis o~ 3i-O-benzoyl-2'-deoxyadenosine from 2'-deoxyadenosine in four steps and 71~ overall yield, and alternatively in 3- steps- -and 38~ overall yield~ Applicant was told that it is-~
preferred to use benzoyl cyanide for selective benzoylation of deoxynucleosides.
Summary of the Invention -In a first aspect, this invention features an improved economical sy~thetic method for the preparation of dideoxyuridine (ddU) which is then converted- lnto dideoxycytidine (ddC) via a 4-triazolyl intermediate. The method is not only cost efficient, but can be scaled up to several hundred gram quantities. The method generally utilizes inexpensive uridine as a starting material which 93/2~13 ~ ;) 8 PCT/US93/04239 is converted in a 7 step reaction sequence to ddC with a yield of about 30~.
The ddC can be used for chemical synthesis of sugar-modified nucleotides; chemical synthesis of DNA
chain terminators; and chemical synthesis of anti-HIV
2',3'-dideoxynucleosides.
Specifically, in this aspect, the invention features a method for chemical synthesis of 2',3'-dideoxycytidine in which 5'-silylated-2',3'-deoxyuridine is used to form ddC, e.q., by reaction with l,2,4-triazole, a phosphoro~s oxychloride, followed by treatment with ammonia, and the silyl group of the resultins product cleaved by use of an ion-exchange resin (specifically, that referred to as the Amberlyst A-26 (F )) in toluene.
In a second aspectl the invention features a method for synthesis of dideoxyguanosine by directly converting N2-isobutyrylguanosine to a 2',3'-unsaturated derivative by action of acetoxyisobutyryl bromide in moist acetonitrile, with subsequent reductive elimination by Zn/Cu/DMF and deacylation by ~aOMe/methanol. Catalytic hydrogenation and ammonolysis of the N2-isobutyryl-2',3'-didehydroguanosine provides 2',3'-dideoxyguanosine in 30%
yield.
The method of this invention utiliæes N2-isobutyrylguanosine as a starting compound, and thusovercomes problems with isolation of unsaturated compounds. The described method is cost efficient and can be scaled up to multi-gram ~uantity.
-Thus, in this aspect, the invention features a method for chemical synthesis of 2',3'-dideoxyguanosine, including contacting N2-isobutyrylguanosine with ace~oxyisobutyryl bromide in acetonitrile with subsequent reductive elimination, deacylation, hydrogenation and ammonolysis.
In a third aspect, the invention features a convenient "one-pot" (i.e., without purification of intermediates~ synthesis of key intermediates: 5'-O-tert-`
G1~3~
W093/234]3 PCT/US~3/04239 butyldiphenylsilyl-3'-O~mesyl(tosyl)-2'-deoxyadenosine, 5'-O-tert-butyldiphenylsilyl-3'-O-mesyl(tosyl)-2'-deoxyinosine, and N2 isobutyryl~O6[(p-nitrophenyl)ethyl]-5'-O-tert-butyldiphenylsilyl-3'-O-mesyl(tosyl)-2'-deoxyguanosine, and their conversion to 2',3'-unsaturated derivatives by simultaneous elimination-deblocking procedures, and subsequent hydrogenation to 2',3'-dideoxynucleosides.
The method of this invention utilizes selective protection of the 5'-OH group of 2'-deoxyadenosine (or 2'-deoxyinosine) and N2-isobutyryl-2'-deoxyguanosine by introducing a tert-butyldiphenylsilyl protecting group.
The high selectivity of this step allows the preparation of the above key compounds in a "one-pot" procedure without isolation of intermediates. Simultaneous removal of the 5'-Q-tert-butyldiphenylsilyl group during the elimination step reduces the total number of steps and gives a high yield of 2',3'-unsaturated nucleosides. The ,-described methods are cost efficient, and can be scaled up to multi-gram quantity.
Thus, in this aspect, the invention features a method for chemical synthesis of 2',3'-dideoxyadenosine, 2',3'-dideoxyinosine, and 2',3'-dideoxyguanosine wlthout purification of intermediate compounds by sequentially treating: (a) N2-isobutyryl 2'-deoxyguanosine with a t- -- -butyl-diphenylsilylchloride, triphenylphosphine, diethyl--- -azodicarboxylate, p-nitrophenylethanol and methanesulfonyl (or p-toluenesulfonyl) chloride; or (b) by treating 2'-deoxyadenosine or 2'-deoxyinosine sequentially witX -t-butyldiphenylsilylchloridet and methanesulfonyl (or p-toluenesulfonyl) chloride.
In a fourth aspect, the invention features a method for synthesis of 3'-O-benzoyl-2'-deoxyadenosine based on the efficient protection of the 5'-hydroxyl group of 2'-deoxyadenosine, using stable t butyldimethylsilyl ether (TBDMS) protection. Acylation with benzoyl cyanide occurs exclusively at the remaining free sugar hydroxyl 7 d~ ~ 8 ~93/2~13 P~T/U~93/~4239 group. Other benzoylating agents (e.q., benzoic anhydride, benzoyl chloride) are not as selective as the chosen benzoylating agent, and yield N-6-benzoylated derivatives, as well as the desired derivative.
The three step procedure of this invention provides high yields of 5'-0 and 3'-O-protection reactions. The procedures used are simple and require no chromatographic purific~tions. Thus, this three step synthesis is a useful improvement over existing methods and provides a high overall yield of about 59%. In addition, the method can be scaled up for synthesis of kilogram quantities of the desired co~pounds~
Thus, in this aspect, the invention features a method for rapid and selective synthesis of 3'-O-~enzoyl-2'-deoxyadenosine, a protected nucleoside intermediate suitable for modification at C-5' and/or N-~ position.
Specifically, as discussed above, the nucleoside is treated with a chemical which provides a protecting group at the 5'-OH group (e.q., TBDMS), and the remaining 3'-OH
g oup is benzoylated using benzoyl cyanide. The protecting group is then removed by treatment with a reagent such as tetrabutylammonium fluoride (TBAF) or its equivalent.
Other features and advantages of the invention will be apparent from the following description of the .
preferred embodiments thereof, and from the claims.
Descri~tion of the Preferred Embodiments The drawings will first briefly be described.
Drawinas:
Fig. 1 is a diagrammatic representation of the chemical synthesis of ddC from uridine by a method of this invention.
Fig. 2 is a diagrammatic representation of the synthesis of 2',3'-dideoxyguanosine.
Fig. 3 is a diagrammatic representation of a method of this invention for synthesis of 2',3'-dideoxyguanosine.
W093/2~13 ~1 3 '~J ~ ~ ~ PCr/~Sg3/Q423~
Fig. 4 is a diagrammatic representation of a method of this invention for synthesis of 2',3'-dideoxyadenosine and 2',3'-dideoxyinosine.
Fig. 5 is a diagrammatic representation of a method for synthesis of 3'-0-Benzoyl-2'-deoxyadenosine.
Synthesis:
The following are specific examples of various aspects of this invention. Those in the art will recognize that equivalent methods within the claims can be readily devised and that the examples are not limiting in the invention.
Exam~le 1: 2' 3'-DideoxYcytidine Referring to Fig. 1, in general, 2'-deoxyuridine is formed from uridine by a method similar to that of Haoqiang and Chu, su pra, and then converted to ddC by the steps shown.
Referring to Fig. 1, uridine (60 g) was treated with acetyl bromide (with or without HBr) in acetonitrile to give 3',5'-di-O-acetyl-2'-bromo-2'-deoxyuridine. The crude material was deoxygenated using tributyltin hydride in the presence of benzoyl peroxide (or 2,2'-azobis-(2-methyl)propionitrile, or azobisisobutyronitrile) to give, after deacetylation with methanolic ammonia, 2'-deoxyuridine in 70% yield. Selective silylation with t-butyldimethylsilyl (TBDMS; triisopropylsilyl chloride ort-butyldiphenylsilyl chloride can be used in place of- - ~~
TBDMS) chloride in pyridine yielded 5'-silylated-2'-deoxyuridine as a crystalline solid in 77% yield. This compound was treated with phenylchloro~hionoformat~ (or 1,1'-thiocarbonyldiimidazole) to give 5'-0-TBDMS-3'-O-phenoxythiocarbonyl-2'-deoxyuridine in 86~ yield. Burto~
d~oxygenation of the above derivative with tributyltin hydride (or tributyltin chloride and sodium borohydride) yielded 5'-silylated-2',3'-dideoxyuridine in 77% yield.
Reaction of this compound with 1,2,4-triazole and phosphorusoxychloride followed by ammonia treatment afforded 5'-silylated-2'3'-dideoxycytidine in 88% yield.
l93/2~13 ? 1 ~ - 4 (') '~ PCT/US93/04239 Cleavage of the 5'-silyl group using ion-exchange resin Amberlyst A-26 (F ) (a macroreticular quaternary ammonium resin, 20-60 mesh particle size) in refluxing toluene gave 2',3'-dideoxycytidine (ddC) in 99% yield. The overall yield (from uridine) was 31%.
Example 2: 2' 3'-DideoxY~uanosine Referring to Fig. 2, the following is an example of a method for synthesis of 2',3'-dideoxyguanosine.
The synthesis of N~-isobutyryl-2',3'-dideoxy-2',3'-didehydroguanosine was formed as follows.
N2-isobutyrylguanosine 6,74 g (20 mM) was suspended in 80 ml of dry acetonitrile. 0.4 ml water was added followed by acetoxyisobutyryl bromide and the mixture stirred -by room temperature (about 24C) for 1 hour. The reaction mix~ure was filtered, dilu~ed with 300 ml of ethylacetate and added to cold saturated NaHCO3 solution. The org~nic layer was separated and washed by water (70 ml x 2). The combined water extracts were washed by ethylacetate (2 x 250 ml). The organic extracts were combined, dried (MgSO4), evaporated to 50 ml, diluted with dry methanol (50 ml), and evaporated to 50 ml; this process was xepeated twice. The remaining solution (50 ml) was added to freshly prepared Zn/Cu couple (from 4.4 g of Cu(OAc)2 and 26.14 of Zn powder according to Mansuri,- ~ ~ 25 54 J. Ora. Chem. 4780, 1989) under vigorous stirring.
After 5 minutes, the reaction mixture was filtered, the filter cake washed with methanol (2 x 75 ml), the filtrate evaporated to dryness, and the residue partitioned between 250 ml of ethylacetate and 70 ml of water. The organic = 30 layer was separated, dried, evaporated, dissolved in t00 ml of dry methanol and treated by 20 mM MeONa in methanol.
After 10 minutes the reaction was quenched by AcOH, evaporated and purified by flash chromatography in CHCl3-CHCl3-methanol (9:1). 2.4 g of N2-isobutyryl-2',3'-dideoxy-2',3'-didehydroguanosine (40%) was isolated as a solid.
2',3'-dideoxyguanosine was formed as follows:
W O 93/23413 ~ P ~ /VS93/04239 2.6 g of N2-isobutyryl-2',3'-dideoxy-2',3'-didehydroguanosine (8.125 mM) in methanol (250 mml) was hydrogenated at 32 psi in the presence of 10% Pd/C (1.3 g) for five hours. The catalyst was filtered and the solvent evaporated. The purification of the crude product by flash chromatography using CHCl3 methanol (10:1~ and subsequent ammonolysis yielded 2.0 g (77%) of 2',3'-dideoxyguanosine as a solid.
Example 3: 2',3'-DideoxYquanosine ~;
Referring to Fig. 3, the synthesis of N~-isobutyryl-6-O-[tp-nitrophenyl~ethyl3-3'-O-mesyl-5'-O-tert-butyldiphenylsilyl-2'-deoxyguanosine was as follows.
N2-isobutyryl-2'-deoxyguanosine 12.9 g (40 mM) dried by coevaporation with pyridine (2 x 200 ml~ was dissolved in dry pyridine ~200 ml) and tert-butyl diphenylsilylchloride 12.00 ml (45.46 mM, 1.1~ equivalents (eq)) was added by use of a syringe over a 10 minute period. The reaction mixture was kept at room temperature (about 24C) for 24 hours, with exclusion of moisture, and 10 ml of methanol added. After an additional 30 minutes the mixture was evaporated to dryness. The residue was coevaporated with toluene (2 x 75 ml), dissolved in chloroform (400 ml), washed with water (2 x 100 ml), dried (MgSO4), evaporated, and coevaporated with dry dioxane (100 ml). The remaining oil was suspended in dry dioxane - -- -(250 ml) and triphenylphosphine 15.73 g (60 mM, 1.5 eq), 2-(4-nitrophenyl~ethanol 10.03 g (60 mM, 1.5 eq), followed by diethyl azodicarboxylate (DEAD) 9.44 ml (60 mM, 1.5 eq) which was added under argon over 20 minutes (slightly exothermic). The reaction mixture was left at room temperature for 4.5 hours, evaporated to about 100 ml, and dry pyridine (100 ml) and methanesulfonyl chloride 9.35 ml (120 mM, 3 e~ added under argon. The reaction mixture was left at the room temperature overnight. Methanol (30 ml) was added and after 1 hour the reaction mixture was evaporated to dryness, and coevaporated with toluene. The residue was dissolved in chloroform (500 ml), washed with 93/23413 ~'~ 3 S ~ 9 ~ PCT/US93/04239 water (3 x loO ml), dried (MgSO4~, evaporated ~nd purified by flash chromatography on silica gel (800 g) in CHCl3-2%
MeOH. N2-isobutyryl-6-O-[(p-nitrophenyl)ethyl]-3'-O-mesyl-5'-0-tert-butyldiphenylsilyl-2'-deoxyguanosine was obtained as a solid in 80% yield.
N2- isobutyryl-2',3'-dideoxy-2',3'-didehydroguanosine was prepared as follows.
N2-isobutyryl-6-O-t(p-nitrophenyl)ethyl)-3'-O-mesyl-5'-0-tert-butyldiphenylsilyl-2'-deoxyguanosine 4.77 g (6 mM) was dissolved in 60 ml of dry N,N-dimethylformamide and 1.62 g (30 mM, 5 eq) sodium methanolate was added. After 1 hour 50 ml methanol was added, and the reaction mixture neutralized by Amberlite IRC 50 (H'3. The resin was filtered, washed by methanol (2 x 25 ml), the filtrate evaporated and partitioned between water (250 ml) and ether (200 ml). The water layer was separated, concentrated to 50 ml and applied on XAD -2 column (200 ml) which was washed with water (800 ml) and eluted with 55% methanol. N2-isobutyryl-2',3'-dideoxy-2',3'-didehydroguanosine 1.26 g (65.6%) was isolated as a solid.
- N2-isobutyryl-2',3'-dideoxyguanosine was prepared as follows.
_ 2.6 g (8.125 mM) N2-isobutyryl-2',3'-dideoxy-~- - 25 2',3'-didehydroguanosine in methanol (250 mml) was hydrogenated at 32 psi in the presence of 10% Pd/C(1.3 g) for 5 hours. The catalyst was filtered off and the - -- solvent evaporated. The purification of the crude product by ~lash chromatography using CHCl3tmethanol (10:1) yielded _ 30 2.0 g (77%) of the title compound as a solid.
Subsequent ammonolysis (NH3/MeOH) of this material led to 2',3'-dideoxyguanosine in 90% yield.
Exam~le 4: DideoxYadenosine and Dideoxyinosine Referring to Fig. 4, dideoxyadenosine and dideoxyinosine can be formed in a similar manner with similar yields to that described in Example 3. In contrast to Example 3, however, there is no need to treat W093/2~13 ~ ~ PCT/US93/0423~
the starting material with triphenylphosphine 2-(4-nitrophenyl)ethanol and diethyl azodicarboxylate after the silylchloride treatment.
The synthesis of 3',0-tosyl-5'-0-t-S butyldiphenylsilyl-2'-deoxyadenosine was performed as follows: ~-10.4 g (40 mM) 2'-deoxyadenosine, dried by coevaporation with pyridine (2 x 100 ml), was dissolved in dry pyridine (100 ml) and 12.66 ml (48 mM, 1,2 eq) -~
t-butyldiphenylsilylchloride added. The mixture was stirred at room temperature (about 24C) for 24 hours.
The reaction mixture was cooled to 0-5C, and 10.0 g (52 mM, 1.3 eq) p-toluenesulfonyl chloride added with ;
additional pyridine (50 ml). The reaction mixture was i kept for 2 hours at 0C, then 24 hours at room temperature (about 24C), with exclusion of moisture, and 30 ml methanol added. After an additional 30 minutes, the ¦ -reaction mixture was evaporated to dryness and coevaporated with toluene. The residue was dissolved in ! -chlorofo~m (500 ml), washed with water (3 x 100 ml), dried (MgSO4), evaporated and purified by flash chromatography on silica gel (800 g~ in CHCl3- 2% MeOH. 3',0-tosyl-5'-0-t-butyldiphenylsilyl-2'-deoxyadenosine was obtained as a- ¦
solid in 85% yield.
2~,3'-dideoxy-2',3~-didehydroadenosine -was~
prepared as follows: - -9.3 g (1~.37 mM) 3',0-tosyl-5'-0-t-butyldiphenylsilyl-2'-deoxyadenosine was dissolved in 150 ml dry N,N-dimethylformamide, and 3.78 g (70 mM, 5 èq) 30 sodium methanolate added. After 1 hour 30 ml methanol was --~
added, and the reaction mixture neutralized by Amber~ite~~-~
IRC 50 (H~). The resin was filtered, washed with methanol ~2 x 50 ml), the filtrate evaporated and partitioned betw~en water (300 ml) and CHC13 (100 ml). The water layer was separated, concentrated to 50 ml and applied on XAD-2 column (300 ml~, which was washed with water (lL) and ~93/2~13 ~1 3 ~ 3 PCT/US93/04239 eluted with 30% methanol. 2.5 g (73.4%) of 2',3'-dideoxy-2',3'-didehydroadenosine was isolated as a solid.
Subsequent hydrogenation of 2',3'-dideoxy-2',3'-didehydroadenosine in methanol at 35 psi of H2 in the presence of 10% Pd/C for 6 hours led to 2',3'-dideoxyadenosine in 80% yield.
The preparation of 2',3'-dideoxyinosine is similar to the above procedure for 2',3'-dideoxyadenosine, with comparable yields.
Example 5: 3'-0-Benzoyl-2'-deoxyadenosine As discussed above, the invention generally features a three step procedure which can be performed without chromatographic purification steps.
2'-Deoxyadenosine (10 g) was dissolved in dry pyridine under reflux, the solution cooled to room temperature (about 24CC) and t-butyldimethylsilylchloride (TBDMSCl; 1.1 equivalents (eq.)) added (t-butyldiphenyl silyl-or triisopropylsilyl chloride could also be used).
Precipitate was formed, and after 1 hour of stirring at room temperature, water and chloroform were added and the mixture filtered. The precipitate was washed successively - with chloroform and water and dried to give 5'-0-TBDMS-2'-deoxyadenosine in 85% yield.
The above product was suspended in acetonitrile- 25 (or dimethylformamide) and benzoyl cyanide (1.1 eq.) added. Triethylamine (5 ml) was added in 5 portions to dissolve all the material. ~fter 20 minutes of stirring a heavy precipitate formed which was filtered off, and washed with acetonitrile. Recrystallization from acetonitrile yielded 3'-0-benzoyl-5'-0-TBDMS-2'-deoxyadenosine in 75% yield.
The above material was suspended in dry tetrahydrofuran (TKF) and 1 M tetrabutylammonium fluoride (TBAF~ in THF added (~mberlyst A-26 (F-) (an ion-exchange resin, speci~ically a mazroreticular quaternary ammonium resin, 2~-60 mesh particle size) in toluene could also be used). The solution was stirred for 1 hour at room WO93/2~13 ~ PCT~US93/04234 temperature and concentrated in vacuo to a syrup.
Methanol was added and the precipitate collected and washed with methanol to give 3'-O-benzoyl-2'-deoxyadenosine in 93% yield. Thus, the overall yield in ~ .
this 3 step reaction was S9%.
Other em~odiments are within the followingclaims.
Claims (24)
1. Method for synthesis of 2',3'-dideoxycytidine comprising the step of providing 5'-silylated-2',3'-deoxyuridine as an intermediate in the process for formation of said 2',3'-dideoxycytidine.
2. The method of claim 1, wherein said 5'-silylated-2',3'-dideoxyuridine is reacted with 1, 2, 4-triazole and phosphorusoxychloride and ammonia to yield a 5'-silylated-2',3'-dideoxycytidine.
3. The method of claim 2, wherein the 5'-silyl group of said 5'-silylated-2',3'-dideoxycytidine is cleaved using an ion exchange resin.
4. The method of claim 3, wherein said ion exchange resin is a macroreticular quaternary ammonium resin with 20-60 mesh particle site.
5. The method of claim 4, wherein said ion exchange resin is used together with toluene to yield 2',3'-dideoxycytidine.
6. Method for synthesis of 2',3'-dideoxyguanosine, comprising the step of contacting N2-isobutyrylguanosine with acetoxyisobutyryl bromide in moist acetonitrile.
7. The method of claim 6 further comprising the step of contacting the product of the step of claim 6 with a zinc/copper couple in methanol or N,N-dimethylphormamide.
8. The method of claim 7 further comprising the step of contacting the product of claim 7 with sodium methylate in methanol.
9. The method of claim 8 further comprising the step of hydrogenating the product of claim 8.
10. The method of claim 9 further comprising the step of ammonolysing the product of claim 9.
11. The method of claim 6 further comprising the steps of reductive elimination, deacylation, catalytic hydrogenation, and ammonolysis to provide said 2',3'-dideoxyguanosine.
12. Method for synthesis of 2',3'-dideoxyadenosine or 2',3'-dideoxyinosine without purification of intermediate compounds, comprising the steps of treating 2'-deoxyadenosine or 2'-deoxyinosine with a t-butyldiphenylsilylchloride, and an alkylsulfonyl chloride.
13. The method of claim 12 further comprising contacting the chemical mixture resulting after the step of claim 12 with sodium methanolate or potassium t-butanoate.
14. The method of claim 13 further comprising the step of hydrogenating the mixture resulting after the step of claim 13.
15. Method for synthesis of 2',3'-dideoxyguanosine without purification of intermediate compounds, comprising the steps of treating N2-isobutyryl-2 ' - d e o x y g u a n o s i n e s e q u e n t i a l l y w i t h t-butyldiphenylchlorosilane, triphenylphosphine, diethyl-azodicarboxylate, 2-(4-nitrophenyl)ethanol and an alkylsulfonyl chloride.
16. The method of claim 15 further comprising contacting the chemical mixture resulting after the step of claim 12 with sodium methanolate or potassium t-butanoate.
17. The method of claim 16 further comprising the step of hydrogenating the mixture resulting after the step of claim 16.
18. The method of claim 17 further comprising the step of ammonolysis of the mixture resulting after the step of claim 17.
19. Method for synthesis of a 3'-0 protected 2'-deoxyadenosine, comprising the steps of:
protecting the 5'-OH group of 2'-deoxyadenosine with a reagent selected from the group consisting of t - b u t y l d i m e t h y l s i l y l c h l o r i d e , t -butyldiphenylsilylchloride, t-butyltriphenylsilylchloride, and triisopropylsilylchloride, benzoylating the 3'-OH group of the protected 2'-deoxyadenosine with a benzoyl cyanide, and deprotecting the benzoylated 2'-deoxynucleoside to yield said 3'-O-benzoyl-protected 2'-deoxyribonucleoside.
protecting the 5'-OH group of 2'-deoxyadenosine with a reagent selected from the group consisting of t - b u t y l d i m e t h y l s i l y l c h l o r i d e , t -butyldiphenylsilylchloride, t-butyltriphenylsilylchloride, and triisopropylsilylchloride, benzoylating the 3'-OH group of the protected 2'-deoxyadenosine with a benzoyl cyanide, and deprotecting the benzoylated 2'-deoxynucleoside to yield said 3'-O-benzoyl-protected 2'-deoxyribonucleoside.
20. The method of claim 19, wherein said benzoylating is performed with benzoyl cyanide in acetonitrile or dimethylformamide.
21. The method of claim 19, wherein said deprotecting step comprises contacting with tetrahydrofuran and tetrabutylammonium fluoride or contacting with a macro reticular quaternary ammonium resin.
22. Method for synthesis of a 3'-0-protected-2'-deoxyadenosine, comprising the steps of:
protecting the 5'-OH group of 2'-deoxyadenosine with a trialkylsilyl chloride, benzoylating the 3'-OH group of the protected 2'-deoxyadenosine with a benzoyl cyanide, and deprotecting the benzoylated 2'-deoxynucleoside to yield said 3'-0-benzoyl-protected-2'-deoxyribonucleoside.
protecting the 5'-OH group of 2'-deoxyadenosine with a trialkylsilyl chloride, benzoylating the 3'-OH group of the protected 2'-deoxyadenosine with a benzoyl cyanide, and deprotecting the benzoylated 2'-deoxynucleoside to yield said 3'-0-benzoyl-protected-2'-deoxyribonucleoside.
23. The method of claim 22, wherein said benzoylating is performed with benzoyl cynaide in acetonitrile or dimethylformamide.
24. The method of claim 22, wherein said deprotecting step comprises contacting with tetrahydrofuran and tetrabutylammonium fluoride or contacting with a macro reticular quarternary ammonium resin.
Applications Claiming Priority (8)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| US88257292A | 1992-05-13 | 1992-05-13 | |
| US07/882,572 | 1992-05-13 | ||
| US07/882,479 US5506349A (en) | 1992-05-13 | 1992-05-13 | Chemical synthesis of 2', 3'-dideoxycytidine |
| US07/882,479 | 1992-05-13 | ||
| US91064992A | 1992-07-08 | 1992-07-08 | |
| US07/910,649 | 1992-07-08 | ||
| US91954492A | 1992-07-24 | 1992-07-24 | |
| US07/919,544 | 1992-07-24 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CA2135498A1 true CA2135498A1 (en) | 1993-11-25 |
Family
ID=27505972
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CA 2135498 Abandoned CA2135498A1 (en) | 1992-05-13 | 1993-05-06 | Synthesis of nucleotide monomers |
Country Status (6)
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|---|---|
| EP (1) | EP0640091A4 (en) |
| JP (1) | JPH07508022A (en) |
| AU (1) | AU4234193A (en) |
| CA (1) | CA2135498A1 (en) |
| MX (1) | MX9302761A (en) |
| WO (1) | WO1993023413A1 (en) |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5627160A (en) * | 1993-05-25 | 1997-05-06 | Yale University | L-2',3'-dideoxy nucleoside analogs as anti-hepatitis B (HBV) and anti-HIV agents |
| TW374087B (en) * | 1993-05-25 | 1999-11-11 | Univ Yale | L-2',3'-dideoxy nucleotide analogs as anti-hepatitis B(HBV) and anti-HIV agents |
| EP1910393A2 (en) * | 2005-07-05 | 2008-04-16 | Hetero Drugs Limited | A novel process for the preparation of didanosine using novel intermediates |
| CN105884846B (en) * | 2016-06-01 | 2018-08-28 | 中南大学 | A kind of synthetic method of 2'-deoxyadenosine |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| US64631A (en) * | 1867-05-14 | Thomas g | ||
| US5099010A (en) * | 1983-01-17 | 1992-03-24 | Research Corporation | Intermediates in the preparation of 3'-amino-2',3'-dideoxycytidine and the pharmacologically acceptable salts thereof |
| US5084445A (en) * | 1986-05-01 | 1992-01-28 | University Of Georgia Research Foundation, Inc. | 3'-azido-2',3'-dideoxy-5-methylcytidine |
| JPS63275597A (en) * | 1987-05-07 | 1988-11-14 | Ajinomoto Co Inc | Production of dideoxycytidine, intermediate therefore and production thereof |
| US4987224A (en) * | 1988-08-02 | 1991-01-22 | University Of Georgia Research Foundation, Inc. | Method of preparation of 2',3'-dideoxynucleosides |
-
1993
- 1993-05-06 WO PCT/US1993/004239 patent/WO1993023413A1/en not_active Application Discontinuation
- 1993-05-06 CA CA 2135498 patent/CA2135498A1/en not_active Abandoned
- 1993-05-06 JP JP6503628A patent/JPH07508022A/en active Pending
- 1993-05-06 AU AU42341/93A patent/AU4234193A/en not_active Abandoned
- 1993-05-06 EP EP93911072A patent/EP0640091A4/en not_active Withdrawn
- 1993-05-12 MX MX9302761A patent/MX9302761A/en unknown
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| Publication number | Publication date |
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| EP0640091A4 (en) | 1995-07-26 |
| WO1993023413A1 (en) | 1993-11-25 |
| MX9302761A (en) | 1994-05-31 |
| JPH07508022A (en) | 1995-09-07 |
| AU4234193A (en) | 1993-12-13 |
| EP0640091A1 (en) | 1995-03-01 |
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