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Bioorganic & Medicinal Chemistry 14 (2006) 955–959

Synthesis and biological evaluation of phosphonated
carbocyclic 2 0-oxa-3 0-aza-nucleosides
Ugo Chiacchio,a Daniela Iannazzo,b,* Anna Piperno,b,* Roberto Romeo,b
Giovanni Romeo,b Antonio Rescifinaa and Monica Saglimbenib
a
Dipartimento di Scienze Chimiche, Università di Catania, Viale Andrea Doria 6, Catania 95125, Italy
b
Dipartimento Farmaco-Chimico, Università di Messina, Via SS. Annunziata, Messina 98168, Italy
Received 29 July 2005; revised 31 August 2005; accepted 6 September 2005
Available online 5 October 2005

Abstract—The synthesis of carbocyclic 2 0 -oxa-3 0 -aza-nucleosides has been described, based on the 1,3-dipolar cycloaddition of a
new phosphonated nitrone with vinyl acetate followed by coupling with silylated nucleobases. The obtained compounds have been
evaluated for their ability to inhibit the reverse transcriptase of avian myeloblastosis retrovirus: no significant activity has been
observed. 
2005 Elsevier Ltd. All rights reserved.

1. Introduction NRTIs may be considered as prodrugs; to become
active, these nucleosides must be phosphorylated by
Since the identification of HIV as the causative agent of cellular kinases to give successively the corresponding
AIDS more than 20 years ago, many efforts have been 5 0 -mono-, di-, and triphosphates. The efficacy of this
made to keep this disease under control and actually process is extremely low (e.g., 0.3% for AZT). Therefore,
19 compounds have been formally approved as anti- many efforts have been made to improve therapeutic
HIV drugs.1 properties by shortening this cascade and bypassing at
least the first phosphorylation step. This approach has
These therapeutic agents act at different steps of the resulted in the synthesis of numerous nucleotide ana-
viral replication cycle and belong to five different catego- logues (NtRTIs).5
ries: the nucleoside reverse transcriptase inhibitors
(NRTIs), the nucleotide reverse transcriptase inhibitors A nucleoside 5-phosphonate is essentially a nucleoside
(NtRTIs), the non-nucleoside reverse transcriptase monophosphate analogue. However, a phosphonate
inhibitors (NNRTIs), the protease inhibitors (PIs), and has the advantage over its phosphate counterpart of
fusion inhibitors (FI).2 being metabolically stable, as its phosphorus–carbon
bond is not susceptible to phosphatase hydrolysis.
Diverse strategies combining and alternating NRTIs, More importantly, the presence of a 5 0 -phosphonate
NNRTIs and PIs have been developed and are object moiety allows the first phosphorylation step required
of ongoing experimentation.3 for nucleoside activation to be skipped, therefore
bypassing this inefficient and often rate-limiting step
The combination of inhibitors of viral proteases and of in the conversion to 5 0 -triphosphate. Like a nucleoside
the RT determined a quantum leap in potency and monophosphate, a nucleoside phosphonate can be
efficacy of anti-HIV therapy in protocols of highly further phosphorylated by cellular nucleotide kinases.6
active anti-retroviral therapy (HAART).4 The concept of nucleoside phosphonate has been
applied to design chain terminators for anti-HIV
chemotherapy and proved to be valid. 9-(2-Phosphonyl-
Keywords: Modified nucleosides; 1,3-Dipolar cycloaddition; Antiviral methoxypropyl) adenine (PMPA) and 9-(2-phosphonyl-
activity, Phosphonated nucleosides. methoxyethyl) adenine (PMEA) are two effective and
* Corresponding authors. Tel.: +39 090 356230; fax: +39 090 potent nucleoside phosphonate chain terminators for
6766562; e-mail: apiperno@pharma.unime.it HIV reverse transcriptase (RT).7

0968-0896/$ - see front matter  2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2005.09.024

20 min. Med.95 Compounds of type 2 show low levels of cytotoxicity (E)-endo 248. Table 1. Nitrone 7 and vinyl acetate in excess were heated at 70 C for 12 h to give. According to these data. Conse- from the sugar moiety (spacer unit) between these two quently. 1). It is interesting to speculate that the biological effects The 1H NMR spectrum of the crude cycloaddition reac- exhibited by nucleoside analogues depend importantly tion shows the presence of two 5-epimeric isoxazolidines on the relative disposition of the phosphate or phospho. (c) CH3NHOHÆHCl. which ilar a-alkoxynitrones. these values agree has been prepared from the commercially available 3.85:1 ratio.551 58. Information on the stereo- Phosphonates 3 preserve the five atom chain length chemical outcome of the cycloaddition process was ten- between the heterocyclic base and the phosphonate tatively inferred by the aid of theoretical supports. they were promptly used for the successive key elements. satisfactorily with the results already reported for their chloro propanaldehyde diethyl acetal 5. 12h.326 39.and a-nu- some days. 100 W. decomposes into an unidentifiable mixture cleosides 3 and 4 were obtained (Scheme 2). in anti-retroviral and anti-HBV chemotherapy. The com- of products. The mixture of two cycloadducts was coupled with sily- nated aldehyde 6. we have described the potent reverse trans.3-dipolar nucleotide type 3 and 4 analogues of compound 1 where cycloaddition reaction was performed under microwave the 5 0 -oxygen atom is substituted by the isosteric meth.8 (E)-exo 248. Chem. we may assume that the major product of the cycloaddition reaction is com- The strategy of the synthetic approach is based on the pound 9 in agreement with the results reported for sim- construction of the new phosphonated nitrone 7. As a consequence. microwave irradiation. reflux.4 equiv of trimethylsilyltriflate (TMSOTf) as catalyst. The Arbuzov lower homologues. after proceeded with good yields: a mixture of b.9 nucleosidation reaction to give the corresponding more stable nucleotides 3 and 4. 95:5) and immediately used for the fur. the yields increased to 65% (Scheme 1). and 5-fluorouracil 12. In particular. moiety of natural nucleosides. Recently. Reagents and conditions: (a) (EtO)3P. 2 h.12 The condensation with silylated thymine in good yields (85%). in a 1. ther reactions. more reactive form of nitrone 7: the (E)-endo transition state leading to cis isomer is about 0.27 and exert. (b) HCl 10%. Nitrone 7 exists as a mixture of E/Z isomers (Z/E ratio.11 Moreover. (d) vinyl acetate. Nitrone 7 was purified through 10. 60 C. Chiacchio et al. The subsequent reaction with N-meth. we describe the synthesis of isoxazolidinyl yield. on RT from almost two different retroviruses. pounds 3c and 4c were further treated with 5% . AM1 calculations results on transition state structures for criptase (RT) inhibitors 2 which represent the lower isoxazolidines 8 and 9 phosphonate homologous of monophosphate com. it water in neutral and acidic conditions and rapidly seems to be important to preserve the distance generated decomposed into a complex mixture products.053 0.22 kcal/mol more stable than the (E)-exo one leading to trans stereoisomer 2. per- aluminum oxide chromatographic column (chloro. 0. 14 (2006) 955–959 Scheme 1. N-acetylcytosine 11.058 0.88 a specific inhibitory activity. 90 C. lated nucleobases according to the Vorbrüeggen meth- yl hydroxylamine afforded nitrone 7 in E/Z mixture and odology. CH2Cl2. Isoxazolidines 8 and 9 were sensitive to nate moiety and the heterocyclic base. this modification could improve the anti-viral activity 6:1). Chemistry (Table 1). Better results were obtained when the 1. which is comparable with (Z)-endo 246. (Z)-exo 246. thus opening a new perspective on their possible use as therapeutic agents. isoxazolines 8 and 9 in 50% global In this paper. Transition state DHf (kcal/mol) Percent calcd pounds 1 where the oxygen atom in C-5 0 is eliminated. irradiation: the reaction time was reduced to 20 min and ylene moiety (Fig. Compound 7 is light stable and. Figure 1.8 reaction of 5 with triethyl phosphite followed by efficient acidic hydrolysis furnished the corresponding phospho. AM1 calculations10 suggest the (E)-isomer as the of 2.89 that of AZT.956 U. formed in acetonitrile at 55 C in the presence of form–methanol. / Bioorg.

any anti-viral activity is forbidden. which does the b-anomer clearly predominates as the nearly exclu. 3a–d and 4a–d. 300 or 500 MHz (1H) and at 75 MHZ (13C) using deute- were carried out by varying the temperature and the rochloroform as solvent. a/b ratio). Conversely.8 No retroviral activity was observed at the higher concentration tested. the insertion of methylene in compounds 2 not only does not potentiate but determines the com- plete lack of biological effects. according to the literature methods. However. while the distance assigned on the basis of 1H NMR and NOE experi. NOE sequence. chemical shifts are given in amounts of catalyst. The best results have been obtained parts per million from TMS as internal standard. Preparative separations were carried out . that is. using 0. allows Pure anomers were isolated by chromatography on the transfer of a diphosphoryl or a nucleotidyl group. alternate scan subtraction of two FIDs in which the sat- sive product (1:9. enhancements of the H10 . and H70 a signals were observed. Relevant NOE effects on compounds 3a and 4a. The insertion of nucleotides in the growing nucleic acid chain. on Molt-3 cells. All solvents were dried ing H70 b (Fig. As expected. General pyrimidine base are in a trans relationship. / Bioorg. for compounds 3 the distance Pa–N3 is the irradiation of H10 increased the resonance of H70 b not compatible with anyone stable 7-membered chelate (the downfield resonance of methylene protons at 7 0 ). U. were tested achieved through Merck silica gel 60-F254 precoated alu- in vitro for their cytotoxicity. MPLC Büchi C-601 using CHCl3–MeOH (99:1) as The nitrogen atom is probably responsible for metal– solvent (10 mL/min). AZT and Scheme 2. in favor of the formation of the b-isomer 3a. while. we aqueous sodium carbonate to give the deacetylated speculate that the nitrogen atom of the isoxazolidine derivatives 3d and 4d.O-nucleoside system. while in the case of thymine uration frequency is moved on-resonance and off-reso- and 5-fluorouracil a significant amount of a-anomers nance. the references. compound 2. The ability to inhibit the reverse transcriptase activity of avian myeloblastosis retrovirus was tested in a cell free assay as previously reported. chosen as model. Several attempts to optimize the reaction yields and con. and. at 1000 lM. in this context. H40 . Biological assays approximately 5· T1 (5s in our case) before acquiring the FID. is assisted by bivalent Figure 2. Pa–N3 in compounds 2 allows the formation of 6-mem- ments. ring would facilitate the breaking between Pa and Pb in the nucleotides triphosphate and. irradiation of H70 a gave rise to a strong 4. NOE experiments were performed by a modified cycle For N-acetylcytosine. and H40 and H6. it is a steady-state NOE in which a single resonance is saturated at low power for 3. minum plates. Thin-layer chromatographic separations were The synthesized compounds. NMR spectra were recorded on a Varian instrument at ditions. was assayed. consequently. Elemental analyses were performed on a Perkin-Elmer 220B micro analyzer. with refer- ence to known anti-viral compounds (AZT and com- pound 2). These data unam- biguously indicate a b-configuration where H40 and the 4. in these experimental conditions. when H70 b was irradiated. 14 (2006) 955–959 957 the inhibitory activity versus retroviral RT. the a-configuration or anomers 4 was supported Commercially available chemicals and solvents were re- by the strong NOE effect observed for H6 when irradiat.13 Although we do not know the exact interaction of compound of type 2 with the RT polymerase. Experimental positive NOE effect for H6 and H70 b . The relative configurations were ion coordinations and. operated by polymerases. experiments because the irradiation is cycled through the lines of the multiplet.1. completely inhibit the RT activity at the 10 nM. None of the above-mentioned synthetized phosphonated nucleosides show any signifi- cant cytotoxicity up to 1000 lM at the higher concentra- tion tested. bered chelates. Chem. Further- more. Chiacchio et al. Surprisingly. Moreover. in this way.4 equiv of TMSOTf and a temperature of 70 C. Power may be reduced from ordinary NOE has been observed (3:7. metallic ions that facilitate the transfer of nucleotide units. implemented in Darmstadt. 2). the synthesis of compounds 3 provides a contribution to the research on the structural features which concur to determine the biological activity of the N. a/b ratio). agent grade and used as received. Med. In particular for compound 3a.

5SR)-5-acetoxy-2-methylisoxazoli. Med. 2H). 6H.05. Diethyl {(1 0 SR. H70 b ).2 and 7. NH). mer 8: dH (300 MHz. 2.85 (t. 4. 1H. 2H.61.03.13. 4.92 (m. over sodium sulfate.10 (br s.79 (s. 4H. 2.17. 47.35 (dd. and the reaction mixture was stirred at 70 C for 5-fluorouracil 11.3 Hz).0 Hz). 1H.10 (q. 1H.12. N-CH3). / Bioorg.4 0 RS)-1 0 -(5-fluoro-2.28. 1H. 1.2 Hz). 3H. 6H. N-CH3). and the residue was purified through aluminum oxide chromato.9 Hz).040– bis(trimethylsilyl)-acetamide (BSA) (2.60 (m. 3H).9 Hz). Z-isomer: dH (300 MHz. 135. 3. 14 (2006) 955–959 by MPLC Büchi C-601 using Merck silica gel 0.89. 39. 160. 2. J = 7 Hz). 110. (Sticky oil.52. H50 ).50. 16.76 (s. the organic phase pyl} phosphonate 7. 1. 61. A suspension of nucleobase 10–12 dihydropyrimidin-1(2H)-yl)-3 0 -methyl-2 0 -oxa-3 0 -azacyclo- (0.4-dioxo-3. 44. 2. 1H.5 Hz). 16. CDCl3): 1. First eluted fractions gave an inseparable mixture 2H.10 (q.78.62 mmol) in dry acetonitrile (3 mL) was treated with pent-4 0 -yl] ethylphosphonate 4b. 68. H60 ).3. 3H.2. 1H. N. 61. 4. H. 1. 1H NH). pent-4 0 -yl] ethylphosphonate 3b.30. 22. 82.4 0 RS)-1 0 -[[5-methyl-2. 3H. J = 6. thymine 10. 61. 15%).72.97. J = 4.91 (br s. 1H.5.7. J = 1. 1. 4.60.95 (ddd. J = 5. H6). 12. Found: 101. 7. 1H. isoxazolidines 8 4. 20. 4. ized by careful addition of aqueous 5% sodium bicar- bonate and then concentrated in vacuo. 4H. dihydropyrimidin-1(2H)-yl]-3 0 -methyl-2 0 -oxa-3 0 -azacyclo- din-3-yl-ethylphosphonate and diethyl 2-(3RS.11.4 0 RS)-1 0 -[[5-methyl-2.15 (q. 11. 148.30.93 (m.30 (dd. for C15H26N3O6P: C.94 (d.74 (s.04. A solution of diethyl 2-form. To the clear solution obtained were added a solution of the epimeric isoxaz- 4. 9. 8. Method B: the solution of nitrone 7 (5.03 (m.22.10. Chiacchio et al. Diethyl [(1 0 RS. N-CH3). E-isomer: dH J = 4. Calcd for C6H18NO4P: C. dC (125 MHz.79 (s. After being cooled at 0 C.5 Hz).70. 4. 21%).88. Diethyl [(1 0 SR. 6. the sol. 4.66.6.68.13 (q. 9. 61.33. 95:5) to give an dihydropyrimidin-1(2H)-yl]-3 0 -methyl-2 0 -oxa-3 0 -azacyclo- mixture E/Z (1:6) of diethyl {(3Z) and (3E)-3-[methyl pent-4 0 -yl]}ethylphosphonate 3a. H50 ). 4H.91. 6 h. H10 ). 36. H.2 (t.5RS)-5.2. 26. dH . wise. 3H.90.92. 11. (m. of 8 and 9 (1. H70 a ). 7. and 13.05 (s.1 Hz.80 (m. 1. 21.90 (t.5 Hz. 6. 4H. 102. 7.46. 1H.67. CDCl3): 1. 3H. J = 6. J = 6.85 (m. CDCl3): 12. 1H.7 Hz. (500 MHz.60 (s.0 Hz). 6. Found: C. 1H. 2.26. addition of dichloromethane (8 mL). H.32. H4).70. 6H. J = 6. The reaction mixture was then evaporated under re.75 (s. Starting materials olidines 8 and 9 (0. H70 a ). 7. 3. N. H50 ).2. 6.97. General procedure for the preparation of nucleo. 2. CDCl3): 1.5. 47. 1. 36. After the 4.91. Büchi C-601 using CHCl3–MeOH (99:1) as solvent ature for 5 h. H. 2H).99 3H. H. 43. 6. Anal.6.1.71.18. 2. 44. vinyl acetate (30 mL) was heated at 70 C for 12 h. 1H). 55%). and N-acetylcytosine 12 were pur. 9. N. H6). dH roform). 49%).94 (d.1 Hz. 7. afforded an oil (50% yield) identified as a mixture of 20.2.99 (d. 3H). N.70 (ddd. CDCl3): 1.4- 4.90 (m. 29. CDCl3): 1. J = 7 Hz). dC (125 MHz. J = 5. 19. N-methylhydroxyl.40. 83. H60 ).91.49 (s.70. N-CH3). 3H. N.7 mmol) in 2H. 1. 40. 1H). 6H. dH (oxido) imino]propyl} phosphonate 7 as a colorless oil (500 MHz. N.38. 135.16. 6.0. tions.2 Hz).4-dioxo-3.5. 9. 11. H60 ). 1. 1. Calcd CDCl3): 15. 82.31 (t.1.75 (m. H10 ). CDCl3): 1. 12. 11. CDCl3): 1.5 and 13. 2H. J = 6.7. 3. J = 6.05. 1H J = 5.50.2. H40 ).21 mmol) drop- 3-Chloro propanaldehyde diethyl acetal 5. 6. 2. 47.70.01.85.03.35. 20.90. 63.00. Anal. 7.1 and (300 MHz. H70 b ).4 0 RS)-1 0 -(5-fluoro-2. Anal. N.85:1) as a colorless oil.40 (m.7 mmol) and vinyl 47.9 Hz). 1H. 2H). 164.74. 1. (10 mL/min).4- graphic column (chloroform–methanol.82. 1.75 (m.42. fluxed for 15 min under stirring. J = 1. 2H. 2. washed with water (2· 10 mL). 2H).95 (m. 3.98.10 (q. (Sticky oil. 3.04. filtered. 1H. 3H. 3H. Diethyl {(1 0 RS.958 U.2 (t. J = 5. 2. J = 6. H70 b ). H4b). (85% yield). 3H.35 (q.07 (ddd. H. (Sticky oil.54 mmol) and re- 0.78 (m. J = 5. dried ylethylphosphonate 614 (20 mmol). Diethyl 2-(3RS.10 (ddd. 6. N. acetate (10 mL) was irradiated under microwave condi- tions at 100 W and 60 C for 20 min.52 (m. J = 1 Hz.0 and 7. CDCl3): 12. 44.2.063 mm. 111. 2H. 1. 1. 2.61 (q. Method A: a solution of nitrone 7 (5.40. H10 ).50.20 (t. 20. Chem.4-dioxo-3.2. 11. 10. 52. 6H. (300 MHz. 4H. J = 1 Hz. (Sticky oil. CDCl3): 16.73. 46. 6.7 (m.0.4-dioxo-3.2.07. H70 a ).1. Major isomer 9: dH 10.05. 44.5 Hz). amine hydrochloride (20 mmol). H6). 1H. 16. 26. H40 ). dH acetoxy-2-methylisoxazolidin-3-yl-ethylphosphonate 8 (500 MHz. 141. 2.13 duced pressure and after rapid purification through alu. Diethyl {(3Z) and (3E)-3-[methyl (oxido) imino]pro. 61.5.73 (m. minum oxide chromatographic column (chloroform). H4). 1H NH). and 9. J = 7.79.2. the solution was neutral- chased from Aldrich Co. J = 6. 1. 2. 163. J = 6.2. Anal. 1H.9 and 1. Found: C. 44. 150. 61. 4. 6. 2H).76 (m. 1H.36.35 (t.9 Hz).86. Calcd for C14H23N3O6PF: C. 3. 6. J = 7 Hz). 4. 4.92. C.40 (br s. 1H. (ddd.78.73 (m.4. 150. CDCl3): 16. N-CH3). 6.02.35. 63. 1H. J = 4. 2.52 mmol) in dry acetonitrile (3 mL) and trimethylsilyltriflate (TMSOTf) (0. and 13. pent-4 0 -yl]}ethylphosphonate 4a.01.10 (q. H4a). 16. 2H). H.10 (dd.36.10. 61.82 (m.89.55 (m. 2H). dC (125 MHz.35 (t.17. 2. and evaporated to dryness.2 Hz). 6.03 (s.1 and 13.7 and 2. 1H J = 5.67. they were promptly used for the successive reac.2 Hz).56. J = 6. 69. dC (75 MHz. 3H.14. 2.3 Hz). 1H.4- and 9 as a mixture have been obtained in 65% yield dihydropyrimidin-1(2H)-yl)-3 0 -methyl-2 0 -oxa-3 0 -azacyclo- and after a rapid purification on aluminum oxide (chlo. 1. 9.55 (m. 4H.75 (m. 1H. J = 4.48. 6.22 (t. isoxazolidines 8 and 9.1 (dd.00.86.56.39. Found: C.75.12 (q. 2. 7. 4. 1H.6. N-CH3). 61. Minor iso. J = 7 Hz).8 Hz). The reaction mixture was filtered. 4H. J = 6. vent was evaporated under reduced pressure. was separated. and trietylamine The residue was purified by chromatography on MPLC (20 mmol) in toluene (70 mL) was stirred at room temper. 16. 11. 2H).88.46.4- tides 3a–c and 4a–c.65 (m.85 (m. 68. Calcd for C15H26N3O6P: C. dC (75 MHz. N-CH3). H. 127.

. H40 ).12 (q. 2.0.. 1H. Rev. A. B. 24. E.60.31 (t. J = 7 Hz). E. 2002.. Ehler. J = 7. 1H. Pauwels. J = 3. J = 7. M. 14. 1H.0 and 14. Pharmacother. 6. 332–336. Zhong. Vorbrüeggen. NH2). CDCl3): 1.. Mc Intee.22. and 14.. 1H.4 and 7. J = 7.50 (d. 5. S. Chem.. 1H NH). J = 6. Soc. 1H. 7.32 (t. H70 a ). dH (500 MHz. B. Saglimbeni. G. CDCl3): 14. De Clercq. V. 1. Shaker. 3.06. dH (500 MHz. 7.. (Progetto FIRB H10 ).14 (br s. sol. Holy. 13. 1999.4 Hz.. M. Calcd for C14H25N4O5P: C. 704–720.5. 6H.11. H70 b ).. 165..4. 1H. 63.2. 54. De Clercq.81. 2. 188. 114. (Sticky oil. Naesens. 1. H10 ). 2H. 1H NH).4 0 RS)-1 0 -[4-(acetylamino)-2-oxo. Anal... 42.99.40 (d. 14.02..0 Hz). (a) Wagner..53. K. 148. 1389–1394. CDCl3): T. J = 7. 1H. 1H. 48.4 Hz). Rescifina. J = 3. Med.4 0 RS)-1 0 -[4-(acetylamino)-2-oxo. 95. Lacor.2. 2. 2002.49.5 and 9. 3. De 7. E.. CDCl3): 16. U. G. 146. Med.85. H 06 ). J = 7. 15. A. 46. 96.10 6. N. 11. C.40 (t. Birhhauser: Basel- Switzerland.85.. 2005.8. 44. Revuelta.07.0. 142. 62. Diethyl [(1 0 SR.51. C. 2. 4H. 141.50. H. 2. and Girardet. C. HIV Med. NH2). Alfano. Mouriès.4 and 7. 417–451. 1H.59. 8%).. Vandamme. Nature 2004. 54. 157. Hallahan. 83–92.00 (br s. Chiacchio. Natl. J = 7 Hz).75.4 J. 1H. T.14 (br s.. B. 20.33. CDCl3): 1. H. H60 ).. J. CDCl3): 14. N.10 (m. Rosemberg. F. 3.. Johns.. H50 ). Acta 4. 1. 9. H70 a .10.99. A.S. J = 3. E.. Med. Calcd for C14H23N3O6PF: C. 171. 2000. 3c or 4c (100 mg). 55.50. pounds 3d or 4d. J. J = 7 Hz). dC (125 MHz.8). 1H.01 (ddd. 13.95 (m. (a) De Clercq. dH (500 MHz.5.82.. Anal. H50 ).60. 127.. Diethyl [(1 0 RS.74 (ddd. (d. J = 7 Hz). J = 3. L. H40 ). 9.. R.72. Rosemberg. 70%). P.6 Hz.50. (Sticky oil. 1. C. In Combination Therapy of AIDS. M.73.. De Wit. J.46.00 (br s. (d. De Clercq. K.80 (s. Vandercam. 84.R.39 phosphonate 4d. 4. 12.68.80. 3H).58. Acta Clin. Krolikiewicz.4 Hz). 6. 1. 577. 15. 6. 9. 6. 23.74 C. P.4 Hz. Calcd for C14H25N4O5P: C. J = 7. Belson. J = 7.19 (ddd.93. N-CH3). D. (d) Clumeck. 2H. 1H.82.71. G. W. 3H.70. 2H. 95:5) to give the deacetylated com. 54. yl} ethylphosphonate 3c. A.. B. 2. 4. 2. Robbins. 1. 1. 2004.4 and 13. Van Wijngaerden.4 and 13.0 (125 MHz.92. (Sticky oil. NH). Calcd for C16H27N4O6P: C. Van 43. 2H. 61.. 8. Baba. 62. 1H. 2H. 6. 13..93 (d.00 (br s. J.01. R. N. H. (c) Dybul. 7. Chem. 2002). 1. 11.78.50.80. M. 148. 86.. Acknowledgment 3H.66. 1H. N. A.A. 1989.76 (s. W. 1H. 22. 21. Sci.. 9. 2. CDCl3): 14. dC (125 MHz. Int. H06 ).63.10. Koh.89. 4H..78 (m.3 Hz). 165.49. Sigel. W.40 (d. 14 (2006) 955–959 959 (500 MHz. L. (e) Colebun- (125 MHz. Biochim. 7. and 15 Hz. Zoebisch. 8. 70. Hong. F. 1H. U.68.80. E.60 (m. 6H.13 (q.15 (d.35 (m. 4. 1(2H)-yl)-3 0 -methyl-2 0 -oxa-3 0 -azacyclopent-4 0 -yl] ethyl- N-CH3).. J = 7 Hz). dC Wit. 9. 2H. H.82 (d. 96.84. 9.98 (ddd. DeGruttola.. Romeo. J = 7 Hz). 14637–14677. Eds. Synthesis of nucleotides 3d and 4d. Clumeck. Yoder. H06 ). 62.70. 3.65 (m. 21.. M. 44. Chem. 1H. 6.16.. 1. H. J.33. 4. E. J = 7 Hz). J. 7. T. E. CDCl3): (ddd. 1H. 6H.. Eymery. 2. Gosselin. 1H.. / Bioorg. 14. 15–29.74 (ddd. 48. 4H. 33. Belg. N. 95%). 15. V.30. L. 2H.4 0 RS)-1 0 -(4-amino-2-oxopyrimidin. Tejero. 4.22. Curr. G.65. Holy.4 Hz). N.40. J = 7. L. H70 a . 95%). 62.1 (dd. Chem.52 (m. 1H.. M.1 (dd. H70 b ). 4. 141.. 3902–3909. 1H. 4. 2. 3H. Found: C.44. L.4. J = 3. 15. Soc.98 (ddd. J. pyrimidin-1(2H)-yl]-3 0 -methyl-2 0 -oxa-3 0 -azacyclopent-4 0 .. H50 ).20. 1H. 2867–2875. N. Acad. 42. F. 14. (Sticky oil.. 48. 84.72. 7. Chem. 6. H6). 2. J. Chiacchio et al. Sciortino. 6. Biophys. 2H. 14.20. 141. J..30 3581–3592. M. D. E. J = 7. Tetrahe- Found: C.70 (s.6 and 9. Anal. in a mixture of aqueous K2CO3 (5%. 44.6 and 9. Healy. 2003. G. 191–200.72. 1. 70%). J = 3.55. J = 3.. J.28. 148. W. Hertogs. K. 1H.68 (m. 2004.2. 43.49.88. Smith.. yl} ethylphosphonate 4c (sticky oil. J = 7.10 (s. 2.85.. 2. Piperno.70 (br s. 2H. 10. R. M. (b) Zemlicka.20 (ddd. 1981. 6.14 (dd. Chiacchio. 3. P. Proc.58. 3. N.0. 157. Am. 13.20 (ddd.. De 4.... 8. dC (125 MHz. 3.32 (t. Sakuma. 2H.2 (dd. U. 48. 14.72 (m. 24. STD AIDS 2003. (CHCl3–MeOH. H10 ). Daucher. Calcd for C16H27N4O6P: L. Csako. 44.0 Hz. 5 mL) and methanol (5 mL) was left to stir for 4 h. 44. Demothy. 63.6.82.33.4 and 13. 464–467. J = 7. Drug Des.82. S.89.01. phosphonate 3d.7 (m.70. 157.53. H40 ). Dewar. 62.. 2. Balestrieri. Anal.20. P. 1.51. Nies- (m. 1H. 46.. G.31 (t. Clercq..3 Hz).74 (ddd. 16. H50 ). 13. Z.46 (d. Macchi. N-CH3). V. vent was then evaporated under reduced pressure. P. 2. 1997. Iyer. 7. Tetrahedron 2003. Merino. H70 b ). N. M.02 (dd. 2H.57. 1H. J = 2. R. J = 3. 6. J = 7. Savignac. A. 46. Balzarini. Diethyl {(1 0 RS. Biomed. 3H. Infect. CDCl3): 1.U. J. J = 3. N. V. 29. 1H. J. 107. A. A. 15. 1H. and 13. 2000. 1234–1255. (e) Murphy. pyrimidin-1(2H)-yl]-3 0 -methyl-2 0 -oxa-3 0 -azacyclopent-4 0 - 2. This work was supported by M.. H.71. 1(2H)-yl)-3 0 -methyl-2 0 -oxa-3 0 -azacyclopent-4 0 -yl] ethyl. Wu. J. H10 ).99. Bennua. Chem. 6. 9.74.99.15.15 (q. 6.99. Med. Poli. H70 a . Ber.5 Hz. Sklar.70.. Med. 6. Med. 62. Found: C. 3H.0 44.1 Hz). References and notes Found: C.13 (q.. J. dron 1998. 9. 6H.. H. N. J = 7. H. 4H. 150. Dis. (b) Leen. Kraske. V. Romeo. 62.12. 2. 389–411.4 0 RS)-1 0 -(4-amino-2-oxopyrimidin- H50 ). the residue was purified by flash chromatography 7. 44.. H. E. 61.0. (c) Perigaud. Rev.8).74.72.66.08 (s.8 Hz.49.5 and 9. I. Chem. 1H.62.66. Rescifina. 1. 141. J. 2. J = 7 Hz). 276–286. Iorga. Maudgal... 11.80. H06 ). dH (500 MHz. Curr.80. J. ders. C. 2004. P. 44. 9.40. Trials 2001. Wanzeele.. 14.66. 1H NH). 69. 6. M.4 Hz).20.90.24. J. 6H. 4. H.76 (s.13 (q. R. H. 44. 70. Imbach. Valveri. 62.55.4 Hz).10 (m. dC 9.5.00 (br s. 4H. 62. 9 and 15 Hz. Anal.. J = H40 ).38. 14.50.24 and 7. 6.. Rev.11. J = 7. 2H. I. Mastino. (a) Smeaton. Nature 1986. Z. 9. A solution of 2002. P.28 Hz. E.5 and 9. 1H.2. N-CH3).80.50. N-CH3). 95. 6. 2. CDCl3): 14.05 (ddd.97 (m.1 Hz).11. Chem. 1985. 23. 1.. C. N.. Iannazzo. Control Clin. H70 a ). Florence. 59.45.2.. 326. 171.07.4 H40 ). G.40. Herdewijn. Merino.. pp 1–24. 46. J. 1. 7. 2. 150...76 (s.8. A. 1587. H10 ). . 144.81. J = 7. H70 b ). S.4 Hz). 3.9..4. H. (b) Balzarini.95 (m. H. 2005.97. 7.I. 2. 2. E. 3H). Diethyl {(1 0 SR.8 Hz. Stewart. Res. 62. 44. 14.. Found: C.80. 388. J = 3. G. Top.-M. 20. et al. 9. H70 b ). 1H. 84. 4. P. 6. Shim. L. U. 1H NH).4.