4 -Azido-2 -deoxycytidine

The cytidine and inosine analogues in the 4 -azido-2 -deoxy series were made by well known procedures from the uridine and adenosine analogues, respectively (Scheme II). Thus, following the method of Reese, 4 -azido-2 -deoxyuridine was acetylated and then converted to the 4-triazolide, which on treatment with ammonium hydroxide furnished 4 -azido-2 -deoxycytidine. Using a procedure of Herdewijn, 4 -azido-2 -deoxyadenosine was enzymatically deaminated to the inosine analogue. 

Additional information <1> (<1>, the deletion mutants rDm-dNKAClO and rDm-dNKAC20 show the same substrate activity pattern as the recombinant wild-type enzyme. Relative phosphorylation of 2 -deoxycytidine and 2-chloro-2 -deoxyadenosine increases with increasing C-terminal truncation. The relative activities of rDm-dNKAClO and rDm-dNKAC20 with deoxyribonucleosides remains largely unchanged, whereas there is a substantial decrease in the phosphorylation of the purine ribonucleosides adenosine and guanosine, as well as of all dideoxyribonucleosides and 3 -azido-2 ,3 -dide-oxythymidine. The relative activities with the pyrimidine ribonucleosides and l-/l-D-arabinofuranosylcytosine and l-/i-D-arabinofuranosylthymine are not affected by the C-terminal deletions [4]) [4]... 

Numerous azido-substituted nucleosides have been evaluated as potential antitumour agents. De Clercq et al. found that of a series of 2 - and 3 -azidonucleoside analogues prepared and evaluated for antitumour activity against murine LI210 cells in vitro, the most potent derivative was 3 -azido-2, 3 -dideoxyadenosine (146), with 3 -azido-2, 3 -dideoxyguanosine (102), 3 -azido-3 -deoxyadenosine (147) and 2 -azido-2 -deoxycytidine (148) proving moderately active [170], the activity of the last compound having been reported previously [171, 172], AZT was also evaluated in this study... 

The 2 -azido group of cytarazid renders the nucleoside more resistant to deamination to the 2 -azidouridine derivative (153) by deoxycytidine deaminase, but was also observed to reduce substrate affinity for the deoxycytidine kinase necessary for anabolic phosphorylation to the active cytarazid 5 -triphosphate [180]. Conversely, cytarazid was a more potent inhibitor of the target DNA polymerases a and K = 0.6 and 0.7 //M, respectively) than the parent ara-C (.A = 10 and 17 fiM, respectively), and the dissimilar spectrum of antitumour activity exhibited by the two compounds was attributed to differences in stability, metabolic activation and inhibitory potency [180, 181]. Interestingly, the instability of cytarazid to thiols present in the assay media, was commented on but not pursued [180]. In view of previous discussions concerning the bioreduction of AZT... 

The form of the enzyme found in the cytosol acts only on deoxythymidine. The thymidine kinase found in the mitochondria acts on deoxythymidine, deoxycytidine, and deoxyuridine. The mitochondrial thymidine kinase s activity is sufficiently broad that it will also act on the anti-HIV drug, 3 -azido-2 3 -dideoxythymidine (AZT). Evidence suggest that deoxyribonucleotides of AZT interfere with mitochondrial function, possibly by inhibiting mitochondrial DNA replication or transcription, which may explain some of the side effects of cardiotoxicity observed with its use. 

Azido and 3 -amino analogues of 2 -deoxy-5-substituted uridine and cytidine, together with 3 -amino-5 -fluoro-thymidine, have been prepared by standard procedures using 3 -0-mesyl ester intermediates the most active anti-cancer compounds were the 3 -amino-5-fluoro analogues of 2 -deoxyuridine and 2 -deoxycytidine cuid the 3 -amino analogue of 2 -deoxycytidine.An alternative doubleinversion sequence at C-3 using a 3 -chloro-3 -deoxyxylo-nucleo-side intermediate has been described for the preparation of 3 -... 

Analogue of deoxycytidine triphosphate, exo-N- 2-[N-(4-azido-2,5-difluoro-3 -chloropyridine-6-yl)-3 -amino-propionyl] aminoethyl -2 -deoxycytidine-5 triphosphate which has 4-azidopyridine as a photoactive site, was applied for photoaffmity modification of DNA binding proteins [111]. 

Replacement of the uracil moiety of the 3 -azido nucleoside analogue 1 with the cytosine to form 7 did not affect its antiviral activity however, when the substituent on carbon-5 of the cytosine moiety was either fluoro 8 or methyl 9, the antiviral activity was markedly reduced. This could be explained if these 3 -azido-2, 3 -dideoxycytidine analogues were required to be deaminated by deoxycytidine deaminase for which the 5-methyl or 5-fluoro analogues are not substrates. Other possibilities include differences in metabolic conversion to the di- and triphosphates analogues, as well as the relative affinities of the triphosphate analogues for the reverse transcriptase. 

Replacement of the 3 -azido moiety of 2 with hydrogen (20) resulted in loss of activity however, subsequent removal of a hydrogen atom from both the 2 - and 3 -carbon produced the active compound 17 (D4T) thus the azido moiety per se is not an absolute requirement for the antiviral activity of the thymidine analogues. However, replacement of the 5-methyl group of 17 with a hydrogen (16) yielded a loss of activity. The azido moiety conferred moderate antiviral activity to 2 -deoxycytidine (7), but this moiety also is not an absolute requirement since its replacement with a hydrogen (24) resulted in a marked increase in activity. Subsequent removal of a hydrogen from the 2 - and 3 -carbon of 24 to produce 18 did not affect the increase in antiviral activity from that of 7. Deamination of 24 produced 19 with a concomitant loss of antiviral activity, which could not be recovered by insertion of a variety of substituents on the carbon-5 of the uracil moiety (20-23). 

Among the 3 -azido analogues of pyrimidine deoxyribonucleosides, 3 -azido-3 -deoxy-thymidine (2, AZT) was the most active against HIV-1 in vitro with an EC50 value of 0.002 jaM. Conversely, 3 -azido-3 -deoxy-6-azathymidine (37) was practically inactive (EC50 >100 aM). The 3 -azido derivatives of 3 -deoxy-3-(3-oxo-l-propenyl)thymidine (36), 2 -deoxyuridine (1), 5-bromo-2 -deoxyuridine (5), 2 -deoxy-5-fluorocytidine (8), 2 -deoxy-5-iodouridine (6), 2 -deoxycytidine (7), 2 -deoxy-5-fluorouridine (4), 2 -deoxy-5-thio-... 

As was the case with the thymidine and uridine nucleosides, the addition of an azido or cyano group to the 4 -position of 2 -deoxycytidine led to very potent compounds. 4 -Azidocytidine, in comparison, was inactive. The 2 -deoxycytidine analogs in general were as potent as the thymidine derivatives and more potent than the 2 -deoxyuridine analogs. 4 -Cyano-2 -deoxycytidine was significantly more potent than was its 2 ,3 -dideoxycytidine counterpart. This trend was also observed in the thymidine series. 

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