5-methyldeoxycytidine and ibacitabine

The incorporation into DNA of 5-bromocytosine and 5-iodocytosine, derived from their respective administered deoxyribonucleoside analogs, has been demonstrated in studies with cells infected with herpes simplex virus types 1 and 2 (HSV-1 and HSV-2) and in cells transformed with the thymidine kinase gene of HSV-1. No significant incorporation of iodocytosine or iodouracil occurred in the DNA of uninfected or nontransformed cells when the deaminating enzymes were inhibited, in accord with past studies in our laboratory with 5-bromodeoxycytidine and tetrahydrouridine. When 2'-deoxytetrahydrouridine, a potent inhibitor of cytidine deaminase and dCMP deaminase, was utilized, all the counts in DNA that were derived from [(125)I]iododeoxycytidine appeared as iodocytosine in HSV-infected cells. In the absence of a deaminase inhibitor, 32 to 45% of the counts associated with DNA pyrimidines appeared as iodocytosine, and 55 to 68% appeared as iodouracil in HSV-infected cells. Substantial incorporation of iodocytosine (16%) occurred in cells transformed with the HSV thymidine kinase gene, suggesting the importance of the specificity of cellular nucleoside kinases and the activity of the deaminases in presenting unmodified bases to an undiscriminating polymerase. Incorporation into DNA of bromocytosine derived from [(3)H]bromodeoxycytidine was demonstrated in HSV-2 infected cells; very little incorporation of bromocytosine compared with bromouracil could be demonstrated in these cells in the absence of inhibition of the deaminases (19% of the total counts associated with pyrimidines with deaminase inhibition and 1.5% without). Limited studies with 5-methyl[5-(3)H]deoxycytidine indicated essentially no (or very little) incorporation of this analog as such in the DNA of HSV-1- and HSV-2-infected and -transformed cells. This suggests an exclusion or repair mechanism preventing inappropriate methylcytosine incorporation in DNA. The addition of nucleoside and deoxyribonucleoside deaminase inhibitors, which leads to the incorporation of 5-halogenated analogs of deoxycytidine into DNA as such, does not impair their antiviral activity. We infer from studies with 4-N-alkyl (ethyl and isopropyl)-substituted analogs of iododeoxycytidine that they are incorporated as such into DNA without deamination and effectively inhibit the virus at concentrations that are marginally toxic. Among the several reasons presented for the heightened potential efficacy of analogs of deoxycytidine

Topics: Animals; Antiviral Agents; Bromodeoxycytidine; Cell Transformation, Viral; Cytidine Deaminase; Deamination; Deoxycytidine; DNA, Viral; Kinetics; Simplexvirus; Structure-Activity Relationship; Thymidine Kinase; Tritium