INTRACELLULAR METABOLISM OF THE N7-SUBSTITUTED ACYCLIC NUCLEOSIDE ANALOG 2-AMINO-7-(1,3-DIHYDROXY-2-PROPOXYMETHYL)PURINE, A POTENT INHIBITOR OF HERPESVIRUS REPLICATION

We investigated the intracellular metabolism of S2242 (2-amino-7-(1,3- dihydroxy-2-propoxymethyl)purine), the only known antivirally active a cyclic nucleoside analogue with the side chain substituted at the N7 p osition of the purine ring. Uptake of S2242 by CEM cells increased lin early with increasing extracellular concentrations of the compound and was blocked by inhibitors of nucleoside transport. S2242 was phosphor ylated in a time-and concentration-dependent manner to its monophospha tes, diphosphates, and triphosphates. Intracellular half-life of the d iphosphates and triphosphates in CEM cells was similar to 3-6 hr. A st rong correlation was found between the cytostatic action of the compou nd and its phosphorylation in different cell lines. In accord with the findings that (1) the cytostatic potential of S2242 is reversed by de oxycytidine (dCyd) and (2) the growth of deoxycytidine kinase-deficien t (dCK(-)) cells is refractory to the inhibitory effect of S2242, the amount of metabolites formed from S2242 in the dCK(-) cell line was ap proximately one hundredth of that in the wild-type cells. The observat ion that purified dCK phosphorylates S2242 to its monophosphate furthe r corroborates these results. The activity of S2242 against herpes sim plex virus, varicella-zoster virus, and human herpesvirus type 6 was r eversed by 50-100-fold on the addition of exogenous dCyd. Compound S22 42 was not preferentially phosphorylated in herpes simplex virus 1-, v ari cella-zoster virus-, or human herpesvirus type 6-infected cells (V ero, human embryonic lung, and HSB-2 cells, respectively), and exogeno usly added dCyd reduced substantially the formation of S2242 metabolit es in these cells. in human cytomegalovirus (HCMV)-infected human embr yonic lung cells, a 5-25-fold increase in S2242 metabolite formation w as observed compared with the noninfected cells, suggesting that an HC MV-encoded or -induced enzyme causes the specific phosphorylation of S 2242. Exogenously added dCyd had little effect on the activity of S224 2 against HCMV and on the phosphorylation of the compound in HCMV-infe cted cells. S2242 was not specifically phosphorylated by the HCMV-enco ded UL-97 kinase in cells infected with a vaccinia/UL-97 recombinant. S2242 was found to be a substrate (K-m, = 90 mu M) for purified human deoxyguanosine kinase; the latter enzyme was stimulated 3-4-fold in HC MV-infected cells.