Phosphorylation of the carbocyclic analog of 2′-deoxyguanosine in cells infected with herpes viruses

The carbocyclic analog of 2t′-deoxyguanosine [(±)-2-amino-1,9-dihydro-9-[(1 α, 3 β, 4 α)-3-hydroxy -4-(hydroxymethyl)cyclopentyl]-6 H-purine-6-one] (2′-CDG) is highly active in cell culture against strains S148 and E377 of herpes simplex virus type 1 (HSV-1), both of which code for thymidine kinase, and much less active against strain BW10168 which is deficient in this enzyme activity. Antiviral activity is associated primarily with the d-enantiomer; the l-enantiomer has much lower but significant activity. The metabolism of racemic 2′-CDG and its d- and l-enantiomers was studied in uninfected HEp-2 cells and in HEp-2 cells infected with the S148 or BW10168 strains of HSV-1. Nucleotides were separated by HPLC, and their elution was monitored by spectrophotometry. The chromatograms of extracts of cells infected with the S148 strain and treated with (±)-2′-CDG or d-2′-CDG included a new peak which appeared in the triphosphate region. This peak, the area of which exceeded that of the GTP peak, was shown to be due to the triphosphate of 2′-CDG. The new peak was not observed by HPLC of extracts of uninfected cells treated with (±)-2′-CDG or either of its enantiomers, cells infected with the S148 strain and treated with l-2′-CDG, or cells infected with the BW10168 strain and treated with (±)-2′-CDG or either of its enantiomers. The results were similar when these studies were performed with uninfected Vero cells and with Vero cells infected with strain S148 of HSV-1. In experiments with d-[8- 3h]-2′-cdg, small amounts of phosphates of 2′-CDG could also be detected in uninfected HEp-2 cells and in cells infected with the BW10168 strain of HSV-1. Thus, 2′-CDG apparently is a good substrate for the virus-coded kinase and a very poor substrate for cellular phosphorylating enzymes. The selective phosphorylation of 2′-CDG by the virus-specific kinase presumably is critical for its antiviral activity as it is for that of acyclovir and other acyclic derivatives of guanine.