Antiviral Activity ofArabinosylthymine inHerpesviral Replication: Mechanism ofAction InVivoandInVitro

(ara-T), ametabolite ofthesponge Tethya crypta, hasshownselective activity against herpes simplex virus (HSV)replication(G.A.Gentry andJ.F.Aswell, Virology 65:294-296, 1975). Analysis of HSV-infected anduninfected cell lysates byCsClisopycnic centrifugation showed thatara-T blocked theincorporation of[3H]hypoxanthine intoviral deoxyribonucleic acid and, toalarge extent, into hostdeoxyribonucleic acid of infected (but notuninfected) cells. Additional experiments with[y-32P]adenosine 5'-triphosphate asaradiophosphate donor demonstrated thatara-T isphosphorylated byextracts ofHSV-infected BHK cells andnotbythose ofuninfected cells. Atanara-T concentration that almost completely inhibited thegrowth of LM cells, whichhadbeentransformed toapyrimidine deoxyribonucleoside kinase+ (dPyK+) phenotype byultraviolet-inac tivated HSV-1,thegrowth of uninfected LMcells wasnotaffected. These results indicate that theviral dPyK isresponsible fortheselective antiviral activity ofara-T. Thisconclusion was further supported byexperiments that showed that thereplication ofavariety of dPyK-mutants ofHSV-1andHSV-2werenotaffected byara-T andthatara-T inhibited thephosphorylation ofdeoxycytidine anddeoxythymidine byHSV-1 dPyK,butnotbyhostdeoxycytidine anddeoxythymidine kinases, respectively. Ara-T also selectively inhibited thereplication ofequine herpesvirus type 1(EHV-1) invitro andwaseffective against EHV-1infection invivoinhamsters. Further, EHV-1wasinhibited byara-T andbybromodeoxyuridine in LM cells lacking acytosol thymidine kinase, suggesting thatEHV-1induces a dPyK.Finally, spectrophotometric assay forthymine suggested thatara-T isnot asubstrate fornucleoside phosphorylase ofhamster liver, andamicrobiological assay indicated that substantial amounts ofara-T wereexcreted intheurine of uninfected hamsters thathadreceived asingle injection of5mgofara-T, the amountgiven ineachinjection intheinvivoexperiments withEHV-1. 1-p-D-Arabinofuranosylthymine (ara-T; spon