The naturally occurring
nucleoside analogue arabinosyl
thymidine is known as an anti-herpes and anti-
cancer agent. The biologically active form is arabinosyl
thymidine triphosphate (
Ara-TTP), which inhibits cellular and
viral DNA-polymerases and thus interferes with DNA replication. Using two murine
erythroleukemia cell lines, Friend cell clone F4-6 and F4-12N, the latter being
thymidine kinase deficient (TK-) cells transformed to a TK+ phenotype with the HSV TK gene, we have determined 1) the role of cellular and herpes simplex virus
thymidine kinase (HSV TK) in the uptake of
Ara-T into the cells; 2) the subsequent phosphorylation of intracellular
Ara-T to
Ara-TMP, Ara-TDP and
Ara-TTP; 3) the incorporation of
Ara-TTP into the
DNA. Incorporation into
DNA was studied under different conditions, including selective inhibition of the different cellular
DNA polymerases by
aphidicolin (that inhibits polymerases alpha and delta) and dideoxythymidine (that preferentially inhibits polymerases beta and gamma). The uptake of
Ara-T into the
methanol soluble pool of the cells depends upon its phosphorylation to
Ara-TMP, which is more efficiently performed by the HSV TK than by the cellular TK, thus explaining the sensitivity of HSV infected cells to
Ara-T. However, using increasing concentrations of
Ara-T, we have shown that phosphorylation also occurs in normal control cells due to the cellular
thymidine kinase. More than 90% of
Ara-T is phosphorylated in the cell, and more than 60% of total
Ara-T(MP, DP, TP) exists in the
triphosphate form.(ABSTRACT TRUNCATED AT 250 WORDS)