TY - JOUR
T1 - Substrate specificity of human deoxycytidine kinase toward antiviral 2prime;,3′-dideoxynucleoside analogs
AU - Kierdaszuk, Borys
AU - Bohman, Christina
AU - Ullman, Buddy
AU - Eriksson, Staffan
N1 - Funding Information:
Acknowledgements-This researchw ass upportedb y Grant MV-492p rovidedt o B.U. by the AmericanC ancerS ociety and by grantst o S.E. from the SwedishM edical Research Council, the Swedish Cancer Society, and the Medical Facultyo f the Karolinska Institute. B.U. is a recipiento f a ResearchC areerD evelopmentA ward from the National Instituteso f Health.
PY - 1992/1/22
Y1 - 1992/1/22
N2 - Deoxycytidine (dCyd) kinase has been purified to homogeneity from human leukemic spleen, and the capacity of the enzyme to phosphorylate 2',3 '-dideoxynucleoside (ddN) analogs that are clinically effective inhibitors of human immunodeficiency virus (HIV) replication was evaluated. Cytosine-containing ddN analogs, such as 2',3'-dideoxycytidine, 2',3'-dideoxy-2',3'-dehydrocytidine, and cytallene, were efficiently phosphorylated by dCyd kinase, while no phosphorylation of purine-containing ddN analogs was detected. dCyd kinase was completely inactive toward 2',3'-dideoxyadenosine (ddAdo), 2',3'-dideoxyinosine, 2',3'-dideoxyguanosine, and adenallene, although it was capable of phosphorylating both 2'-deoxyadenosine (dAdo) and 2'-deoxyguanosine (dGuo). The abilities of wild type and mutant human T lymphoblastoid CEM cells to accumulate ddAdo in situ and in vitro were also ascertained. Comparison of the abilities of intact wild type CEM cells and derivatives deficient in nucleoside transport, dCyd kinase, and/or adenosine (Ado) kinase to accumulate [3H]ddAdo-derived radioactivity revealed no significant differences among the wild type and mutant strains. However, ddAdo phosphorylating activity was decreased in extracts from Ado kinase-deficient cells but not in lysates prepared from cells genetically deficient in dCyd kinase activity. In comparative growth rate experiments, wild type, nucleoside transport-deficient, and dCyd kinase-deficient CEM cells were equally sensitive to ddAdo toxicity, while, interestingly, a deficiency in Ado kinase correlated with a 5-fold decreased growth sensitivity to the purine ddN. Insertion of an adenine phosphoribosyltransferase deficiency into the CEM cell lines did not influence ddAdo toxicity or incorporation rate. These results imply that Ado kinase may be an important factor in ddAdo phosphorylation by CEM cells. Furthermore, these studies demonstrate that cytosine- and purine-containing ddNs are transported and activated by independent pathways and, therefore, have important implications for anti-HIV therapy in that pyrimidine and purine ddNs might be used in combination for the treatment of acquired immunodeficiency syndrome.
AB - Deoxycytidine (dCyd) kinase has been purified to homogeneity from human leukemic spleen, and the capacity of the enzyme to phosphorylate 2',3 '-dideoxynucleoside (ddN) analogs that are clinically effective inhibitors of human immunodeficiency virus (HIV) replication was evaluated. Cytosine-containing ddN analogs, such as 2',3'-dideoxycytidine, 2',3'-dideoxy-2',3'-dehydrocytidine, and cytallene, were efficiently phosphorylated by dCyd kinase, while no phosphorylation of purine-containing ddN analogs was detected. dCyd kinase was completely inactive toward 2',3'-dideoxyadenosine (ddAdo), 2',3'-dideoxyinosine, 2',3'-dideoxyguanosine, and adenallene, although it was capable of phosphorylating both 2'-deoxyadenosine (dAdo) and 2'-deoxyguanosine (dGuo). The abilities of wild type and mutant human T lymphoblastoid CEM cells to accumulate ddAdo in situ and in vitro were also ascertained. Comparison of the abilities of intact wild type CEM cells and derivatives deficient in nucleoside transport, dCyd kinase, and/or adenosine (Ado) kinase to accumulate [3H]ddAdo-derived radioactivity revealed no significant differences among the wild type and mutant strains. However, ddAdo phosphorylating activity was decreased in extracts from Ado kinase-deficient cells but not in lysates prepared from cells genetically deficient in dCyd kinase activity. In comparative growth rate experiments, wild type, nucleoside transport-deficient, and dCyd kinase-deficient CEM cells were equally sensitive to ddAdo toxicity, while, interestingly, a deficiency in Ado kinase correlated with a 5-fold decreased growth sensitivity to the purine ddN. Insertion of an adenine phosphoribosyltransferase deficiency into the CEM cell lines did not influence ddAdo toxicity or incorporation rate. These results imply that Ado kinase may be an important factor in ddAdo phosphorylation by CEM cells. Furthermore, these studies demonstrate that cytosine- and purine-containing ddNs are transported and activated by independent pathways and, therefore, have important implications for anti-HIV therapy in that pyrimidine and purine ddNs might be used in combination for the treatment of acquired immunodeficiency syndrome.
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U2 - 10.1016/0006-2952(92)90278-Q
DO - 10.1016/0006-2952(92)90278-Q
M3 - Article
C2 - 1739408
AN - SCOPUS:0026537061
SN - 0006-2952
VL - 43
SP - 197
EP - 206
JO - Biochemical Pharmacology
JF - Biochemical Pharmacology
IS - 2
ER -