TY - JOUR
T1 - Cytotoxicity of 3′-azido-3′-deoxythymidine correlates with 3′-azidothymidine-5′-monophosphate (AZTMP) levels, whereas antihuman immunodeficiency virus (HIV) activity correlates with 3′-azidothymidine-5′-triphosphate (AZTTP) levels in cultured CEM T-lymphoblastoid cells
AU - Törnevik, Yvonne
AU - Ullman, Buddy
AU - Balzarini, Jan
AU - Wahren, Britta
AU - Eriksson, Staffan
PY - 1995/3/15
Y1 - 1995/3/15
N2 - Activation of the anti-human immunodeficiency virus (HIV) compound 3′-azido-3′-deoxythymidine (AZT) is dependent on its 5′-phosphorylation by cellular nucleoside and nucleotide kinases. Azidothymidine 5′-triphosphate (AZTTP) is considered to be the metabolite responsible for both the anti-HIV effect of AZT, via inhibition of reverse transcriptase, and cytoxicity by interference with cellular DNA polymerases. During the characterization of AZT metabolism in cultured human T-lymphoblastoid CEM cells, a spontaneously occurring variant cell line, CEM/Ag-1, was found that showed approximately 10-fold resistance to AZT growth inhibition as compared to wild type (wt) cells (EC50 = 2 mM as compared to 350 μM for wt cells). CEM/Ag-1 cells had a 3-fold reduced capacity to accumulate azidothymidine monophosphate (AZTMP) compared to wt cells whereas similar levels of AZTTP were found in both cell lines. The intracellular half-life of AZTMP was approximately 70 min in both wt and CEM/Ag-1 cells. A 3-fold lower specific activity of cytoplasmic thymidine kinase was observed in CEM/Ag-1 extracts as compared to wt. The reduced thymidine kinase activity was not correlated to a decreased level of thymidine kinase mRNA. Syncytium formation of CEM/Ag-1 cells infected with HIV-2 as well as HIV-1 antigen production was inhibited at the same concentrations of AZT (approx. 0.01 μM) as were HIV-1 and HIV-2 infected wt cells. Thus, minor decreases in cellular thymidine kinase levels may markedly affect the cytoxicity of AZT but have no major effect on the antiviral activity of AZT. Our results strongly suggest that AZTMP is responsible for a major part of the growth inhibitor effects, while AZTTP mainly mediates the antiviral activity of AZT.
AB - Activation of the anti-human immunodeficiency virus (HIV) compound 3′-azido-3′-deoxythymidine (AZT) is dependent on its 5′-phosphorylation by cellular nucleoside and nucleotide kinases. Azidothymidine 5′-triphosphate (AZTTP) is considered to be the metabolite responsible for both the anti-HIV effect of AZT, via inhibition of reverse transcriptase, and cytoxicity by interference with cellular DNA polymerases. During the characterization of AZT metabolism in cultured human T-lymphoblastoid CEM cells, a spontaneously occurring variant cell line, CEM/Ag-1, was found that showed approximately 10-fold resistance to AZT growth inhibition as compared to wild type (wt) cells (EC50 = 2 mM as compared to 350 μM for wt cells). CEM/Ag-1 cells had a 3-fold reduced capacity to accumulate azidothymidine monophosphate (AZTMP) compared to wt cells whereas similar levels of AZTTP were found in both cell lines. The intracellular half-life of AZTMP was approximately 70 min in both wt and CEM/Ag-1 cells. A 3-fold lower specific activity of cytoplasmic thymidine kinase was observed in CEM/Ag-1 extracts as compared to wt. The reduced thymidine kinase activity was not correlated to a decreased level of thymidine kinase mRNA. Syncytium formation of CEM/Ag-1 cells infected with HIV-2 as well as HIV-1 antigen production was inhibited at the same concentrations of AZT (approx. 0.01 μM) as were HIV-1 and HIV-2 infected wt cells. Thus, minor decreases in cellular thymidine kinase levels may markedly affect the cytoxicity of AZT but have no major effect on the antiviral activity of AZT. Our results strongly suggest that AZTMP is responsible for a major part of the growth inhibitor effects, while AZTTP mainly mediates the antiviral activity of AZT.
KW - antiviral chemotherapy
KW - cytotoxicity
KW - deoxynucleoside kinases
KW - metabolism
KW - nucleoside analogs
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U2 - 10.1016/0006-2952(94)00453-S
DO - 10.1016/0006-2952(94)00453-S
M3 - Article
C2 - 7702641
AN - SCOPUS:0028920558
SN - 0006-2952
VL - 49
SP - 829
EP - 837
JO - Biochemical Pharmacology
JF - Biochemical Pharmacology
IS - 6
ER -