Correlation between intracellular pharmacological activation of nucleoside analogues and HIV suppression in vitro.
Following intracellular activation of HIV nucleoside analogue reverse transcriptase inhibitors, their triphosphates (ddNTPs) compete with endogenous nucleoside triphosphates (dNTPs) for incorporation into proviral DNA. In this study we have examined the effect of combinations of two thymidine analogues, stavudine (d4T) and zidovudine (ZDV), and two cytidine analogues, lamivudine (3TC) and zalcitabine (ddC) on intracellular drug activation and on the relevant competing dNTP in uninfected and persistently HIV-infected cells. Endogenous triphosphates of deoxycytidine (dCTP) and deoxythymidine (dTTP) were measured using a template primer assay and the ratio of ddNTP:dNTP was calculated. Antiviral activity of two-drug combinations was also assayed by p24 ELISA. A significant reduction in d4T triphosphate (d4TTP) [0.11+/-0.09 pmol/10(6) cells to undetectable (<0.01); P=0.039] in the presence of equimolar concentrations of ZDV and d4T, resulted in a decrease in the d4TTP/dTTP ratio of 90%. ZDVTP/dTTP was not significantly altered in the presence of d4T. 3TC (10 microM) reduced total ddC phosphates by 57% and ddCTP/dCTP by 27%. 3TC phosphorylation was comparatively unaffected by ddC, up to a concentration of 10 microM ddC (>100 times the plasma concentration achieved following standard dosing). 3TC plus ddC resulted in greater p24 inhibition than 3TC or ddC alone (P<0.001). Combining one thymidine analogue (ZDV or d4T) with one cytidine analogue (3TC or ddC) resulted in greater inhibition of p24 inhibition than with any single agent. From a pharmacological viewpoint, the combination of ZDV plus d4T should be avoided, but in vitro the combination of 3TC plus ddC confers modest benefit over either drug alone. This in vitro study illustrates that decreases in ddNTP/dNTP are consistent with a reduction in antiviral effect.
Hoggard
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PG
Department of Pharmacology & Therapeutics, University of Liverpool, UK. patrick@liverpool.ac.uk
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D019380:Anti-HIV Agents / Q000494:pharmacology / Q000627:therapeutic use; D015658:HIV Infections / Q000188:drug therapy* / Q000201:enzymology / Q000821:virology; D015497:HIV-1 / Q000187:drug effects* / Q000502:physiology; D006801:Humans; D018894:Reverse Transcriptase Inhibitors / Q000494:pharmacology* / Q000627:therapeutic use; D018119:Stavudine / Q000494:pharmacology / Q000627:therapeutic use; D014779:Virus Replication / Q000187:drug effects; D016047:Zalcitabine / Q000494:pharmacology / Q000627:therapeutic use; D015215:Zidovudine / Q000494:pharmacology / Q000627:therapeutic use
D019380:Anti-HIV Agents; D018894:Reverse Transcriptase Inhibitors; D015215:Zidovudine; D016047:Zalcitabine; D018119:Stavudine