Abstract
The purpose of this study was to investigate the mechanism of acquired cellular resistance to AZT, a mechanism that has been described as a potential source of drug resistance in addition to viral mutations.
To study this phenomenon the kinetics parameters of thymidine kinase (TK) activity have been defined in CEMazt, a cell line previously selected for resistance to AZT, in comparison with the parental AZT-sensitive CEM cells.
The results revealed that the value of the maximum velocity (V max) of TK activity for deoxythymidine (dThd) phosphorylation is decreased in CEMazt as compared to the wild-type cell line (V max: CEM = 105.3 ± 17.6 nmol/hr/mg of protein; CEMazt = 0.3 ± 0.02 nmol/hr/mg of protein; p < 0.001). Furthermore, the enzyme affinity versus dThd is lower in CEMazt as compared to CEM (K m: CEM = 0.9 ± 0.2 μM; CEMazt = 1.6 ± 0.2 μM; p < 0.01). Consequently phosphorylation efficiency, expressed as the ratio between V max and K m, is also reduced in CEMazt (p < 0.001).
To evaluate whether such a phenomenon may also occur in patients, ex vivo experiments were carried out by using PBMCs from HIV-infected patients, treated or not treated with AZT. The results (mean values from 10 patients for each group) indicate that a prolonged treatment (>6 months) with AZT may modify the enzymatic kinetics of TK, leading to a significant reduction in the phosphorylation efficiency of the enzyme (4.07 ± 1.7 in treated patients versus 13.5 ± 1.7 in untreated patients; p < 0.001).
These results indicate that AZT treatment can also induce a defect in TK activity in patients.
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