title: Long-term exposure to zidovudine affects in vitro and in vivo the efficiency of phosphorylation of thymidine kinase.
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 (Vmax) of TK activity for deoxythymidine (dThd) phosphorylation is decreased in CEMazt as compared to the wild-type cell line (Vmax: 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 (Km: CEM = 0.9 +/- 0.2 microM; CEMazt = 1.6 +/- 0.2 microM; p < 0.01). Consequently phosphorylation efficiency, expressed as the ratio between Vmax and Km, 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.
authors.affiliation: Department of Biomedicine, University of Pisa, Italy.
authors.forename: G
authors.initials: G
authors.lastname: Antonelli
authors.forename: O
authors.initials: O
authors.lastname: Turriziani
authors.forename: A
authors.initials: A
authors.lastname: Verri
authors.forename: P
authors.initials: P
authors.lastname: Narciso
authors.forename: F
authors.initials: F
authors.lastname: Ferri
authors.forename: G
authors.initials: G
authors.lastname: D'Offizi
authors.forename: F
authors.initials: F
authors.lastname: Dianzani
mesh_terms: D002460:Cell Line; D015658:HIV Infections / Q000097:blood / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D015497:HIV-1; D006801:Humans; D007700:Kinetics; D007963:Leukocytes, Mononuclear / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D010766:Phosphorylation; D013601:T-Lymphocytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D013936:Thymidine / Q000378:metabolism; D013937:Thymidine Kinase / Q000187:drug effects* / Q000378:metabolism; D013997:Time Factors; D014316:Tritium; D015215:Zidovudine / Q000494:pharmacology*
chemical_list: D014316:Tritium; D015215:Zidovudine; D013937:Thymidine Kinase; D013936:Thymidine