| Autor: |
Stegmann, A.P.A., Honders, W.H., Willemze, R., Haperen, V.W.T.Ruiz van, Landegent, J.E. |
| Zdroj: |
Blood; March 1995, Vol. 85 Issue: 5 p1188-1194, 7p |
| Abstrakt: |
The AraC-resistant rat leukemic cell line R0/1-A has been shown to have a typical deoxycytidine kinase (DCK)-defi-cient phenotype and cannot metabolize the antileukemic drugs cytarabine (AraC) and decitabine (DAC). To investigate the relative contribution of mutations in the dckgene to the development of in vitro–induced AraC-resistance, a neomycin selectable plasmid construct harboring the wild-type dckcoding region was transfected into RO/1-A. Polymerase chain reaction analysis confirmed the presence of vector DNA in the target cells (RO/1-ADCK) that were stably transfected and monitored over a period of 14 weeks. Northern and Western blot analysis showed restoration of dckmRNA and protein expression. Initial rate measurements of DCK activity showed that Kmvalues for dckwere only slightly altered as a result of transfection, whereas strongly increased Vmaxvalues were observed, resulting in a 12-fold increased phosphorylation efficiency for both dC and AraC, compared with the AraC-sensitive parental cell line RO/1 from which the RO/1-A was originally derived. In vitro sensitivity to AraC- and DAC-mediated cytotoxicity was fully restored in RO/1-ADCK. The data pinpoint acquired DCKdeficiency caused by mutations of the dckgene as the major cause of AraC resistance in this model. |
| Databáze: |
Supplemental Index |
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