Summary Dexrazoxane combined with doxorubicin (+ 5-fluorouracil + cyclophosphamide -the FAC regime) leads to a significant decrease in doxorubicin cardiotoxicity and a significant increase in median survival time for patients with advanced breast cancer responsive to FAC. The reason for this increase in survival may be due to interference with the mechanism involved in the emergence of multidrug resistance (MDR). In order to test this hypothesis, we induced resistance to doxorubicin in the K562 cell line by growing cells in increasing concentrations of doxorubicin (10-30 nM) in the presence and absence of dexrazoxane (20 nM). The doxorubicin sensitivity of all resultant sublines was measured using the MTT assay. Flow cytometry was used to assess the MDR1 phenotype, measuring P-glycoprotein expression with MRK 16 antibody and drug accumulation in the presence and absence of PSC 833 for functional P-glycoprotein. Long-term growth in doxorubicin increased the cellular resistance (IC 50 ) of K562 cells in a concentration-dependent manner (r 2 = 0.908). Doxorubicin resistance was not induced in the presence of dexrazoxane (P < 0.0001) for several months. In parallel, the expression of functional P-glycoprotein was delayed after concomitant addition of dexrazoxane to the selecting medium (P < 0.001). Dexrazoxane did not act as a conventional modulator of P-glycoprotein. These results suggest that dexrazoxane may delay the development of MDR1, thus allowing responders to the FAC regime to continue to respond. (7), 959-964 © 2001 Cancer Research Campaign doi: 10.1054/ bjoc.2001.1697, available online at http://www.idealibrary.com on http://www.bjcancer.com develops an MDR1 phenotype within a few weeks (Kato et al, 1990). K562 cells have a doubling time of approximately 16 hours, were maintained in RPMI 1640 culture medium plus 10% FCS, 100 1U ml -1 penicillin, 100 µg ml -1 streptomycin (all from Sigma Aldridge, UK) and were passaged every 48 hours to 1.0 × 10 5 cells ml -1 .A pilot study was carried out in order to optimize the concentrations for long-term exposure to dexrazoxane (Cardioxane, Chiron, UK) and doxorubicin. We tested dexrazoxane at a range of concentrations up to 5 µM and established that concentrations up to 200 nM were commensurate with long-term growth. Doxorubicin resistant sublines were then generated by incubating K562 parental cells at 1.0 × 10 5 cells ml -1 in the presence of sublethal concentrations of doxorubicin ± dexrazoxane. The starting concentration used for doxorubicin was 10 nM and it was possible to increase this stepwise by 10 nM amounts to 40 nM in the presence of 20 nM dexrazoxane. Cells selected in doxorubicin without dexrazoxane showed typical signs of drug damage manifested by cell swelling, alterations in cell shape and granularity. This became detectable after 4 days and when the cells had adapted to this environment (approximately 2-4 weeks) the concentration of doxorubicin was increased. The cells were examined microscopically every 48 hours before passaging and if low growt...
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