The multidrug resistance proteins (MRPs) MRP1, MRP2, MRP3, MRP5 and P-glycoprotein (P-gp) act in concert with each other to give a net resultant pump function in acute myeloid leukemia (AML). The aim of the present study was to analyze the activity of these proteins, which might be upregulated at relapse as compared with de novo AML due to clonal selection. The mRNA expression and activity of P-gp and the MRPs were determined with RT-PCR and flow cytometry, in conjunction with phenotype, as measured with the monoclonal antibodies CD34, CD38 and CD33, in 30 paired samples of de novo and relapsed AML. P-gp and MRP activity varied strongly between the cases (rhodamine 123 efflux-blocking by PSC833: 5.4 ؎ 7.7, and carboxyfluorescein efflux-blocking by MK-571: 4.3 ؎ 6.7, n ‫؍‬ 60). P-gp and MRP activity were increased in 23% and 40% of the relapse samples, and decreased in 30% and 20% of the relapse samples, respectively (as defined by a difference of Ͼ2 ؋ standard deviation of the assays). Up-or downregulation of mRNA expression was observed for MDR1 (40%), MRP1 (20%), MRP2 (15%), MRP3 (30%), and MRP5 (5%). Phenotyping demonstrated a more mature phenotype in 23% of the relapsed AML cases, and a more immature phenotype in 23% of the relapses, which was independent of the karyotypic changes that were observed in 50% of the studied cases. P-gp and MRP activity correlated with the phenotypic changes, with higher P-gp and MRP activities in less mature cells (r ‫؍‬ −0.66, P Ͻ 0.001 and r ‫؍‬ ؊0.31, P ‫؍‬ 0.02, n ‫؍‬ 58). In conclusion, this study shows that P-gp and MRP activity are not consistently upregulated in relapsed AML. However, P-gp and MRP activities were correlated with the maturation stage as defined by immune phenotype, which was observed to be different in 46% of the relapses. Leukemia (2001) 15, 1544-1553.

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Deletion of the multidrug resistance gene MRP1has been demonstrated in acute myeloid leukemia (AML) patients with inversion of chromosome 16 (inv[16]). These AML patients are known to have a relatively favorable prognosis, which suggests thatMRP1 might play an important role in determining clinical outcome. This study analyzed MRP1 deletion by fluorescent in situ hybridization (FISH), with a focus on inv(16) AML patients. Functional activity of multidrug resistance protein (MRP) was studied in a flow cytometric assay with the use of the MRP substrate carboxyfluorescein (CF) and the inhibitor MK-571. MRP1, MRP2, and MRP6 messenger RNA (mRNA) expression was determined with reverse transcriptase–polymerase chain reaction (RT-PCR). The results were compared with normal bone marrow cells. MRP1deletion was detected in 7 AML patients; 2 cases showed no MRP1FISH signals, and 5 cases had 1 MRP1 signal, whereas in 4 AML patients with inv(16) no MRP1 deletions were observed. A variability in MRP activity, expressed as CF efflux–blocking by MK-571, was observed (efflux-blocking factors varied between 1.2 and 3.6); this correlated with the number of MRP1 genes (r = 0.91, P < .01). MRP activity in the AML cases was not different from normal hematopoietic cells. MRP1 mRNA was detected in patients with 1 or 2 MRP1 FISH signals, but not in patients with no MRP1 signals. MRP2 and MRP6 mRNA were expressed predominantly in AML samples with 1 MRP1 signal, whereas in normal bone marrow cells no MRP2 and MRP6 mRNA was observed. In conclusion, this study shows that MRP activity varies among inv(16) AML cases and does not differ from that in normal hematopoietic cells; this might be in part due to the up-regulation of other MRP genes.