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Quality control of multidrug resistance assays in adult acute leukemia:
correlation between assays for P-glycoprotein expression and activity
HJ Broxterman, P Sonneveld, N Feller, GJ Ossenkoppele, DC Wahrer, CA Eekman, M Schoester, J Lankelma, HM Pinedo, B Lowenberg and GJ Schuurhuis
Department of Medical Oncology, Academisch Ziekenhuis Vrije Universiteit,
Amsterdam, The Netherlands.
We have compared multiple assays for the P-glycoprotein (Pgp/MDR1)
phenotype in fresh and thawed adult acute leukemia to validate and
quantitate measures for the expression and function of Pgp. The results are
related to the Pgp-expressing KB8 and KB8-5 call lines. The most sensitive
assay was the measurement of modulation of the rhodamine 123 (R123)
fluorescence by 2 micromol/L PSC833, followed by the modulation of the
probe calcein-AM. We also found a good intralaboratory and interlaboratory
correlation between the values of the R123/PSC833 assay for fresh as well
as thawed samples. In addition, the affects of PSC833 on 3H-daunorubicin
(DNR) accumulation, DNR fluorescence, and 3H- vincristine accumulation were
very similar. The correlation between the DNR/PSC833 and R123/PSC833 test
was r = .86 (N = 51). The modulation of drug accumulation by 8 micromol/L
verapamil was the some as the PSC833 effect for DNR (117%, N = 21), but was
higher for vincristine in every single case (161% v 121%, N = 22; P<
.001), indicating additional verapamil effects, not related to Pgp. The
correlation of the staining of viable cells for Pgp with the monoclonal
antibody MRK16 was r = .77 (N = 52) for the R123/PSC833 functional test and
r = .84 (N = 50) for the DNR/PSC833 test. From these results it could be
calculated that a maximal increase of the mean DNR accumulation of about
50% can be achieved by blocking Pgp pump activity with PSC833 in leukemic
blast samples with the highest mean Pgp expression. Subpopulations of blast
calls with higher Pgp activity are likely to be present. Their relevance
has to be studied further. The methods outlined here allow the reliable,
quantitative monitoring of the Pgp/MDR1 phenotype in leukemias in
multicentered, clinical Pgp modulation studies.
Volume 87,
Issue 11,
pp. 4809-4816,
06/01/1996
Copyright © 1996 by The American Society of Hematology
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