Functional characterization of minimal residual disease for P-glycoproteinand multidrug resistance protein activity in acute myeloid leukemia

Relapse is common in acute myeloid leukemia (AML) due to persistence of res idual leukemia cells: minimal residual disease (MRD). In 102 out of 127 pat ients (80%), cells at diagnosis displayed one or more leukemia-associated p henotypes (LAP), le combinations of cell surface markers which are absent i n normal cells and can thus be used to detect MRD at followup. Functional c haracterization of MRD cells for P-glycoprotein (Pgp) and multidrug resista nce protein (MRP) activity is essential to investigate the role of these dr ug transport proteins in multidrug resistance in AML. A fluorescent probe a ssay using Syto16/PSC833 and calcein-AM/probenecid as substrate/modulator o f the Pgp and MRP pump, respectively, and subsequent labeling of cells with monoclonal antibodies for LAP detection allowed simultaneous detection of LAP and Pgp or MRP activity. Validation of this assay is shown for 30 newly diagnosed AML and 11 MRD situations. In addition, no significant differenc es were found when comparing fresh and cryopreserved de novo AML for LAP ex pression (n = 43), Pgp (n = 30) and MRP (n = 24) function and for MRD sampl es for simultaneous LAP expression and Pgp/MRP activity (n = 10). This appr oach enables longitudinal and multicenter studies on the detection, quantif ication and functional characterisation of MRD cells.