ACTIVITY OF P-GLYCOPROTEIN IN B-CELL CHRONIC LYMPHOCYTIC-LEUKEMIA DETERMINED BY A FLOW CYTOMETRIC ASSAY

Background: Chemoresistance in some hematologic malignancies has been associated with overexpression of P-glycoprotein, which is encoded by the MDR1 gene (also known as PGY1). However, inconsistencies in data o n frequency and clinical relevance of multidrug resistance in B-cell c hronic lymphocytic leukemia (B-CLL) may reflect a need for improved te chniques to detect this overexpression. Purpose: Our purpose was to me asure P-glycoprotein activity in peripheral blood cells of B-CLL patie nts and to analyze possible clinical correlations (disease duration, p rior treatment, Rai disease stage, lymphocyte counts, and disease prog ression). Methods: P-glycoprotein activity was assayed in peripheral b lood cells of 42 consecutive B-CLL patients (22 treated and 20 untreat ed). We used dual fluorescence in a flow cytometric assay that detects efflux of the fluorescent dye rhodamine 123, which is transported fro m the cell by the P-glyprotein pump. Leukemia cells were co-stained wi th monoclonal antibody Leu12/CD19, and rhodamine 123 efflux was measur ed. Expression of MDR1 and MDR3 (also known as PGY3) messenger RNA (mR NA) was quantitatively evaluated by polymerase chain reaction (PCR) in 26 cases. Results: Marked rhodamine 123 efflux was observed in 34 (81 %) of the 42 cases and was abolished in the presence of multidrug resi stance inhibitors. Rhodamine 123 efflux was not associated with Rai st age, lymphocyte counts, duration of disease, or disease progression. A lthough rhodamine 123-negative cases were about equally distributed am ong untreated and previously treated patients, the percentage of cells with rhodamine 123 efflux was significantly lower for untreated patie nts than for those treated with chemotherapy regimens including at lea st one multidrug resistance-associated drug. MDR1 mRNA was detected in 25 of 26 cases and MDR3 mRNA in all 26. MDR1 mRNA expression was sign ificantly correlated with rhodamine 123 efflux, whereas MDR3 mRNA expr ession was not significantly correlated; MDR1 and MDR3 mRNA expression was not significantly associated with Rai stage, prior treatment, or disease progression. Conclusions: These findings suggest that P-glycop rotein overexpression in B-CLL is intrinsic rather than acquired and t hat P-glycoprotein activity is enhanced after exposure to multidrug re sistance-associated drugs. This enhanced activity does not seem to be associated with more aggressive disease. Our results also indicate tha t an assay of P-glycoprotein function combined with PCR is suitable fo r clinical multidrug resistance screening. Implications: Additional st udies are needed to determine whether functional activity of P-glycopr otein, measured by rhodamine 123 efflux, is directly related to clinic al drug resistance.