Detection of P-glycoprotein in cell lines and leukemic blasts: failure of select monoclonal antibodies to detect clinically significant Pgp levels inprimary cells

Multidrug resistance (MDR) is a salient feature of chemotherapy failure in pediatric patients. One of the most common and well-studied mechanisms impl icated in causing MDR is P-glycoprotein (Pgp), an ATP-dependent, transmembr ane drug efflux. pump. Accurate and reproducible detection of this MDR prot ein is necessary as it may have important clinical implications. In this st udy comparing the directly conjugated anti-Pgp monoclonal antibodies UIC2-P E and 15D3-PE to the unconjugated anti-Pgp mAb MRK16, we analyzed cell line s, normal peripheral blood cells, and bone marrow cells from pediatric pati ents diagnosed with acute myeloid leukemia and acute lymphoblastic leukemia ; all samples were also analyzed for Pgp function using rhodamine 123 in or der to correlate results from antibody staining with functional activity. F or all patient samples evaluated, only MRK16 correlated well with the rhoda mine 123 assay. Both the directly conjugated antibodies UIC2-PE and 15D3-PE failed to detect Pgp in almost all cases. Pre-treatment of cells with neur aminidase did not provide a consistent enhancement of antigen detection. Ba sed on these results, we suggest that while UIC2-PE and 15D3-PE may be able to detect the very high levels of Pgp expressing laboratory-cultured cell lines, they are not suitable for clinical application in their currently av ailable conjugated form. When assaying patient samples for Pgp expression a nd function using flow cytometry, the rhodamine 123 functional assay should be performed in concert with staining with MRK16. Published by Elsevier Sc ience Ltd.