Monoclonal antibodies (MAbs
) recognizing external epitopes of the human MDRl P-glycoprotein have been used both for the detection of multidrug-resistant cells and as specific inhibitors of P-glycoprotein-mediated multidrug resistance. Using a panel of recently developed transfected or transgenic cell lines containing variants of the human MDRl and MDR3 P-glycoproteins, we have compared the specificity and binding properties of the previously isolated MAbs MRKl6, HYB-241, UlC2 and 4E3, and of the newly isolated MAb 7G4. The removal of I, 2 or all 3 of the N-glycosylation sites present in the first extracellular loop of MDRl P-glycoprotein did not significantly affect the binding of these MAbs. In contrast, a 20 amino acid deletion in the first extracellular loop of MDRl P-glycoprotein completely abolished binding of UICZ, whereas the binding of all other MAbs was hardly affected. None of the MAbs tested bound detectably to cell lines containing a high level of the human MDR3 P-glycoprotein. The differences in the binding specificity between UIC2 and the other tested antibodies parallel the reported functional differences in the ability of these antibodies to inhibit P-glycoprotein-mediated drug efflux, o 1993 Wilqv-Liss, Inc.
Multidrug resistance (MDR) is the phenomenon that, after selection with a single cytotoxic drug, cells can become resistant to a whole range of drugs with different structures and cellular targets. In mammalian cells, one of the established causes of MDR is over-production of P-glycoproteins (Endicott and Ling, 1989; Schinkel and Borst, 1991). P-glycoproteins are large, glycosylated plasma membrane proteins that can function as active, ATP-hydrolyzing drug efflux pumps. They are about 1280 amino acids long, composed of 2 similar halves, each containing 6 predicted transmembrane segments and a putative intracellular nucleotide binding domain (see Fig. la). P-glycoproteins are encoded by small gene families. In humans, 2 members are present, respectively MDRl and MDR3 (also called MDR2). Transfection experiments have shown that the human MDR1, and the related mouse mdrla and mdrlb complementary DNAs (cDNAs) can confer multidrug resistance. Despite extensive efforts, attempts to confer multidrug resistance with the closely related human MDR3 or mouse mdr2 cDNAs have failed (Van der Bliek et aL, 1988;Gros et al., 1988; Schinkel etal., 1991), making it likely that the encoded P-glycoproteins are not able to transport hydrophobic drugs. These proteins probably transport some other substrate.
Recent studies indicate that the human MDRl P-glycoprotein may play a role in the clinical resistance of at least some tumors against chemotherapy (Goldstein et al., 1989;Chan et al., 1991). Monoclonal antibodies (MAbs) that specifically recognize P-glycoproteins are important in the fundamental and clinical analysis of MDR. They have been used for characterization of the protein itself (