Identification of novel drug resistance-associated proteins by a panel of rat monoclonal antibodies.

Since some multidrug-resistant (MDR) tumor cell lines show drug accumulation defects but do not over-express Pgp or MDR protein (MRP), a search was made for novel MDR-related transporter proteins by immunizing rats with non-small cell lung cancer SW- 1573/2R120 cells to produce monoclonal antibodies (MAbs). Five rat MAbs (LMR-4, -12, -42, -44 and -94) were generated, showing strong membranous staining of non-Pgp MDR SW- 1573/2R120 tumor cells and minimal reactivity to the corresponding parental and revertant cell lines. In addition, a 6th MAb (LMR-5) was isolated, recognizing the MDR-related lung resistance protein (LRP), previously identified as the major vault protein. The first 5 LMR MAbs show predominantly membranous staining of several non-Pgp MDR tumor cell lines of different histogenetic origins, except for LMR-4, which recognizes only MDR sublines of the SW- 1573 cell line. Flow-cytometric analysis revealed that all MAbs, except LMR-4 and -5, detect outside epitopes. Functional studies showed that these MAbs did not restore the daunorubicin accumulation defect. All but one of the MAbs (LMR-42) showed staining of distinct normal human tissues, notably epithelial cells lining the airways and digestive tract. In addition, staining of vascular endothelial cells was found with all MAbs except LMR-4. Three MAbs (LMR-12, -44 and -94) showed remarkable immunoreactivity with vincristine-selected SW- 1573 sublines. By immunoblotting and precipitation, the LMR antigens were found to be in the 42-69 kDa range.