AMPLIFICATION AND EXPRESSION OF THE ABC TRANSPORTERS ARA AND MRP IN ASERIES OF MULTIDRUG-RESISTANT LEUKEMIA-CELL SUBLINES

E1000, the most drug-resistant subline from the E-series (CCRF-CEM/E16 to E1000), has been previously shown to express high mRNA levels from two ABC transporter genes associated with multidrug resistance, ARA a nd MRP, The expression and amplification of both genes has now been ch aracterized for each member of the E-series of drug-resistant sublines and is reported here. Both ARA [detected by reverse transcriptase pol ymerase chain reaction (RT-PCR)] and MRP (detected by Northern blot an alysis) were expressed at low levels in the sensitive parental CEM cel l line. An equivalent level of MRP mRNA expression was detected throug hout the GEM, E16, E25 and E50 sublines, and there was increasing expr ession in the E100, E200 and E1000 sublines. ARA expression was not de tected in the E16, E25, E50 and E100 sublines but was detected by both RT-PCR and Northern blot analysis in the E200 and E1000 sublines, Sou thern blot analysis indicated the increased levels of MRP and ARA expr ession resulted from gene amplification and that MRP was first amplifi ed in the E100 subline and ARA in the E200 subline, suggesting that th e two genes were not initially co-amplified. Cytogenetic analysis of E 1000 cells demonstrated a large addition to chromosome 16p, around the region where the ARA and MRP genes are located. Increased expression of ARA is associated with increased colchicine resistance in the E-ser ies of sublines and combined with MRP may account for their resistance phenotype.