Molecular cloning of cDNAs which are highly overexpressed in mitoxantrone-resistant cells: Demonstration of homology to ABC transport genes

Reports of multiple distinct mitoxantrone-resistant sublines without overex pression of P-glycoprotein or the multidrug-resistance associated protein h ave raised the possibility of the existence of another major transporter co nferring drug resistance, In the present study, a cDNA library from mitoxan trone-resistant S1-M1-80 human colon carcinoma cells was screened by differ ential hybridization, Two cDNAs of different lengths were isolated and desi gnated MXR1 and MXR2. Sequencing revealed a high degree of homology for the cDNAs with Expressed Sequence Tag sequences previously identified as belon ging to an ATP binding cassette transporter. Homology to the Drosophila whi te gene and its homologues was found for the predicted amino acid sequence, Using either cDNA as a probe in a Northern analysis demonstrated high leve ls of expression in the S1-M1-80 cells and in the human breast cancer subli ne, MCF-7 AdVp3000. Levels were lower in earlier steps of selection, and in partial revertants. The gene is amplified 10-12-fold in the MCF-7 AdVp3000 cells, but not in the S1-M1-80 cells, These studies are consistent with th e identification of a new ATP binding cassette transporter, which is overex pressed in mitoxantrone-resistant cells.