CHARACTERIZATION OF MOUSE LEUKEMIA-L1210 CELLS RESISTANT TO THE RIBONUCLEOTIDE REDUCTASE INHIBITOR 4-METHYL-5-AMINO-1-FORMYLISOQUINOLINE THIOSEMICARBAZONE

Mouse leukemia L1210 cells were generated for resistance to 4-methyl-5 -amino-1-formylisoquinoline thiosemicarbazone (MAIQ), a potent inhibit or of ribonucleotide reductase that is directed at the nonheme iron su bunit (NHI) of the enzyme, The resistant cells, MQ-580, showed an 8-fo ld increase in IC50 toward MAIQ, a 4-fold increase in IC50 toward hydr oxyurea, and also showed resistance to other ribonucleotide reductase inhibitors. In addition, the MQ-580 cell line was resistant to nonribo nucleotide reductase inhibitors such as etoposide, daunomycin and vinb lastine, but not to cisplatin, The mRNA for the NHI subunit was increa sed 7-fold in the MQ-580 cells with essentially no change in the mRNA level for the effector-binding subunit, The ribonucleotide reductase a ctivity in the cell-free extracts prepared from the MQ-580 cells was o nly slightly elevated (30%). However, passage of the cell-free extract from the MQ-580 cells over Sephadex G-25 resulted in a 4.8-fold incre ase in specific activity over that of the wild-type cells. While the r eductase activity in the cell-free extract from the MQ-580 cells did n ot show altered sensitivity to MAIQ, the reductase activity in the cel l-free extract from the MQ-580 cells was much more sensitive to the ef fects of the iron-chelating agents Desferal and EDTA. The cell pellets from the MQ-580 cells were much darker in color than the pellets from the wild-type cells or hydroxyurea-resistant cells. The supernatant f raction from the MQ-580 cells after-SDS-PAGE showed the appearance of a strong Coomassie blue-staining band at 50 kDa that was not apparent in either the wild-type or hydroxyurea-resistant cells. This new resis tant cell line offers an opportunity to explore differences in resista nce mechanisms of drugs (e.g. MAIQ and hydroxyurea) that are directed at the same subunit of ribonucleotide reductase. Copyright (C) 1996 El sevier Science Inc.