RAPID EMERGENCE OF METHOTREXATE RESISTANCE IN CULTURED MOUSE CELLS

  • 1 January 1984
    • journal article
    • research article
    • Vol. 44  (8) , 3303-3306
Abstract
The time required for mouse [fibroblast] 3T6 cells to become resistant to 200 nM methotrexate was examined by 3 selection protocols: a single-step 0-200 nM dose; a 2-step 0-80-200 nM dose; and a multistep 0-40-80-120-160-200 nM dose. An initial inoculum of 5 .times. 105 cells was grown to 106 cells at each increment of methotrexate, reduced to 5 .times. 105 cells and again grown to 106 ells at the next increment. The total elapsed time required for an initial inoculum of 5 .times. 105 cells to grow to 1 .times. 106 cells resistant to 200 nM methotrexate was 45, 21 and 6.5 days, respectively, for the 3 drug dosage schedules. The single-step resistant variants did not contain amplified dihydrofolate reductase genes, whereas cells resistant to 200 nM methotrexate by the 2 stepwise selections were resistant as a result of a 6-fold amplification of the dihydrofolate reductase gene. The resistance to 200 nM methotrexate resulting from gene amplification did not preexist in the initial population but was generated during the selection process. These results are discussed in terms of the emergence of drug resistance during the course of chemotherapy of tumors.