Resistance to the microtubule inhibitor podophyllotoxin: Selection and partial characterization of mutants in CHO cells

Abstract

Stable mutants two- to fourfold more resistant to the microtubule inhibitor podophyllotoxin (Pod^RI)have been selected in a single step in Chinese hamster ovary (CHO) cells. Mutants showing higher levels of resistance to the drug can be obtained in a second-step selection using the Pod^RI mutants (Pod^RII class).Prior treatment of cells with the mutagen ethylmethane sulfonate markedly enhanced the frequency of both Pod^RI and Pod^RII mutants in culture. The spontaneous rate of mutation of Pod^RI as determined by fluctuation analysis was found to be 1−2×10⁻⁵ and 4× 10⁻⁷ mutations/cell/generation at 20 and 30 ng/ml podophyllotoxin, respectively. Somatic cell hybrids between Pod^RI × Pod^RII and Pod^RII × Pod^S showed intermediate levels of resistance to the drug, indicating that the Pod^R phenotype behaves codominantly under these conditions. Cross-resistance studies with various Pod^RI and Pod^RII mutants show that they are not cross-resistant to other microtubule inhibitors, e.g., colchicine, colcemid, griseofulvin, vinblastin, that have been examined, or to a compound such as VP16-213, which is a podophyllotoxin derivative which lacks microtubule inhibitory activity. These results indicate that the lesion in Pod^R mutants is highly specific and probably does not involve a permeability alteration.