In vitro studies on the cellular mechanisms of resistance to cytotoxic compounds with special reference to multidrug resistance and cancer chemotherapy

Abstract

The development of resistance to antineoplastic drugs is a major obstacle in the treatment of cancer. When resistance develops to natural product, lipophilic antitumour agents the resistance is also conferred to structurally and functionally unrelated compounds. This phenomenon is termed multidrug resistance (MDR). The development of MDR in vitro is associated with a host of phenotypic changes which are believed to be involved in the cellular mechanism of MDR.
In this study a murine mammary tumour cell line, EMT-6, and the previously uncharacterised doxorubicin-selectetj derived subline were utilised to investigate the role of some the MDR-associated phenotypic changes in the mechanism of resistance.
The EMT-6/R cell line, permanently maintained in 5.0 j.tM doxorubicin, was found to differ from the parental cell line, EMT-6/S, in a number of basic growth parameters such as size, doubling time, cloning efficiency and stationary phase cell density. The plasma membrane fluidity and total protein content of the cell lines were not significantly different. Immunoprecipitation revealed the presence of P-glycoprotein in the EMT-6/R cell line but not in any detectable amounts in the parent cell line. No significant difference in the glutathione content was observed although a significant increase in glutathione-S-transferase (GST) activity was detected in the EMT-6/R cell line.
Although resistance was selected using doxorubicin, the EMT-6/R cell line also displayed cross-resistance to vincristine and colchicine, but not to Cis-platin nor methotrexate. The EMT-6fR cell line, however, displayed hypersensitivity (compared
to the parental cell line) to a number of lipophilic compounds used as resistance reversing agents. The same reversing agents were shown, at non-toxic concentrations, to potentiate the cytotoxicity of doxorubicin in the EMT-6R cell line but not by any appreciable amount in the drug-sensitive parent cell line. Assessment of cellular was also revealed, the nature of which could not be determined except that it was closely related to P-glycoprotein.
These studies confirm that MDR is multifactorial in origin and that the overexpression of P-glycoprotein is likely to be a major component of the resistance mechanism. Further work is required to determine the extent to which the control of P-glycoprotein can be mediated through phosphorylation and to find specific effectors of this process which could be useful in cancer chemotherapy.