ORGAN-SPECIFIC MODULATION OF STEADY-STATE MDR GENE-EXPRESSION AND DRUG-RESISTANCE IN MURINE COLON-CANCER CELLS

Background: The major cause of death from cancer is metastases that ar e resistant to conventional therapies. The resistance of metastatic tu mor cells to chemotherapy can be caused by their intrinsic properties, such as increased expression of the mdr genes. Purpose: The purpose o f our present study was to determine some of the mechanisms by which t he organ microenvironment influences the response of tumor cells to ch emotherapy. Methods: Murine CT-26 colon cancer cells growing in contin uous culture (parental cells) were harvested and injected subcutaneous ly into the lateral flank (to produce subcutaneous tumors) or the late ral tail vein (to produce experimental lung metastases) of 10 8-week-o ld syngeneic male BALB/c mice. Seven days after tumor-cell injection, the mice were given intravenous injections of either doxorubicin (10 m g/kg) or 0.9% NaCl (controls). This in vivo injection was repeated 7 d ays later. Mice with subcutaneous tumors and lung metastases were kill ed by cervical dislocation on day 21, and tumor samples from control m ice were harvested and adapted to culture. The sensitivity of the cult ured cells to doxorubicin and fluorouracil (5-FU) was determined at mu ltiple time points. Levels of mdr-1 DNA were measured by slot-blot and Southern-blot analyses. mdr mRNA expression levels were measured by N orthern-blot analysis using mdr-1- and mdr-3-specific hybridization pr obes, and P-glycoprotein level was determined by fluorescente-activate d cell sorting using different monoclonal antibodies. Results: Treatme nt with doxorubicin produced 80% growth inhibition of CT-26 subcutaneo us tumors but had little effect on the number (and size) of experiment al lung metastases. Collectively, the results suggest that the multidr ug-resistant phenotype developed in CT-26 cells growing in the lung en vironment. Cultures established from lung metastases were initially re sistant to doxorubicin (but not to 5-FU) and showed elevated expressio n of mdr-1 mRNA transcripts and P-glycoprotein. This resistance could be overcome by verapamil and disappeared after 21 days in culture. No mdr gene amplification was detected. The expression level of mdr-speci fic mRNA (predominance of mdr-1) and P-glycoprotein was directly assoc iated with resistance to doxorubicin. Conclusions: Results of this stu dy have demonstrated that the in vivo sensitivity of murine CT-26 colo n carcinoma cells to doxorubicin depends on the organ environment. The organ environment can influence the P-glycoprotein-mediated multidrug -resistant phenotype in tumor cells, and the increased expression of P -glgcoprotein is transient; once removed from the environment (lung), the cell's resistance reverts to that of the sensitive parent cells.