Intrinsic and acquired multidrug resistance and its relationship to the expression of P-glycoprotein
Separate mechanisms underlie the MDR phenotype found in tumour cells selected for resistance to low concentrations of daunorubicin and vincristine sulphate, singly and in combination. The vincristine sulphate selected VCR series were resistant to 4,8,16ng/ml; the daunorubicin selected DNR series were resistant to 10,15,30ng/ml and finally the combination series (DNR:VCR) series were resistant to 1:0.04; 5:0.2 and 10:0.4ng/ml of daunorubicin and vincristine sulphate respectively. The three resistant cell line series thus derived were characterised with respect to the following: a) growth characteristics; b) in vitro cross resistance using the MTT cytotoxic assay; c) amplification of MDR1 and MDR3 genes; d) level of MDR1 and MDR3 transcripts using RNA dot blot analyses; e) overexpression of P-glycoprotein was carried out using a panel of antibodies, C219, MRK-16 and JSB-1; f) drug accumulation studies using the Rhodamine assay, and finally an attempt was made to g) identify any novel amplified sequences. The combination drugs (DNR:VCR) series exhibited characteristics of the classical MDR phenotype at lower levels of resistance; marginally increased levels of P-glycoprotein and its transcript MDR1, and cross resistance to a wide range of structurally and functionally dissimilar drugs. This classic phenotype however gave way to an atypical phenotype in HSB-2 10:0.4DV cell line, with no overexpression of P-glycoprotein and its transcript MDR1 and a cross resistance profile indicative of DNA Topoisomerase II involvement; resistant to anthracyclines, etoposide and only marginally resistant to vincristine, vinblastine and colchicine. In comparison, the single drug exposure cell line series, demonstrated a progressive increase of P-glycoprotein and its transcript MDR1.