The Effect of Multidrug Resistance Modulators on Cytotoxicity of Adriamycin upon Human Renal Cell Carcinoma Cell Lines

Kim, Hyeon Hoe; Ahn, Han Jong; Song, Kyuyoung; Lee, Eun Sik; Lee, Chong wook

Abstract

It is well known that renal cell carcinoma shows poor responses upon chemotherapy, and a multidrug-resistance has been suggested as one of the possible mechanisms of these resistances to chemotherapeutics of renal cell carcinoma as well as other malignancies. We tried to measure the expressions of the multidrug resistance gene and p-glycoprotein in human renal cell carcinoma cell lines, and to evaluate whether various multidrug resistance modulators could enhance the cytotoxicity of adriamycin. Four human renal cell carcinoma cell lines, A-498, A-704, Caki-1, Caki -2, were used and verapamil, cyclosporin A, cefoperazone, quinidine were used as the multidrug resistance modulating agents. Polymerase chain reaction was used to detect the expression of MDR1 mRNA, and measurement of p-glycoprotein was done by FACScan using JSB-1 monoclonal antibody. Cytotoxicity of adriamycin was measured by MTT colorimetric assay. Expression of MDR1 mRNA was observed in A-704, and A-498, but was not detected in Caki-1 and Caki-2. Expression of p-glycoprotein was found in A-498 and A-704, but not in Caki-1 and Caki-2. The relative expression rates of MDR1 and p-glycoprotein in A-498, A-704, Caki-1 and Caki-2 comparing to KB-3-1, the negative control cell line were 1.75, 2.44, 0.98, 0.97 and 1.31, 1.12, 1.08, 0.78 respectively. From these observations, A-498 was selected as MDR positive cell line and Caki-2 as MDR negative cell line, and then a study was performed to evaluate the effect of multidrug resistance modulators on the cytotoxicity of adriamycin upon these cell lines. Verapamil, in concentration of 0.1/microM, did not enhance the anticancer effect of adriamycin on A -498 cells, but with the concentration of 1 microM and 10microM, it decreased IC(50) of adriamycin from 0.43 microgram/ml when only adriamycin was used to 0.21 and 0.16, showing the dose modification effect of 2. 05 and 2.68 respectively (p<0.05, by Mann Whitney test). Cyclosporin A, in all the concentration of 0.3, 1, 3 microM, also decreased IC(50) of adriamycin on A-498 cells to 0.17, 0.14, 0.15 microgram/ml respectively, showing the dose modification effect of 2.53 to 3.07 (p<0.05, by Mann Whitney test). But the cytotoxicity of adriamycin was not influenced by cefoperazone and quinidine. In Caki-2 cells, in which MDR1 and p-glycoprotein expression were barely detected, verapamil and cyclosporin A as well as cefoperazone and quinidine did not show any effect upon the cytotoxicity of adriamycin. Considering above results, verapamil and cyclosporin A seem to be an effective multidrug resistance modulating agents to enhance the cytotoxicity of adriamycin in renal cell carcinoma showing MDR1 and p-glycoprotein expression, and further studies including in vivo study are needed before clinical trials to improve chemotherapeutic effect upon renal cell carcinoma.