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Doxorubicin sensitizes human bladder carcinoma cells to Fas-mediated cytotoxicity
Y Mizutani1, Y Okada, O Yoshida
1Department of Urology, Faculty of Medicine, Kyoto University, Japan.
Background:
The resistance of bladder carcinoma to anticancer chemotherapeutic agents remains a major problem. Hence, several immunotherapeutic approaches have been developed to treat the drug-resistant cancer cells. Fas antigen (Fas) and Fas ligand participate in cytotoxicity mediated by T lymphocytes and natural killer cells. Like Fas ligand, anti-Fas monoclonal antibody (MoAb) induces apoptosis of the cells expressing Fas. This study examined whether bladder carcinoma cells are sensitive to cytotoxicity mediated by anti-Fas MoAb and whether anticancer agents synergize with anti-Fas MoAb in cytotoxicity.
Methods:
Cytotoxicity was determined by a 1-day microculture tetrazolium dye assay. Synergy was assessed by isobolographic analysis.
Results:
The T24 human bladder carcinoma cell line constitutively expressed the Fas on the cell surface; however, T24 line was resistant to anti-Fas MoAb. Treatment of T24 cells with anti-Fas MoAb in combination with mitomycin C, methotrexate, or 5-fluorouracil did not overcome their resistance to these agents. However, treatment of T24 cells with a combination of anti-Fas MoAb and doxorubicin resulted in a synergistic cytotoxic effect. In addition, the doxorubicin-resistant T24 cells were sensitive to treatment with a combination of anti-Fas MoAb and doxorubicin. Synergy was also achieved in three other bladder carcinoma cell lines and four freshly derived human bladder carcinoma cells. Treatment with anti-Fas MoAb in combination with epirubicin or pirarubicin also resulted in a synergistic cytotoxic effect on T24 cells. The mechanisms of synergy were examined. Anti-Fas MoAb did not affect the intracellular accumulation of doxorubicin, the expression of P-glycoprotein, or the expression of the antioxidant glutathione S-transferase-pi mRNA. However, treatment with doxorubicin enhanced the expression of Fas on T24 cells.
Conclusions:
This study demonstrated that treatment of bladder carcinoma cells with doxorubicin sensitized the cells to lysis by anti-Fas MoAb. The synergistic effect obtained with established doxorubicin-resistant bladder carcinoma cells and freshly isolated bladder carcinoma cells suggests that drug-resistant bladder carcinoma cells can be sensitized by doxorubicin to Fas- and Fas ligant-mediated cytotoxicity by lymphocytes. Furthermore, the sensitization required low concentrations of doxorubicin, thus supporting the in vivo application of a combination of chemotherapy and immunotherapy in the treatment of drug-resistant and/or immunotherapy-resistant bladder carcinoma.
Insights
Doxorubicin sensitizes drug-resistant bladder cancer cells to Fas-mediated apoptosis. This combination therapy, using low-dose doxorubicin, offers a promising approach for treating resistant bladder carcinoma.
Area of Science:
- Oncology
- Immunology
- Pharmacology
Background:
- Bladder carcinoma often exhibits resistance to chemotherapy, necessitating alternative treatment strategies.
- Immunotherapy, utilizing Fas antigen (Fas) and Fas ligand, offers a potential approach to overcome drug resistance.
- Anti-Fas monoclonal antibody (MoAb) can induce apoptosis in Fas-expressing cells, similar to Fas ligand.
Purpose of the Study:
- To investigate the sensitivity of bladder carcinoma cells to anti-Fas MoAb-mediated cytotoxicity.
- To determine if anticancer agents can synergize with anti-Fas MoAb to enhance cytotoxicity.
- To explore the potential of combining chemotherapy with immunotherapy for drug-resistant bladder cancer.
Main Methods:
- Cytotoxicity was assessed using a 1-day microculture tetrazolium dye assay.
- Synergistic effects were evaluated through isobolographic analysis.
- Mechanisms of synergy were investigated by examining doxorubicin accumulation, P-glycoprotein expression, and glutathione S-transferase-pi mRNA levels.
Main Results:
- T24 human bladder carcinoma cells expressed Fas but were resistant to anti-Fas MoAb alone.
- Combination therapy with anti-Fas MoAb and doxorubicin demonstrated synergistic cytotoxicity in T24 cells and other bladder cancer lines.
- Doxorubicin treatment enhanced Fas expression on T24 cells, contributing to sensitization.
- Synergistic effects were also observed with epirubicin and pirarubicin in combination with anti-Fas MoAb.
Conclusions:
- Doxorubicin sensitizes bladder carcinoma cells, including drug-resistant and freshly isolated cells, to Fas-mediated apoptosis.
- The combination of doxorubicin and anti-Fas MoAb shows potential for treating drug-resistant bladder carcinoma.
- Low-dose doxorubicin requirement supports the in vivo application of this combined chemotherapy and immunotherapy strategy.