Induction of Chemoresistance by Transfection of the Full‐Length Canine mdr1 Gene in Canine Cell Lines

Abstract

Introduction: In the chemotherapy for treatment of lymphoid tumors in dogs, myelosuppression is a frequently encountered dose‐limiting factor. One possible approach to overcome myelosuppression is induction of chemoresistance in hematopoietic stem cells through expression of the mdr1 gene. A full‐length canine mdr1 cDNA clone was isolated in our laboratory. The present study was carried out to assess whether it confers multidrug resistance in canine cell lines. Materials and methods: The full‐length canine mdr1 cDNA was inserted into an expression plasmid vector. A canine mammary tumor cell line (CIPP) and osteosarcoma cell line (OOS) were transfected with the canine mdr1 expression vector. Expression of P‐gp was examined by immunoblotting. Function of ATP‐dependent drug efflux was measured by flow cytometric analysis using Rhodamine 123. Sensitivity to chemotherapeutic drugs was shown by estimation of 50% inhibitory concentrations (IC50) of vincristine or doxorubicin. Results: Immunoblotting of the transfected cells revealed a strong band of P‐gp detected by a monoclonal antibody directed to P‐gp. Rhodamine 123 efflux test showed an apparent drug efflux activity in the transfected cells. From the IC50 of the chemotherapeutic agents, the transfected cells showed a remarkable increase (20 to 60‐fold) in the resistance to vincristine or doxorubicin. Conclusion: Transfection of canine mdr1 gene induced P‐gp expression and strong drug resistance in canine cell lines.