Multidrug resistance (MDR) due to the expression of the MDR1 gene and its P-glycoprotein (Pgp) product is a major factor in the prognosis and clinical outcome of patients with refractory lymphomas and other malignancies. The aim of our study was to establish a lymphoma, cellular system where a de novo acquisition of multidrug resistance is specifically related to overexpression of a transgenic, human MDR1. A multidrug sensitive lymphoma cell line (LM1) was established from a sporadic T-cell lymphoma of BALB/c mouse and was transduced by a retroviral vector containing the human MDR1 cDNA. The resultant cell variant (LM1/MDR) was characterized in comparison to the parental LM1 cells. The LM1/MDR cell variant is cross-resistant to DOX, COL, ACT D and VBL. This cell variant expresses the human MDR1 and exhibits de novo functional Pgp activity that can be blocked by the Pgp-modulators VRP and KT-5720. The acquired MDR of LM1/MDR is not accompanied with gene amplification, alternative splicing or up-regulation of the murine endogenous mdr1a, mdr1b, mrp1, mrp2 and mrp3 transporter-genes. Therefore, the acquired MDR is, specifically, human MDR1-dependent as it has been found in malignant cells of most lymphoma patients. Moreover, this system can be used as a model to study MDR and the efficacy of drugs and modulators on malignant cells where human Pgp is a major factor of multidrug resistance.