Interferon alpha (IFN-alpha), an important physiological immunomodulator, possesses direct cytotoxic and cytostatic effects on tumour cells, antiangiogenic effects, and activates anti-tumour immunity. Herein the human lung carcinoma cell line A549, a model of NSCLC in vitro, was used to pursue the effect of IFN-alpha on A549 cell proliferation and differentiation together with the effect on protein expression and activity of three ATP-transporters mediating multi-drug resistance (MDR). IFN-alpha significantly inhibited the proliferation of A549 cells which was not connected with arrest in a particular cell cycle phase. IFN-alpha-mediated differentiation of A549 was observed based on an increase in alkaline phosphatase activity. Simultaneously, IFN-alpha increased the expression and activity of ATP-transporters mediating MDR. The IFN-alpha down-regulation of NSCLC cell proliferation was accompanied by a potential of cells to exclude potential therapeutic substances such as chemotherapeutic agents.