Abstract: Objective: To study the function of apoptosis induction of arsenic trioxide (As₂O₃) in Ewing sarcoma cell line RD-ES and down-regulation of EWS-FLi1 fusion protein in vitro. Methods: Apoptosis was assessed by means of MTT, morphologic study, TUNEL assay and flow cytometry. RTPCR was used to investigate the expression of c-myc gene at mRNA level. Expression of the EWSFLi1 fusion protein in Ewing sarcoma was assessed by means of immunocytochemistry and western blotting. Results: The growth of Ewing sarcoma cell line was markedly inhibited by As₂O₃ in vitro. When the RD-ES cell line was cultured with 2umol/L As₂O₃, typical morphological apoptotic changes were found in morphological observation after cultured with 2 mol/L As₂O₃ for 72h. The positive rate of TUNEL assay was 43.2%. The result of RT-PCR and immunocytochemistry and western blotting studies revealed that less c-myc mRNA and EWS-Flil fusion protein expressed as time going on in the early stage of apoptosis induced by As₂O₃ in RD-ES cell. Conclusion: As₂O₃ can inhibit growth and induce apoptosis of Ewing sarcoma RD-ES cell line in vitro, and the mechanism is associated with the change of mitochondria transmembrane protiential and loss of EWS-FLi1 fusion protein which caused the inhibition of c-myc gene expression.