To study the resistant mechanisms of cisplatin in human lung adenocarcinoma cell line A₅₄₉DDP. A₅₄₉DDP cells was established by stepwise increasing concentration of cisplatin (CDDP) in medium. Interstrand crosslinked DNA (ICL) was measured by ethidium bromide fluorescence assay. The intracellular and intranuclear accumulation of cisplatin was measured by atomic absorption spectrometry. The removal of GS-X was determined by FCM and fluorescence microscopy. Results: The As49DDP cell line was 8.9-fold resistance relative to the parental A₅₄₉ cell line. The formation of ICL in A₅₄₉ was 6.28 times higher than that in A₅₄₉DDP cells. The intracellular and intranuclear accumulation of cisplatin in A₅₄₉ cells was 5.9 times and 4.1 times higher than that in A₅₄₉DDP cells, respectively. The ability of GS-X pump pumped GS-X complex (GS-Pt) in A₅₄₉DDP cells was higher than that in A₅₄₉. The repair rate in A₅₄₉DDP cells was 2 times higher than that in A₅₄₉. Conclusions: Decreased accumulation and increased export of cisplatin might be the main mechanism of cisplatin resistant A₅₄₉DDP cells while the enhanced repair capacity of DNA may play a role in CDDP resistance.