Background Gallbladder cancer (GC) is considered a relatively rare malignancy with extensively poor prognosis. To guide clinicians in selecting treatment options for GC patients, reliable markers predictive of poor clinical outcome are desirable. This study analyzed the correlation of NF-E2-related factor 2 (Nrf2), heme oxygenase-1 (HO-1), and multidrug resistance-related protein 3 (MRP3) in GC and their relationships to clinicopathologic features and survival. Material and Methods We immunohistochemically investigated 59 specimens of gallbladder adenocarcinoma tissues using Nrf2, HO-1, and MRP3 antibodies. Results There were significant correlations between the high level of Nrf2, HO-1, and MRP3 expression and the tumor differentiation, Nevin staging, and metastasis. Significant positive correlations were found between the expression status of Nrf2 and that of HO-1 and MRP3 (r = 0.38, P = 0.008 and r = 0.59, P < 0.001, respectively). High Nrf2 expression was significantly associated with shorter overall survival times in univariate analysis (log-rank test, P < 0.001), being also identified as an independent prognostic factor in multivariate analysis (P = 0.035). Conclusions Nrf2, HO-1, and MRP3 were associated with certain clinicopathologic parameters in GC. Evaluation of Nrf2 expression may be an important factor in identifying a poor prognostic group of GC.