Glioblastoma multiforme (GBM) is one of the most aggressive types of brain malignancy, with resistance to chemotherapy being a primary treatment obstacle. ATPase copper transporting beta (ATP7B) is involved in multidrug resistance; however, its expression in GBM remains to be evaluated. In the present study, GBM specimens from 79 patients who underwent gross total tumor removal followed by concomitant temozolomide (TMZ) chemotherapy and radiotherapy were assessed immunohistochemically. The association between the overall survival times of patients and the expression of ATP7B in neoplastic cells was evaluated. In 12/79 tumors (15.2%) >10% of neoplastic cells were immunohistochemically-positive for ATP7B, and categorized as high-ATP7B GBM. In the remaining 67 tumors (84.8%) the rate of ATP7B-positive cells was <10% and recorded as low-ATP7B GBM. The median overall survival times of patients with high-and low-ATP7B GBM were 14.6, and 24.7 months, respectively. High expression of ATP7B was identified to be associated with shorter overall survival times (hazard ratio, 0.452; 95% confidence interval, 0.206-0.994; P=0.048). Of the 79 patients, 12 underwent a second operation due to recurrence. These tissue samples were also subjected to immunohistochemical study. The ATP7B positivity rate of tumor cells obtained during the second surgery was significantly higher compared with that in the first surgery (9.17 +/- 2.56 vs. 2.75 +/- 0.55%; P=0.008). In addition, two ATP7B-transfected GBM cell lines were identified to be significantly resistant (3.8- and 1.7-fold, respectively) to TMZ compared with the control cell line. The findings of the present study suggest that ATP7B influences GBM resistance to TMZ.