Background: Astrocytoma is the most common primary tumor of the brain. High-grade astrocytomas are aggressive tumors with poor prognosis. Biomarkers that predict patient outcome and response to a specific treatment will provide patients with personalized estimation of prognosis and customized therapeutic regimens with maximal effectiveness and minimal toxicity. Aim: The aim of this work was to explore the ability of the expression of IDH1 R132H, pSTAT3, and Tbx2 proteins to predict outcome of patients with high-grade astrocytoma. Materials and methods: The expression of IDH1 R132H, pSTAT3, and Tbx2 proteins was assessed by immunohistochemistry in 41 high-grade astrocytoma cases. They were correlated with patients’ clinicopathological features, recurrence-free survival, overall survival (OS), and response to therapy. Results: A significant direct association was observed between IDH1 R132H immunoreactivity and response to TMZ therapy (P=0.01), better progression-free survival (PFS) as well as OS (P<0.001). The expression of both Tbx2 and pSTAT3 proteins was inversely correlated with response to TMZ therapy (P=0.019 and 0.014, respectively). Survival analysis revealed that Tbx2 and pSTAT3 immunoreactivity was significantly associated with shorter PFS (P=0.021 and 0.047, respectively), as well as poor OS (P=0.005 and 0.007, respectively). Conclusion: IDH1 R132H protein expression and loss of Tbx2 and pSTAT3 proteins are associated with prolonged PFS and OS and better response to TMZ chemotherapy.