Temozolomide (TMZ) is widely used as a chemotherapeutic agent in the treatment of glioma; however, the development of drug resistance remains a major obstacle in the effective treatment of glioblastoma. Increasing evidence has indicated that microRNAs (miRs) are involved in the drug resistance of glioma; however, the role of miR‑186‑5p in the TMZ resistance of glioblastoma remains unknown. In the present study, the role of miR‑186‑5p in the resistance of glioblastoma to TMZ was investigated. mRNA and protein expression levels were detected via reverse transcription‑quantitative PCR and western blot analysis, respectively. It was determined that miR‑186‑5p was significantly downregulated in glioblastoma tissues and cell lines. Additionally, the expression of miR‑186‑5p was decreased, whereas that of Twist1 was upregulated during the development of drug resistance in glioma cells. The introduction of miR‑186 into glioblastoma cells via transfection decreased the proliferation and TMZ resistance of glioblastoma cells, as determined via 5‑ethynyl‑2'‑deoxyuridine and Cell Counting Kit‑8 assays, whereas the inhibition of miR‑186‑5p induced opposing effects. Furthermore, luciferase reporter and expression rescue assays revealed that miR‑186‑5p bound to the 3'‑untranslated region of Twist‑related protein 1 (Twist1). In conclusion, the present study demonstrated that downregulation of miR‑186‑5p may contribute to the proliferation and drug resistance of glioblastoma cells via the regulation of Twist1 expression. These results suggested that miR‑186‑5p may be a novel therapeutic target in the treatment of glioblastoma.