Characterized by invasive growth and infiltrative dissemination, glioma is poorly diagnosed and prognosed at present. The mammalian diaphanous-related formin 1 (mDRF1), which is involved in a number of actin-related biological processes, has been found to participate in the process of invasion and metastasis in human breast cancer cells and to show abnormal expression under pathological conditions. However, the role of mDRF1 in glioma is not clear. In this study, we carried out a comprehensive analysis of the effects of mDRF1 on human glioma. We used siRNA to knock down mDRF1 expression in highly invasive U87 malignant glioma (MG) cells and examined the changes in cell proliferation, apoptosis, invasion and migration. Atomic force microscopy was used to examine invadopodia formation. Immunohistochemical and immunocytochemical assays were used to analyze the cellular localization and the expression levels of mDRF1 in human glioma tissue and in the U87 MG cells. Following the transfection of U87 MG cells with siRNA-mDRF1, their in vitro proliferation was significantly decreased, apoptosis was markedly increased, and invasion and metastasis were significantly inhibited. The results from atomic force microscopy revealed that invadopodia were formed at leading the edge of the U87 MG cells. However, following the silencing of mDRF1 by siRNA, the edge of the cells became smooth and the invadopodia disappeared. For in vivo experiments, nude mice were transplanted with tumor cells and then treated with siRNA-mDRF1. The results revealed that treatment with siRNA-mDRF1 significantly inhibited tumor growth and led to a decrease in the weight of the transplanted tumor. In conclusion, our data demonstrate that mDRF1 is highly expressed in human glioma tissue. The knockdown of mDRF1 in U87 MG cells led to a sharp decline in their invasive and metastatic ability, which effectively reduced the spread of glioma cells into the surrounding areas. To our knowledge, this is the first report showing that mDRF1 is a promising target for the treatment of malignant gliomas.