Background: MGMT gene promoter methylation in tumor tissue of gliomas is now recognized as a predictive biomarker for response to temozolomide. Up to date there is lack of studies investigating MGMT methylation status in the circulating DNA of glioma patients undergoing chemotherapy with temozolomide. In this prospective study on a cohort of patients with gliomas of different grades of malignancy we aimed to: (1) evaluate the concordance between MGMT methylation status in tumor tissue and plasma; (2) monitor MGMT methylation status in plasma before and during temozolomide, and explore its value as a predictive biomarker. Methods: We enrolled 58 patients who underwent surgical resection, followed by radiotherapy and/or temozolomide. Blood samples were collected at baseline and every 3 cycles of temozolomide up to 12 months. MGMT promoter methylation status was assessed in both paraffin-embedded tumor tissue and plasma by real-time PCR. Results: MGMT promoter methylation status was concordant in tumor tissue and plasma at baseline in 42 out of 48 patients (87.5%, Kappa=0.75, 95%CI: 0.57-0.93). When considering glioblastomas only we obtained a 93.7% concordance and a Kappa of 0.87 (95%CI: 0.70-1.00). Mortality was higher for patients with MGMT unmethylated promoter both in tumor tissue (HR=2.21, 95%CI: 0.99-4-95) and plasma (HR=2.19, 95%CI: 1.02-4.68). Progression-free survival was shorter for patients with unmethylated MGMT promoter both in tumor tissue (HR=2.30, 95%CI: 1.19-4.45) and plasma (HR=1.77, 95%CI: 0.95-3.30). The cumulative incidence of unmethylated MGMT promoter in plasma was 58% at baseline and reached virtually 100% at 12 months. Conclusions: The assessment of MGMT promoter methylation in plasma can be useful when adequate tumor tissue (i.e. in stereotactic biopsy or in inoperable tumors) is not available. Moreover, the analysis of MGMT promoter methylation in plasma could be particularly useful to monitor methylated patients, helping to establish when the tumor switches from a methylated to an unmethylated status, thus predicting the emerging of a treatment resistance.