The PTEN gene, recently identified on chromosome 10q23, has been proposed to be a candidate tumor suppressor gene inactivated in multiple cancers including glial tumors. We investigated 47 glioblastomas (GBM), 14 anaplastic astrocytomas (AA), 6 non-pilocytic low-grade astrocytomas (LGA), 21 low-grade and anaplastic oligodendrogliomas (O) and oligoastrocytomas (OA), and 3 ependymomas (E) for mutation of the PTEN gene using denaturing gradient gel electrophoresis (DGGE) followed by DNA sequencing. These tumors have been previously screened for loss of heterozygosity (LOH) on chromosome 10q, p53 mutations and EGFR amplification. Overall, PTEN mutations, detected in 14 of 91 tumors, were present in 13 of 47 GBM and 1 of 14 AA. In contrast, mutations were absent in other glioma subtypes (0/30). In all informative cases, PTEN mutations occurred in tumors showing LOH on chromosome 10q, confirming the inactivation of this gene by a 2-hit mechanism. No correlation was observed between the presence of PTEN mutation and p53 mutation and EGFR amplification. Our results indicate that biallelic PTEN inactivation plays an important role in the pathogenesis of high-grade astrocytomas as a late event. Moreover, they suggest that PTEN alterations are equally involved in the 2 glioblastoma pathways defined by the presence of EGFR amplification and p53 mutation. Finally, correlation analysis with clinical data did not show that PTEN mutation was linked to survival of the patients.