The PTEN protein is a negative regulator of the Akt pathway, leading to suppression of apoptosis and increased cell survival. Its role as a tumor-suppressor gene has been adequately substantiated, and homozygous mutations have been demonstrated in familial and sporadic cancers. In breast cancers, expression of PTEN protein is lost/reduced in 38% of cases. Somatic mutations are, however, rarely found. Our study was therefore designed to determine if differential methylation of the PTEN promoter region has a role in the transcriptional inactivation of the gene in invasive breast carcinomas. A total of 44 samples of invasive human breast cancer, 5 breast cancer cell lines and 16 samples of normal human breast tissue from young and elderly women were studied for methylation of the PTEN promoter by methylation-specific PCR and PTEN protein expression by immunohistochemistry. PTEN methylation occurred in 34% of breast cancers, and 60% of these samples were associated with loss of PTEN protein. Analyzed from a different perspective, 34% of breast cancers had reduced expression of PTEN and 60% had a methylated PTEN promoter. None of the breast cancer cell lines and normal breast tissues showed methylation. In summary, methylation of the PTEN promoter leads to PTEN inactivation in a subset of human breast cancers.