The nature of genetic alterations in bilateral breast cancer (BBC) associated with the distinctive development of a second primary tumor or a metastatic lesion is not clearly established. In this study, patterns of promoter methylation and gene copy number changes were assessed for their utility in the distinction of two types of BBC (synchronous and metachronous). Seven cases of synchronous and five cases of metachronous breast cancer tissues were used in X chromosome inactivation assay to assess the methylation pattern of human androgen receptor gene. X chromosome inactivation assay alone did not provide enough information to distinguish the genetic origins of synchronous and metachronous BBC. When four pairs of paraffin-embedded BBC tissues were used in cDNA array-based CGH with placenta DNA as a reference, higher DNA copy number changes were observed from metachronous pairs (9.0%) than from synchronous pairs (3.1%). From the two cases of metachronous pairs tested, 44 genes were found to be commonly modulated in gene copy numbers in a cancer detected later.