Borderline epithelial ovarian tumors (BEOTs) possess clinical and pathologic features intermediate between cystadenomas and cystadenocarcinomas. Although the clinical and pathologic characteristics of BEOTs are well described, the molecular aspects are poorly understood. Three regions of loss of heterozygosity (often referred to as allelic imbalance [AI] when identified by polymerase chain reaction) on chromosome 17p13, one of which includes the p53 gene, have been implicated in the development of ovarian and breast cancers. To provide evidence that genes in these regions also may be involved in the development of BEOTs, we undertook a detailed analysis of AI at all three loci in BEOTs from 21 patients. Seventeen of the BEOTs were serous and four were mucinous. Five of 21 tumors (24%) had AI at one or more loci. Four tumors had AI using the D17S695 marker, two of which showed AI only at this locus. In addition, three tumors exhibited AI at the D17S654 locus, one of which showed AI only at this locus. These data suggest that there may be two tumor suppressor genes distal to p53 involved in the development of at least a subset of BEOTs. Peritoneal implants from a subset of serous BEOTs also were evaluated for AI and were found to be concordant with the primary tumor in all cases. Their genetic similarity is consistent with the implantation theory of peritoneal spread of serous BEOTs in these cases.