Tumor cell invasion requires expression of degradative enzymes such as plasminogen activator, collagenase, and cathepsins. Cathepsin D, a lysosomal aspartic protease produced constitutively in human breast cancer cell lines, also has mitogenic activity in breast cancer cells. Additionally, high cathepsin D expression is associated with increased risk of metastasis in patients with node-negative breast cancer. Recently, a novel aspartic protease gene, ALP56 (aspartic-like protease 56kDa), has been identified. To examine possible interrelationships we quantitated ALP56 mRNA and cathepsin D mRNA in breast cancers using reverse transcription polymerase chain reaction. ALP56 mRNA expression was greater in cancers than in noncancerous tissues (p < 0.0001), as was expression of cathepsin D mRNA. ALP56 gene expression was dose-dependently down-regulated in T-47D breast cancer cells treated with estradiol, while cathepsin D was up-regulated. Expression of ALP56 mRNA in estrogen receptor (ER)-positive breast cancers was less than that in ER-negative cancers, and mRNA expression for ALP56 and cathepsin D did not correlate with one another. Thus ALP56 as well as cathepsin D may be a useful target molecule in breast cancer treatment.