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Invasive tumor cells and their microenvironments are enriched with a broad spectrum of different proteases. Legumain, a novel asparaginyl endopeptidase, has been observed to be highly expressed in several types of solid tumors. However, there is no data available identifying the relationship of legumain expression and clinicopathologic or biological variables in invasive breast cancer. For the first time, the prevalence of legumain expression in invasive breast cancer (n = 432) and non-neoplastic breast tissues (n = 128) was investigated by immunohistochemistry. Three staining patterns were observed in the cytoplasm: diffuse positivity, tiny dots and vesicles. Whereas vesicular positivity in the majority of tumor cells was significantly correlated to an adverse outcome, cytoplasmic and dot-like staining showed no prognostic effect. Vesicular positivity was observed in 24% of carcinomas, but only in one case of non-neoplastic breast tissue (<1%; proliferative mastopathy). This staining pattern was found to be independent of other factors analysed as grading, nodal status or HER2 expression. Besides being of prognostic value, legumain might prove to be an important predictive factor in breast cancer, since its unique cleavage specificity is already used in prodrug activation strategies.