Granzyme B (GrB) is the prototypic member of a serine protease family primarily used by cytotoxic lymphocytes to kill target cells. We report here that, by immunohistochemical staining of paraffin-embedded tumour sections, GrB protein was unexpectedly detected in malignant cells of a subset of breast cancers and their adjacent reactive endothelial and mesenchymal cells in which endogenous retinoblastoma protein (pRB) is overexpressed. The identity of the endogenous GrB was further confirmed experimentally in RB-deficient breast carcinoma cell culture upon overexpression of ectopic pRB. Our finding extends the recent paradigm-shifting trend for a more diverse biological role of granzyme B, and might provide a rational basis for exploring its potential prognostic value in a variety of human cancers.