The MUC1 protein was found to be up‐regulated in a spectrum of malignant tumors. T‐cell responses to the MUC1 extracellular tandem repeat array (TRA) were observed in murine models as well as in breast‐carcinoma patients. In the present study, we evaluated the anti‐tumor potential of HLA‐A2.1‐motif‐selected peptides from non‐TRA domains of the molecule. Peptide immunogenicity was examined in the D^b−/⁻ × β2 microglobulin (β2m) null mice transgenic for a modified HLA‐A2.1/D^b‐β2 microglobulin single chain (HHD mice). Our results show the existence of 3 novel HLA‐A2.1‐restricted MUC1‐derived cytotoxic T‐lymphocyte (CTL) epitopes. These peptides are processed and presented by the HHD‐transfected breast‐tumor cell line MDA‐MB‐157. Moreover, CTL induced by these 3 peptides show higher lysis of target cells pulsed with breast‐carcinoma‐derived peptides than of targets pulsed with normal breast‐tissue‐derived peptides. These data suggest an important role for non‐TRA MUC1‐derived peptides as inducers of a MHC‐restricted CTL reaction to a breast‐carcinoma cell line and patient‐derived tumor extracts. Int. J. Cancer 85:391–397, 2000. ©2000 Wiley‐Liss, Inc.