MAGE-4 is considered as an attractive cancer-testis (CT) antigen for tumour immunotherapy, and it is overexpressed in oesophageal carcinoma (EC). To identify MAGE-4-derived HLA-A2 restricted epitopes, native peptides and their analogues were predicted with prediction programs. The native peptide, p286 (KVLEHVVRV), and its analogues, p286-1Y2L and p286-1Y2L9L, showed potent binding affinity and stability towards HLA-A*0201 molecule. Cytotoxic T lymphocytes (CTLs) induced by p286-1Y2L9L could release IFN-γ in ELISPOT assay. In cytotoxicity assay, p286-1Y2L9L showed the capability to induce specific CTLs which could lyse the target cancer cells from both PBMCs of healthy donors and HLA-A2.1/Kb transgenic mice. Our results indicated that the peptide p286-1Y2L9L could serve as a novel candidate epitope to develop peptide vaccines against oesophageal carcinoma.