Natural antigen processing and presentation of antigen is thought to be important for the generation of a broad functional repertoire of antigen-specific T cells. In this study, the T-cell repertoire to an immunodominant human leukocyte antigen A2 (HLA-A2) binding peptide epitope of HER-2/neu, p369-377, was examined in a patient following immunization with a peptide-based vaccine consisting of helper peptides encompassing HLA-A2 peptide epitopes. The responding T-cell repertoire generated was both phenotypically and functionally diverse. A total of 21 p369-377 clones were generated from this patient. With the exception of two clones, all clones were CD3(+). Sixteen of the clones were CD8(+)/CD4(-). Five of the clones were CD4(+)/CD8(-), despite being generated with an HLA-A2 binding peptide. Nineteen of 21 of clones expressed the alpha beta-T-cell receptor (TCR). The remaining two clones expressed the gamma delta T-cell response (TCR). Selected alpha beta-TCR clones, both CD8(+) and CD4(+), could lyse HLA-A2 transfected HER2 overexpressing tumor cells and p369-377-loaded B-lymphoblastic cell line. In addition to their lytic capabilities these clones could be induced to produce interferon-gamma (IFN-gamma) specifically in response to p369-377 peptide stimulation. The 2 gamma delta-TCR clones expressed CD8 and lysed HLA-A2(+) HER-2/neu(+) tumor cells, but not HLA-A2(-) HER-2/neu(+) tumor cells. One of gamma delta-TCR clones also released IFN-gamma directly in response to p369-377 stimulation. These results suggest that a tumor antigen TCR, directed against a specific epitope, can be markedly polyclonal at multiple levels including CD4/CD8 and TCR.