Purpose-Alveolar rhabdomyosarcoma (ARMS) frequently contains the fusion transcription factor PAX3/FKHR. Therefore, clinical studies have been initiated to utilize the PAX3/FKHR translocation point area as a peptide vaccine against ARMS. Our study was directed at identifying antigenic T-lymphocyte epitopes at the PAX3/FKHR translocation point area.Experimental design-The peptide sequence surrounding the PAX3/FKHR translocation point was evaluated by MHC binding algorithms for potential T-lymphocyte antigenic epitopes (class I molecules HLA-A1, -A2 and -A3; class II molecules HLA-DR1, -DR4 and -DR7). Using in vitro techniques, dendritic cells loaded with PAX3/FKHR peptides were used to stimulate na¿ve Tlymphocytes. T-lymphocyte activity was then evaluated by 51 Cr release and 3 H-thymidine uptake assays.Results-Only one HLA-A3-restricted epitope was predicted by the algorithms. The peptide was prepared and tested for its ability to stimulate na¿ve cytotoxic T-lymphocytes (CTLs). Unfortunately, the peptide was unsuccessful at stimulating na¿ve CTL. However, induction of na¿ve helper Tlymphocytes (HTL) to recognize and respond to the PAX3/FKHR translocation peptide was successful. Yet, this HTL peptide activity did not translate into recognition of PAX3/FKHRcontaining ARMS tumor cells.Conclusions-It appears that the fusion area of PAX3/FKHR may not be a good source of antigenic anti-tumor peptide epitopes. These results raise serious concerns about the success and applicability of future peptide-based vaccine immunotherapy directed at the PAX3/FKHR translocation point.