ANTIGENICITY OF FUSION PROTEINS FROM SARCOMA-ASSOCIATED CHROMOSOMAL TRANSLOCATIONS

Worley BS, Goletz TJ, Helman LJ, Berzofsky, JA (National Institutes of Health, Bethesda, MD 20892)

Synovial sarcoma (SS), clear cell sarcoma (CCS), and desmoplastic small round cell tumor (DSRCT) are soft-tissue malignancies occurring primarily in adolescents and young adults. These tumors contain specific chromosomal translocations which fuse the 5’ region of one gene with the 3’ region of another, forming characteristic fusion proteins. The affected genes encode transcription factors, such that the resulting fusion product retains the DNA binding specificity of one gene while inappropriately activating or repressing transcription through the transactivation domain of the other gene. These translocations are unique to tumor cells, may be required for persistence, and thus provide targets for immunotherapy. We hypothesized that the fusion breakpoint sequences associated with SS, CCS, and DSRCT can serve as tumor specific neoantigens. To test this, we designed peptides representative of each type of fusion breakpoint and assessed the ability of the peptides to bind to various class I HLA molecules. This was done using immunofluorescent assays with HLA-transfected T2 cells and C1R cells. Two peptides designed from the SS breakpoint specifically bind HLA-B7, and a 10 amino acid minimal epitope has been identified for this interaction. Specific binding of an SS peptide and a CCS peptide to HLA-B27 has also been observed. Finally, a peptide designed from the DSRCT breakpoint specifically binds HLA-A3. To test whether these peptides can elicit specific T cell responses, normal HLA-specific donor lymphocytes were stimulated in vitro with autologous, monocyte-derived dendritic cells pulsed with peptide. T cell proliferation and cytokine production were measured. In the case of HLA-B7+ lymphocytes stimulated with SS peptide, one out of four donors produced IFN-g in an antigen-specific manner, indicating a specific T cell response. Efforts are currently underway to test other HLA-B7+ donor samples with SS peptide, as well as HLA-A3+ donor samples with DSRCT peptide. These results suggest that sequences in the fusion region of sarcoma-associated chimeras can bind class I HLA molecules and potentially serve as neoantigens. This may be useful for the development of novel immunotherapies for HLA-matched sarcoma patients with tumors bearing these translocations.