Osteosarcoma is a primary bone tumour that generally occurs in the long extremity bones. Although the localised tumours are mostly manageable, the high-grade forms are often accompanied by metastasis, recurrence, and limited treatment options. T-cell-based immunotherapy can offer new possibilities for the treatment of osteosarcoma. It uses the body’s immune system to survey and target peptide antigens presented in complex with human leukocyte antigens (HLAs) on the tumour surface. Utilising in-depth immunopeptidomics of osteosarcoma, we can identify targets more specific to the tumour surface and exploit them in T-cell-based immunotherapy. Over the years, cancer testis antigens (CTAs) have gained attention as potentially valuable tumour-associated antigens. Since most normal cells don’t express them, CTAs are intriguing targets for T-cell-based therapies and cancer vaccines.
In this study, U2OS and SJSA1 cells were used as established osteosarcoma models. To confirm their HLA-I expression, flow cytometry was performed. Afterwards, using a pan HLA-I antibody (W6/32), we isolated peptides from a 5e7 pellet of each osteosarcoma cell line. The eluted peptides were subsequently analysed by LC-MS/MS on the Orbitrap Exploris 480 mass spectrometer. Data analysis was performed by PEAKS12.5 studio.
These two osteosarcoma cell lines expressed a total of 11 different alleles, including HLA-A*02:01, the most common HLA type in Caucasian populations. Collectively, these 11 HLA alleles cover over 56% of the world population. In total, >8400 for U2OS and >2400 for SJSA1 HLA-bound peptides (pHLAs) were identified. More than 90% of peptides were predicted to bind to their respective HLA-I alleles. Altogether, we found 28 different pHLAs across 9 different HLA-I alleles that were derived from the MAGE family, including MAGE1, MAGE4, and MAGD1, some of which are currently in clinical trials for other malignancies. Overall, our data shows that MAGE family-derived antigens are potential targets for T-cell immunotherapy in osteosarcoma.