Major Histocompatibility Complex Class II+ Invariant Chain Negative Breast Cancer Cells Present Unique Peptides that Activate Tumor-specific T Cells from Breast Cancer Patients

摘要

The major histocompatibility complex (MHC) class II-associated Invariant chain (Ii) is present in professional antigen presenting cells where it regulates peptide loading onto MHC class II molecules and the peptidome presented to CD4⁺ T lymphocytes. Because Ii prevents peptide loading in neutral subcellular compartments, we reasoned that Ii⁻ cells may present peptides not presented by Ii⁺ cells. Based on the hypothesis that patients are tolerant to MHC II-restricted tumor peptides presented by Ii⁺ cells, but will not be tolerant to novel peptides presented by Ii⁻ cells, we generated MHC II vaccines to activate cancer patients' T cells. The vaccines are Ii⁻ tumor cells expressing syngeneic HLA-DR and the costimulatory molecule CD80. We used liquid chromatography coupled with mass spectrometry to sequence MHC II-restricted peptides from Ii⁺ and Ii⁻ MCF10 human breast cancer cells transfected with HLA-DR7 or the MHC Class II transactivator CIITA to determine if Ii⁻ cells present novel peptides. Ii expression was induced in the HLA-DR7 transfectants by transfection of Ii, and inhibited in the CIITA transfectants by RNA interference. Peptides were analyzed and binding affinity predicted by artificial neural net analysis. HLA-DR7-restricted peptides from Ii⁻ and Ii⁺ cells do not differ in size or in subcellular location of their source proteins; however, a subset of HLA-DR7-restricted peptides of Ii⁻ cells are not presented by Ii⁺ cells, and are derived from source proteins not used by Ii⁺ cells. Peptides from Ii⁻ cells with the highest predicted HLA-DR7 binding affinity were synthesized, and activated tumor-specific HLA-DR7⁺ human T cells from healthy donors and breast cancer patients, demonstrating that the MS-identified peptides are bonafide tumor antigens. These results demonstrate that Ii regulates the repertoire of tumor peptides presented by MHC class II⁺ breast cancer cells and identify novel immunogenic MHC II-restricted peptides that are potential therapeutic reagents for cancer patients.