A pipeline for identification and validation of tumor-specific antigens in a mouse model of metastatic breast cancer

ABSTRACT

Cancer immunotherapy continues to make headway as a treatment for advanced stage tumors, revealing an urgent need to understand the fundamentals of anti-tumor immune responses. Noteworthy is a scarcity of data pertaining to the breadth and specificity of tumor-specific T cell responses in metastatic breast cancer. Autochthonous transgenic models of breast cancer display spontaneous metastasis in the FVB/NJ mouse strain, yet a lack of knowledge regarding tumor-bound MHC/peptide immune epitopes in this mouse model limits the characterization of tumor-specific T cell responses, and the mechanisms that regulate T cell responses in the metastatic setting. We recently generated the NetH2pan prediction tool for murine class I MHC ligands by building an FVB/NJ H-2q ligand database and combining it with public information from six other murine MHC alleles. Here, we deployed NetH2pan in combination with an advanced proteomics workflow to identify immunogenic T cell epitopes in the MMTV-PyMT transgenic model for metastatic breast cancer. Five unique MHC I/PyMT epitopes were identified. These tumor-specific epitopes were confirmed to be presented by the class I MHC of primary MMTV-PyMT tumors and their T cell immunogenicity was validated. Vaccination using a DNA construct encoding a truncated PyMT protein generated CD8 + T cell responses to these MHC class I/peptide complexes and prevented tumor development. In sum, we have established an MHC-ligand discovery pipeline in FVB/NJ mice, identified and tracked H-2D^q/PyMT neoantigen-specific T cells, and developed a vaccine that prevents tumor development in this metastatic model of breast cancer.

KEYWORDS:

• Tumor immunology
• breast cancer
• MMTV-PyMT
• MHC Class I
• peptide prediction
• NetH2pan

Acknowledgments

We would like to thank Dr. Lauren Zenewicz and Alisha Chitrakar for technical assistance with the tetramer staining, Sean Osborn and Steven Cate for providing sequencing services, and the Flow Cytometry Core facilities at OUHSC and University of Utah. This work was supported by funding from a Department of Defense Breast Cancer Research Program Innovator and Scholar Concept Award (W81XWH-12-1-0499 to ALW), a Susan G. Komen Leadership Award (SAC160078 to ALW), a NIH NRSA NIAID Training Grant (T32AI007633 to CID), a NIH R01 Grant (CA205348 to WRC),and the Oklahoma Center for Advancement of Science and Technology Grant (HR16-085 to WRC).

Declaration of Interest

W.H. Hildebrand is chief scientist at, has ownership interest in, and is a consultant/advisory board member for Pure MHC. W.R. Chen has ownership interest in and is a member of the Board of Directors for Immunophotonics, Inc. No potential conflicts of interest were disclosed by the other authors.

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Additional information

Funding

This work was supported by the National Institute of Allergy and Infectious Diseases [T32AI007633];National Institutes of Health [CA205348];Oklahoma Center for the Advancement of Science and Technology [HR16-085];Susan G. Komen [SAC160078];U.S. Department of Defense [W81XWH-12-1-0499].