LANDscape MApping of Epitopes and T Cell Receptors for Selected Cancers

The cloning of genes encoding the T cell receptor (TCR), the identification of tumor-associated antigens and the subsequent characterization of the first HLA-restricted T cell-defined antigenic epitope, were key findings illustrating direct recognition of cancer cells by T cells. These discoveries provided a mechanistic foundation for ensuing work examining the dynamic nature of lymphocyte-dependent recognition and elimination of neoplastic cells. Furthermore, preclinical and clinical investigations illustrate an important role for T cell mediated anti-tumor immunity in human disease, and have characterized the complexity of cancer-associated immune responses that are not always sufficient for tumor elimination. Importantly however, therapeutic targeting of the immune system has demonstrated the power of immunomodulatory drugs for the restoration of anti-tumor immune responses for cancer treatment.

Presence of lymphocytes in a variety of human cancers is well documented, and T cells isolated from tumors that recognize cancer antigens can be harnessed for effective treatment. T cells isolated from patient tumors can be ex vivo expanded, and reinfused back into patients in a regime of cellular therapy termed adoptive cell transfer (ACT). This ACT therapy can be further directed to specific tumor antigens with the genetic manipulation of T cells to express TCR recognizing known p-HLA epitopes with dominant expression on cancer cells. However, the peptide-HLA (p-HLA) epitope landscape of tumor associated antigens, and their cognate TCR are not well described, and those which have been described are primarily limited to the class I HLA-A\*02:01 allele dominant only in European-Caucasian populations. The purpose of this study is to further document the cancer epitope landscape and provide a comprehensive characterization of TCR specificities in a range of malignancies, for a wide variety of class I and class II HLA alleles. In addition, we aim to elucidate not only TCR repertoires important for anti-tumor immunity, but further clarify the role antigen presenting cells play in shaping these T cell repertoires.

The objectives involve the identification of cancer-associated/specific antigen p-HLA epitopes and their cognate TCR, and the subsequent structural and functional characterization these TCR. To meet this objective, immune cells of T, B and myeloid lineage will be analyzed. Phenotypic characterization of these cell subsets will be performed using standard immunological procedures such as immunofluorescence, immunohistochemistry, ELISA, ELISpot, qRT-PCR, analytic cytometry, CyTOF (cytometry time of flight), and in vitro stimulation. To relate immune responses to cancer cell intrinsic biology, RNA and DNA will be sequenced to identify cancer cell transcriptome and mutations, as well as T cell receptor unique sequences. Results from all laboratory analysis will be combined with relevant clinical data. Any confirmation of diagnosis, tissue types and other clinical data will be provided as available from the pathologists of the relevant disease site at UHN.

An incomplete understanding of T cell responses to cancer impedes the development of more effective immunotherapeutics. Discovery using tumor specimens from cancer patients will clarify how the complexity of the tumor environment shapes T cell specificity to induce effective immune responses and facilitate our development of better immune modulating therapeutics.

• Characterizing tumor-antigen specific TCR repertoire diversity across diverse HLA alleles, and further provide a comprehensive functional analysis that identifies immunodominant epitopes important for tumor control [Time frame: 63 months]