Identification of antigenic epitopes recognized by tumor infiltrating lymphocytes in high grade serous ovarian cancer by multi-omics profiling of the auto-antigen repertoire.

Immunotherapeutic strategies aimed at enhancing tumor cell killing by tumor-specific T cells hold great potential for reducing tumor burden and prolonging survival of cancer patients. Although many potential tumor antigens have been described, identifying relevant targets when designing anti-cancer vaccines or targeted cell therapies remains a challenge. To identify novel, potentially immunogenic candidate tumor antigens, we performed integrated tumor transcriptomic, seromic, and proteomic analyses of high grade serous ovarian cancer (HGSC) patient tumor samples. We identified tumor neo-antigens and over-expressed antigens using whole exome and RNA sequencing and examined these in relation to patient-matched auto-antibody repertoires. Focusing on MHC class I epitopes recognized by CD8+ T cells, HLA-binding epitopes were identified or predicted from the highly expressed, mutated, or auto-antibody target antigen, or MHC-associated peptides (MAPs). Recognition of candidate antigenic peptides was assessed within the tumor-infiltrating T lymphocyte (TIL) population expanded from each patient. Known tumor-associated antigens (TAA) and cancer/testis antigens (CTA) were commonly found in the auto-antibody and MAP repertoires and CD8+ TILs recognizing epitopes from these antigens were detected, although neither expression level nor the presence of auto-antibodies correlated with TIL recognition. Auto-antibodies against tumor-mutated antigens were found in most patients, however, no TIL recognition of the highest predicted affinity neo-epitopes was detected. Using high expression level, auto-antibody recognition, and epitope prediction algorithms, we identified epitopes in 5 novel antigens (MOB1A, SOCS3, TUBB, PRKAR1A, CCDC6) recognized by HGSC patient TILs. Furthermore, selection of epitopes from the MAP repertoire identified 5 additional targets commonly recognized by multiple patient TILs. We find that the repertoire of TIL specificities includes recognition of highly expressed and immunogenic self-antigens that are processed and presented by tumors. These results indicate an ongoing autoimmune response against a range of self-antigens targeted by HGSC TILs.

Mutually exclusive mutation profiles define functionally related genes in muscle invasive bladder cancer.

Muscle Invasive bladder cancer is known to have an abundance of mutations, particularly in DNA damage response and chromatin modification genes. The role of these mutations in the development and progression of the disease is not well understood. However, a mutually exclusive mutation pattern between gene pairs could suggest gene mutations of significance. For example, a mutually exclusive mutation pattern could suggest an epistatic relationship where the outcome of a mutation in one gene would have the same outcome as a mutation in a different gene. The significance of a mutually exclusive relationship was determined by establishing a normal distribution of the conditional probabilities for having a mutation in one gene and not the other as well as the reverse relationship for each gene pairing. Then these distributions were used to determine the sigma-magnitude of standard deviation by which the observed value differed from the expected, a value that can also be interpreted as the 'p-value'. This approach led to the identification of mutually exclusive mutation patterns in KDM6A and KMT2D as well as KDM6A and RB1 that suggested the observed mutation pattern did not happen by chance. Upon further investigation of these genes and their interactions, a potential similar outcome was identified that supports the concept of epistasis. Knowledge of these mutational interactions provides a better understanding of the mechanisms underlying muscle invasive bladder cancer development, and may direct therapeutic development exploiting genotoxic chemotherapy and synthetic lethality in these pathways.