Date of Award

2019

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Graduate Group

Cell & Molecular Biology

First Advisor

John M. Maris

Abstract

Recent breakthroughs in biotechnology and immunology have allowed the immune system to be harnessed in the cure of some cancers. Thus far clinical successes using immunotherapy have largely been limited to liquid tumors and solid tumors bearing high mutational burden, while the majority of cancers currently remain poor candidates for these therapies. Here we demonstrate a proof-of-principle in addressing one of the main bottlenecks currently impeding widespread application of immunotherapy, the paucity of tumor specific targets in low mutational solid tumors, and the engineering of T cells to recognize tumor-specific self-antigens. Using neuroblastoma, an aggressive, low mutational, cold tumor, we developed a process combining proteomics of MHC-presented antigens with computational methods and genomic/transcriptomic analyses of normal and tumor tissues, to identify and prioritize tumor-specific antigens derived from lineage restricted oncogenes crucial to tumor biology (derived from PHOX2B, IGFBPL1, HMX1, TH, CHRNA3, and GFRA2). We then developed methodologies for engineering CAR and TCR receptors, using phage display and single-cell sequencing of functionally-expanded antigen-specific T cells, respectively. Using these methods, we demonstrate specific binding of CAR and TCR receptors to tumor antigens and potent killing of neuroblastoma cells using engineered T cells. We describe new methods of multiplexing targets for accelerated T cell engineering and propose a vision for personalized target discovery and T cell engineering.

Additionally, we explore the ability of the immune system to recognize and eliminate neoantigens arising from early cancer driver genes. We developed a model of immunoediting using TCGA data to measure the underrepresentation of HLA alleles in tumors predicted to present neoantigens as a measure of immunoediting. We show that the most common tumor driver mutations such as KRAS G12D and BRAF V600E are immunogenically silent, such that peptides derived from these mutations are not presented on HLA in the vast majority of the population, thus conferring additional evolutionary advantages to these mutations in the evasion of the immune system in addition to their oncogenic properties. We show that HLA alleles offer varying degrees of immunogenic protection against cancer, and thus propose HLA as a factor contributing to cancer susceptibility.

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