Author ORCID Identifier
https://orcid.org/0000-0001-5664-9659
Date of Graduation
5-2024
Document Type
Dissertation (PhD)
Program Affiliation
Cancer Biology
Degree Name
Doctor of Philosophy (PhD)
Advisor/Committee Chair
Eduardo Vilar-Sanchez, MD, PhD
Committee Member
Alexandre Reuben, PhD
Committee Member
Paul Scheet, PhD
Committee Member
Scott Kopetz, MD, PhD
Committee Member
Gregory Lizee, PhD
Committee Member
Linghua Wang, PhD
Abstract
Lynch Syndrome (LS) is the most common inherited colorectal cancer (CRC) syndrome. It constitutes the perfect model to understand DNA mismatch repair deficient (MMRd) carcinogenesis, which underlies 15% of early-stage CRC. LS patients develop MMRd tumors with high loads of shared neoantigens (neoAgs), which are recognized by the immune system. Previous research has concentrated on discovering neoAgs and their potential as targets for vaccines in LS patients. However, these studies have primarily identified shared neoAgs from cancers, lacking detailed information on targetable neoAgs present in precancerous lesions. Understanding this landscape of pre-cancer derived neoAgs is crucial for intercepting cancer development effectively. Furthermore, the T-cell receptor (TCR) landscape of LS patients, particularly those that recognize these neoags, has never been investigated. Thus, there is an excellent opportunity for exploring immune-based preventive strategies and gaining insights into the patterns of these recurrent neoAgs as well as the corresponding TCRs that recognize them.
Our neoAg study consisted of whole exome sequencing (WES) and mRNA sequencing (mRNAseq) on 69 colorectal lesions, that included 12 cancers and 8 advanced pre-cancers, 41 pre-cancers and 8 polyps without malignant potential (PWOMPs). Using this data, an in silico pipeline was developed, which predicted, ranked, and identified the top 100 most recurrent and immunogenic neoAgs that were then validated in vitro using ELISpot and tetramer assays. Our results showed that 65% of our top 100 predicted neoAgs were immunogenic in vitro, and were detected in 92% of cancers, 50% of advanced pre-cancers, and 29% of pre-cancers, while no neoAgs were detected in PWOMPs.
For the TCR assessment, I performed TCR sequencing of 277 PBMCs/blood samples from 102 LS survivors with history of MMRd cancers, 130 LS previvors and 45 controls. Moreover, 14 colorectal lesions were subjected to TCR sequencing. My results revealed that
LS survivors have significantly less diverse circulating TCR repertoires compared to previvors and controls. Also, we found that largely expanded MMRd CRC-specific public TCRβs are detectable in the peripheral blood of LS cancer survivors, while LS previvors and controls exhibited detectable levels of MMRd CRC-specific public TCRβs with medium and minimal expansion, respectively. Importantly, we developed a classification model capable of distinguishing LS individuals from controls with excellent performance, based on the presence of a Lynch-associated TCR signature. Among these Lynch-associated TCRβs, several were confirmed to exhibit specificity towards colorectal cancer and a neoantigen originating within these lesions. Finally, we identified a set of TCRβs with some ability to discriminate LS survivors from previvors, better than random chance.
In summary, this study is the first to describe neoAg generation in pre-cancers as well as the circulating TCR repertoires of LS patients. Our results move the field closer towards the development of a universal LS cancer-vaccine and the identification of TCR clones in the peripheral blood of LS carriers, that could serve as biomarkers of cancer.
Keywords
Lynch Syndrome, Neoantigen, T-cell Receptors, Colorectal Cancer, DNA Mismatch Repair Deficiency, cancer vaccine, TCR signatures, Immunogenomics
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