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Nierves, Lorenz Arthur

University of British Columbia

Chemotherapy remains the primary treatment option for pediatric B cell acute lymphoblastic leukemia (B-ALL). Although effective, it is also harsh and indiscriminate; often leading to acute side effects or secondary diseases and malignancies that manifest later. There is, therefore, a need to seek out more specific therapeutic targets for pediatric B-ALL. Proteins on the cell surface are prime therapeutic targets due to their accessibility and role in cellular growth, nutrient uptake, and intercellular interactions. Furthermore, the emergence and efficacy of immunotherapy highlights the need for new therapeutic targets situated on the cell surface. However, current immunotherapy targets still utilize proteins that are also present on normal cells thus leading to challenges such as “on-target off-tumour” effects. To overcome these challenges, I was interested in the mass spectrometric exploration of cell surface proteolytic proteoforms—cell surface protein forms that arise due to the irreversible action of proteases. I hypothesized that the unique microenvironment in ALL leads to new and cancer-specific proteolytic proteofoms on the cell surface. Identification of these alternate proteoforms has the potential to expand the list of potential therapeutic targets for pediatric B-ALL. In this thesis, I developed a mass spectrometric workflow that allowed for the comprehensive assessment of the cell surface N terminome. After determining that current global mass spectrometric workflows do not sufficiently provide coverage to cell surface proteins—and more so—N termini, I optimized a biotin-based strategy to enrich for cell surface N termini. Biotinylation is a widely-used strategy for previously reported workflows that enriched for the cell surface proteome. I optimized it for cell surface N terminome enrichment by adopting the use of anti-biotin antibodies—which enabled direct detection of biotinylated peptides—and conducting additional work to improve the detectability of biotinylated peptides via mass spectrometry. The optimized cell surface N terminome strategy developed here was proven to be effective for relatively limited amounts of starting material. Finally, my work also included a multi-omic approach that comprehensively profiled the B-ALL microenvironment. Integration of the results from this study allowed for the monitoring of multiple analytes to describe changes and differences in the microenvironment.

Text

2024-09-30

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/85697

Pathology and Laboratory Medicine

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/4.0/