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Bakteria, Sam

To this day, cancer has remained one of the most prominent causes of death. Despite there being great treatments such as radiotherapy, chemotherapy, and surgery, almost one out of every six deaths was due to cancer, as reported in 2020. This shortcoming is because we have not fully understood the cell signaling pathways involved in producing the hallmarks of cancer. A big hurdle the academic community has faced recently is the reproducibility of published results. In recent studies, it has been suggested that only 11% to 50% of the time were scientists able to reproduce findings from other academic laboratories. This can be due to using poorly characterized tools, such as antibodies with unknown specificities for Western blotting and other immunochemical analyses. For this reason, Kinexus Bioinformatics Corporation believes in developing its own tools and databases for its continued research on cancer cell signaling pathways. This thesis aims to address gaps in knowledge regarding cancer signaling pathways, such as those involving mammalian targets of Rapamycin (mTOR) and extracellular signal-regulated kinases (ERKs). Reports indicated that mutations in mTOR and ERK signaling pathways often lead to uncontrolled cell growth, which is why they are the focus of this thesis. Using Kinexus' new KAM-2000 antibody microarray slides, we identified new potential signaling pathways activated in Henrietta Lacks (HeLa) cells in response to stress inducing chemicals such anisomycin and arsenate. Once critical protein substrates were identified in these signaling pathways, novel antibodies were developed for selected target proteins at crucial phosphorylation sites. These antibodies were then characterized for their specificity using SPOT peptide membranes. Western blots were developed using HeLa cells exposed to mitogenic and stress stimuli to confirm treatment effects on target proteins. This methodology identified cell signaling pathways that lead to uncontrolled cell proliferation without activating mTOR-related pathways. Ribosomal protein S6 (RPS6) was used as our marker for detecting cell proliferation. The experiments identified RPS6 activation through epidermal growth factor receptors, protein kinase A (PKA), protein kinase C, and insulin receptor pathways. Furthermore, novel peptide inhibitors were developed against PKA, and with limited success, they could inhibit PKA activation, resulting in inhibited cell proliferation. The identified signaling pathways have proven to be excellent targets for cancer therapeutics and further expanded our knowledge regarding involved signaling pathways.

Text

University of British Columbia. PHAR 491

Vancouver : University of British Columbia Library

10.14288/1.0445153

Pharmaceutical Sciences, Faculty of

Undergraduate

DSpace