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Thesis
Modeling the cooperative effects of GNAS^[R201C], KRAS^[G12V], and TP53^[R175H] in human pancreatic ductal epithelial cells
Master of Science (M.S.), Drexel University
Jul 2025
DOI:
https://doi.org/10.17918/00011125
Abstract
Pancreatic ductal adenocarcinoma (PDAC) is a deadly disease that can arise from clinically detectable precursor lesions known as intraductal papillary mucinous neoplasms (IPMNs). However, the precise molecular events that drive IPMN progression towards malignant transformation are unclear. Genomic analysis indicates that oncogenic mutations in KRAS and GNAS are found at high frequencies within both early-stage IPMNs and IPMN-derived PDAC. Previous studies using human stem-cell derived pancreatic organoid models demonstrate that expression of oncogenic GNAS, either alone or in combination with oncogenic KRAS, is not sufficient to form IPMN-like lesions in the mouse pancreas, suggesting additional mutations such as inactivation of tumor suppressor genes may be necessary. Mutations in TP53 are observed in 38.1% of high-grade IPMNs and are considered a hallmark of IPMN-derived PDAC. In this thesis, we generated combinatorial plasmids to investigate the effect of GNAS^[R201C], KRAS^[G12V], and TP53^[R175H] expression in human pancreatic ductal epithelial (HPDE) cells. Using western blot analysis and fluorescence imaging, we found that expression levels of TP53 and KRAS are heterogenous in both GNAS^[R201C]/TP53^[R175H] and GNAS^[R201C]/TP53^[R175H]/KRAS^[G12V]-expressing HPDE cells. Our findings also showed that the expression of GNAS^[R201C]/TP53^[R175H]/KRAS^[G12V] activates MAPK signaling, whereas expression of GNAS^[R201C]/TP53^[R175H] activates GNAS-associated PKA signaling. Interestingly, 3D growth assays showed that expression of GNAS^[R201C]/TP53^[R175H]/KRAS^[G12V] leads to a significant increase in total surface area of HPDE spheroids that exhibit elevated expression of oncogenic KRAS. Our findings, therefore, provide a rationale to further investigate these combinatorial mutations in human stem-cell derived pancreatic organoid models and assess their ability to develop IPMN lesions in vivo.
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Details
- Title
- Modeling the cooperative effects of GNAS^[R201C], KRAS^[G12V], and TP53^[R175H] in human pancreatic ductal epithelial cells
- Creators
- Erik Michael Schubert
- Contributors
- Ridhdhi Desai (Advisor)
- Awarding Institution
- Drexel University
- Degree Awarded
- Master of Science (M.S.)
- Publisher
- Drexel University; Philadelphia, Pennsylvania
- Number of pages
- x, 62 pages
- Resource Type
- Thesis
- Language
- English
- Academic Unit
- Biochemistry and Molecular Biology; College of Medicine; Drexel University
- Other Identifier
- 991022074528404721