Exploring The Phenotypic Effects of Toll Signaling Pathways In Tumor Progression in EGFR-driven Glioblastoma Models Restricted; Files Only
Gonzalez Varela Saborio, Julia (Spring 2023)
Published
Abstract
The treatment options for Glioblastoma (GBM) are currently limited to resection and chemotherapy approaches, but resistance to chemotherapy is common in patients. There is an urgent need for further understanding of the underlying biology of tumorigenesis in order to develop new treatment options. Previous genomic work on GBMs has shown that the epidermal growth factor receptor (EGFR) is frequently amplified and constitutively active in GBM tumors. To discover new factors that contribute to progression of EGFR-driven GBMs, we used our Drosophila GBM model in which GBM-like glial neoplasia can be induced by glial-specific overexpression of constitutively active forms of EGFR and dp110, a subunit of Phosphoinositide 3-kinase (PI3K), using the UAS-Gal4 gene expression system. Using our Drosophila GBM model system, we performed a modifier screen in which we searched for genes and pathways that worsened or ‘enhanced’ glial neoplasia when specifically overexpressed or activated in neoplastic glia. We also performed a transcriptome analysis comparing wild-type larval brains to those expressing a repo > dEGFR λ ;dp110 CAAX genotype. From our screen and RNAseq data, we identified the Toll pathway, which in Drosophila activates an NF-ΚB signaling pathway. Furthermore, the Toll pathway normally functions in innate immune and injury responses in the brain as well as in development and cell proliferation. Using our Drosophila GBM model, we are studying how the Toll signaling pathway and its effectors, including NF- ΚB, contribute to enhanced growth of neoplastic glial cells. We will use genetic manipulation approaches such as RNAi constructs under the UAS-Gal4 system to manipulate expression of Toll components in glia, and then evaluate phenotypic changes through volumetric analysis, cell counting, confocal microscopy, and immunohistochemical staining.
Table of Contents
Table of Contents
Introduction and Background
Glioblastoma presentation in the clinic and epidemiology 1 EGFR-driven Glioblastoma biology 1 Innate immune and injury response signaling in glial cells 4Scope of the Thesis 8
Materials and Methods
Phenotypic characterization of Drosophila melanogaster 3rd instar larval brains 9 RNA sequencing 10 In vitro transformed cultures derived from a Ntv-a;Cdkn2a−/−;EGFRvIIIfl-stop-fl; 11 Ptenfl/fl mousemodel In vitro assays 12Results
RNA seq gene expression profiling in a Drosophila model of EGFR-Pi3K driven 14 glioblastoma shows enrichment of innate immune pathways Toll pathway activation leads to an enhanced dEGFR;dp110CAAX phenotype 2010. Ntv-a;Cdkn2a−/−;EGFRvIII fl-stop-fl;Ptenfl/fl mammalian model of EGFR-Pi3K driven 25 glioblastoma
Transformed murine neural stem cells expressing EGFRvIII and PTEN knockout 28 show TLR pathway activation in vitro EGFR Tyrosine Kinase Inhibitor Erlotinib is associated with canonical NF-κB 32 pathway components in Ntv-a;Cdkn2a−/−;EGFRvIII;Pten-/- murine neural stem cellsDiscussion and Future Directions
13. Conclusions 36 14. Future directions in our Drosophila repo>dEGFR;dp110CAAX model 37 15. Future directions in our Ntv-a;Cdkn2a−/−;EGFRvIII;Pten-/- murine model 38 16. NF-κB in glioma 40 17. Closing remarks 40
References 42 Appendices 46
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File download under embargo until 22 May 2024 | 2023-04-28 12:55:04 -0400 | File download under embargo until 22 May 2024 |
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