DOI
https://doi.org/10.25772/5D38-C421
Author ORCID Identifier
https://orcid.org/0000-0003-4975-3364
Defense Date
2024
Document Type
Dissertation
Degree Name
Doctor of Philosophy
Department
Human and Molecular Genetics
First Advisor
Senthil Radhakrishnan
Abstract
Cancer cells rely on cellular pathways that promote proliferation, inflammation, and resistance to normally programmed cell death in order to survive and form tumors. In our work we have discovered that inhibiting the expression of the gene Ddi2 (DNA damage inducible 1 homolog 2) results in a reduction in the activity of specific pro-cancer pathways and in cancer cell growth. We observed that loss of DDI2 expression resulted in dramatic decrease in tumor size in in vivo mouse xenograft models utilizing triple negative breast cancer (TNBC) cell line MDA-MB-231 and Ewing sarcoma cell lines EW16 and EW22. These DDI2-deficient cancer cells also exhibited significantly reduced proliferation, cell size, and transformative capacity in vitro. Using bulk RNA-sequencing and subsequent pathway analysis, we identified that the NF-κB pathway is strongly downregulated in DDI2-deficient TNBC and Ewing sarcoma cells. Additionally, we discovered a marked protein synthesis rate defect in DDI2-deficient cells, which has the capacity to affect the expression of a variety of cellular proteins and processes related to cancer cell growth. Our results indicate that DDI2 supports high proliferation and tumor growth by regulating protein synthesis levels and pro-tumorigenic pathways such as NF-κB in the human cancer cell. I also investigated candidate chemical inhibitors for DDI2. Herein, I describe approaches we utilized to test chemical inhibitors for DDI2 and present evidence that Nelfinavir, a recently purported DDI2 inhibitor, results in loss of NRF1 expression and therefore is unlikely to be a direct inhibitor of the protein DDI2 as has been previously reported. In a third project, we aimed to investigate the utility of proteasome inhibitors in breast cancer treatment using a systematic analysis of TCGA breast cancer datasets, which revealed that proteasome subunit transcript levels are elevated in all intrinsic subtypes (luminal, HER2-enriched, and basal-like/triple-negative) when compared to normal breast tissue. Although these observations suggest a pan-breast cancer utility for proteasome inhibitors, further experiments with breast cancer cell lines and patient-derived xenografts (PDXs) pointed to TNBC as the most sensitive subtype to proteasome inhibition. Additionally, we observed that depletion of transcription factor NRF1, as master regulator of proteasome subunit genes, significantly sensitizes TNBC xenograft mouse model tumors to proteasome inhibition. Finally, I discuss our work investigating ubiquitin ligase expression dynamics in the context of proteolysis targeting chimeras’ (PROTACs) efficacy in degrading neo-substrate targets.
Rights
© The Author
Is Part Of
VCU University Archives
Is Part Of
VCU Theses and Dissertations
Date of Submission
5-28-2024
Included in
Bioinformatics Commons, Cancer Biology Commons, Genetic Processes Commons, Medical Cell Biology Commons