Date on Master's Thesis/Doctoral Dissertation
Pharmacology and Toxicology
Pharmacology and Toxicology, PhD
Committee Co-Chair (if applicable)
EGFR; cell death; apoptosis; protein kinase G; STAT3
The Epidermal Growth Factor Receptor (EGFR) is a 170-kilodalton transmembrane protein that belongs to the ErbB family of receptor tyrosine kinases. Upon ligand-mediated activation, the EGFR is responsible for cell growth, proliferation, and tissue homeostasis; however, the EGFR is overexpressed in many human malignancies, including MDA-MB-468 cells, a metastatic breast epithelial cell line. Studies within this cell line, and other cell lines characterized with high EGFR levels, have shown that EGF stimulation results in the induction of apoptosis. However, the mechanisms and signaling effectors implicated in this process have yet to be elucidated. The overarching research goal of this dissertation was to better understand the molecular mechanisms employed by the receptor that result in the induction of apoptosis in MDA-MB-468 cells. Identifying key regulators/mediators of EGF-induced apoptosis will help elucidate new pharmacological targets that better attenuate cell growth in cancers characterized by EGFR hyperexpression. Through an extensive review of the literature, we have identified Cyclic GMP dependent protein kinase (PKG) and Signal Transducer and Activator of Transcription 3 (STAT3) as potential signaling intermediates of EGF-induced apoptosis. EGF stimulation resulted in a dose-dependent increase in PKG activity. Agonists of PKG did not induce EGFR activation, confirming that PKG activity occurred downstream of receptor activation. RNA Interference (RNAi) targeting PKG resulted in a significant reduction in EGF-induced cell death, as well as a reduction in apoptosis. Similarly, we used siRNA to ablate STAT3 activity, which resulted in a significant attenuation of apoptosis, as measured by PARP cleavage. Additionally, cytokine-mediated (EGFR-independent) activation of STAT3 induced PKG activation, and resulted in a significant induction of apoptosis that was sustained over time. Together, our results indicate that PKG and STAT3 are molecular targets for cancers with high levels of EGFR, and that activation of proteins that induce cell death may be more beneficial, therapeutically, than antagonizing proteins that promote cell growth.
Jackson, Nicole Marion, "Identifying the signaling mechanisms of EGFR-mediated apoptosis." (2017). Electronic Theses and Dissertations. Paper 2715.