Date of Award

2014

Document Type

Thesis

Degree Name

Bachelors

Department

Natural Sciences

First Advisor

Clore, Amy

Keywords

Tumor Necrosis Factor-Related Inducing Ligand, Chemotherapy, Lung Cancer, Ligands

Area of Concentration

Biology

Abstract

Mucin 1 (MUC1) is a glycoprotein expressed on the apical surface of epithelial cells in organs such as the lung. Typically upregulated during cases of acute inflammation, an overexpression of MUC1 has been observed in cases of acquired resistance to Tumor Necrosis Factor-Related Apoptosis Inducing Ligand (TRAIL) in non-small cell lung cancer. The ability of cancer cells to gain resistance to chemotherapy treatments has been attributed to their ability to evade apoptosis initiated by ligands like TRAIL through the dysregulation of otherwise harmless proteins, such as MUC1. A decrease in catalase expression, coupled by an increase in the presence of reactive oxygen species in TRAIL-resistant cells, partially accounts for the overexpression of MUC1. However, the pathway leading to MUC1 overexpression has not been fully elucidated, and the proteins with which MUC1 associates in order to disrupt TRAIL signaling within the cell are not all known. Two TRAIL-resistant non-small cell lung cancer cell lines, A549 and H460, were established to study MUC1’s role in acquired chemo-resistance. The levels at which MUC1 and catalase are regulated were examined and possible mechanisms responsible for this regulation explored. MUC1 and catalase were both regulated at either the transcriptional or post-transcriptional level, suggesting a potential role for miRNA regulation of these genes. Three candidates potentially responsible for this regulation, miRNA 943, miRNA 146a, and miRNA 551b, were tested for their expression in non-resistant and TRAIL-resistant cells. Only miRNA 551b was found to be upregulated in A549 TRAIL-resistant cells, but not in H460 TRAIL-resistant cells. To further investigate its role in catalase regulation, levels of miRNA 551b were ‘knocked down’ (reduced), and MUC1 and catalase levels observed. Transient knockdown of miRNA 551b in A549 TRAIL-resistant cells did not conclusively lead to recovery of catalase levels, and MUC1 remained overexpressed. It was concluded that while MUC1 and catalase are regulated prior to translation, miRNA 551b may not be responsible for this regulation. MUC1 transient knockdown was used to observe the effect on various pro-survival proteins, including phosphorylated epithelial growth factor receptor (pEGFR), unphosphorylated EGFR, phosphorylated protein kinase B (pAkt), cellular FADD-Like Interleukin-1β-Converting Enzyme Inhibitory Protein (c-FLIP), cyclooxygenase-2 (COX-2), and Myeloid cell leukemia sequence 1 (Mcl-1). MUC1 knockdown in A549 TRAIL-resistant cells potentially attenuated expression of MUC1 and c-FLIP, but not pEGFR, EGFR, pAkt, COX-2 or Mcl-1. Based on these results, MUC1 appears to potentially be involved in the regulation of c-FLIP, a well-established pro-survival protein involved in the termination of TRAIL signaling, suggesting a possible pathway for inhibition of apoptosis in chemo-resistant cells.

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