Defense Date

2014

Document Type

Thesis

Degree Name

Master of Science

Department

Pharmacology & Toxicology

First Advisor

David Gewirtz

Abstract

The primary treatment options for head and neck cancer are radiation therapy or surgery, or both combined; chemotherapy is often used as an additional, or adjuvant, treatment. Patients treated with radiotherapy are exposed to a high cumulative dose of radiation over a period of time and there is a 17-33% chance of recurrence. High cumulative doses of radiation, a long time course of treatment, side effects and the possibility of recurrence provide the rationale for developing approaches for radiation sensitization, which could be helpful to patients in decreasing the dose, duration of radiation, side effects, or the chance of recurrence. Radiation induces autophagy, which is a catabolic process involving the degradation of the cell’s own components to generate energy under conditions of stress. Autophagy can be cytoprotective helping the cell to survive during stress such as nutrient deprivation or it can be cytotoxic, leading the cell toward death. We investigated whether blocking autophagy by the use of the antimalarial drug, chloroquine, could sensitize head and neck cancer cells to radiation. Studies were performed using the HN30 human head and neck cancer line (p53 wild type) derived from the pharynx as well as HN6 human cells (p53 mutant) derived from the base of the tongue. Cell viability was determined by cell counting and clonogenic survival assays, autophagy was monitored based on acridine orange staining accompanied by flow cytometry, while western blotting, DAPI and TUNEL staining and PI/annexin/FACS were utilized for determination and quantification of apoptosis. Senescence was monitored by beta-galactosidase staining/ FACS analysis. Radiation alone produced a transient growth arrest followed by proliferative recovery in both the HN30 and HN6 cancer cells. Radiation also promoted autophagy in both cell lines. The combination of chloroquine with radiation inhibited autophagy and promoted apoptotic cell death and suppression of proliferative recovery for the HN30 cells, but had little effect on sensitivity to radiation and proliferative recovery in the HN6 cells. The data suggest that autophagy induced by radiation serves a protective function in the HN30 cells and that a blockade to autophagy by chloroquine drives the cell toward apoptosis and death. In contrast, autophagy in HN6 cells appears to be non-protective as a pharmacological blockade did not sensitize the HN6 cells to radiation. These studies support the premise that autophagy induction by radiation need not necessarily have a cytoprotective function and further indicates that caution should be exercised in efforts to sensitize head and neck cancer to radiation through the clinical suppression of autophagy.

Rights

© The Author

Is Part Of

VCU University Archives

Is Part Of

VCU Theses and Dissertations

Date of Submission

May 2014

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