Date of Graduation

12-2014

Document Type

Thesis (MS)

Program Affiliation

Biomedical Sciences

Degree Name

Masters of Science (MS)

Advisor/Committee Chair

Anil K Sood, M.D.

Committee Member

Lois Ramondetta, M.D.

Committee Member

Gary Gallick, Ph.D.

Committee Member

Menashe Bar-Eli, Ph.D

Committee Member

Robert Dantzer, Ph.D.

Abstract

Background: Chronic stress and sustained adrenergic signaling are known to promote tumor progression. The underlying mechanisms behind this process are not well understood. We examined the effects of sustained adrenergic signaling on cervical cancer progression through increased expression of HPV oncogenes, E6 and E7.

Materials and Methods: ADRβ expression levels were examined in patient-derived cervical cancer samples. We used an orthotopic model of cervical cancer to investigate the effects of restraint stress on tumor growth and metastasis. We evaluated the in vivo effects of a β-blocker, propranolol, and HPV E6/E7 siRNA. In vitro, ADRβ positive cervical cancer cells were treated with norepinephrine (NE) or isoproterenol (ISO) to examine intracellular responses. Invasion and anoikis assays were performed to elucidate the biological effect of NE. Furthermore, the significance of secreted HPV was examined.

Results: Among tumor samples evaluated from cervical cancer patients, 61% had increased ADRβ2 expression which correlates with decreased overall survival (p=0.038). In an orthotopic model of cervical cancer, chronic stress led to increased tumor weight and nodules. This effect was abrogated with propranolol. Further, treatment with HPV E6/E7 siRNA in the restraint stress model decreased tumor growth and metastases. In vitro, HPV E6/E7 mRNA was elevated in cell lysates and corresponding supernatant after treatment with NE or ISO. NE exposure resulted in increased invasion and migration of SiHa cells, while E6/E7 siRNA abrogated these effects. After NE exposure, ME-180 cells showed a 45% reduction in anoikis compared to controls. Fibroblasts cultured with supernatant from SiHa cells had increased migration and elevated mRNA of pro-inflammatory genes CXCL2 and IL-8. Further, the fibroblasts took up cervical cancer cell derived exosomes that contain HPV E7. The conditioned fibroblasts demonstrated the ability to increase cervical cancer cell invasion when co-cultured.

Conclusion: This study shows increased adrenergic signaling promotes cervical cancer growth and progression. Disruption of this pathway with β-blockers could provide a novel complement to current therapies. In addition, we show HPV oncogenes E6 and E7 can be influenced by catecholamines. We also show exosomes as a potential mean of communication between cervical cancer cells and fibroblasts in vitro. The functional implications of this study needs to be explored further.