Date of Graduation
12-2016
Document Type
Dissertation (PhD)
Program Affiliation
Cancer Biology
Degree Name
Doctor of Philosophy (PhD)
Advisor/Committee Chair
Eugenie S. Kleinerman, MD
Committee Member
Nino Rainusso, MD
Committee Member
Candelaria Gomez-Manzano, MD
Committee Member
Dianna Milewicz, MD Ph.D.
Committee Member
Gary Gallick, Ph.D.
Abstract
Metastatic disease to the lungs is the primary cause of death for patients with pediatric osteosarcoma (OS). OS has a high degree of heterogeneity and genomic instability, making understanding the pathogenesis and drivers of metastasis of this disease challenging. In an effort to explain tumoral heterogeneity, the tumor initiating cell model (TIC) states that tumors are composed of cells that form the majority of the tumor and are terminally differentiated. This model however, attributes tumorigenesis, metastasis and chemoresistance to a distinct cell population with a stem-like phenotype that can be identified using selective markers. OS appears to follow this model where OS cells with tumor initiating potential can be identified by expression of Stro1, CD117 and embryonic stem cell transcription factors such as Sox2, Nanog and Oct3/4. Additionally, OS stem-like cells display high aldehyde dehydrogenase activity and sarcosphere formation under limited nutrient media and anchorage independence. These markers are not feasible targets for therapy due to their expression on normal tissue stem cells; however, upstream regulators of this phenotype may be targetable. Therefore, we investigated other modulators of the stem-like phenotype.
Her4, a transmembrane receptor of the EGFR family, has been recently studied for its role in cancer. Previously, we demonstrated that Her4 is highly expressed in neuroblastoma, and OS, while others have shown its importance in Ewing sarcoma. This receptor is induced and required to survive stressors, like anchorage independence, serum starvation and chemotherapy treatment, which are similar in vitro conditions used to enrich for cells with tumor initiating potential. Therefore, we hypothesized that Her4 expression is an important regulator of a stem-like phenotype in OS.
In sarcosphere culture, Her4 expression is induced and precedes the induction of CD117 and Stro1. OS cells with Her4 deleted by CRISPR/Cas9 have decreased aldehyde dehydrogenase activity and cannot upregulate the pluripotency transcription factors Sox2, Oct3/4 and Nanog even when in sarcospheres. Overexpression of exogenous Her4 was able to cause upregulation of these transcription factors and increase expression of CD117 in monolayer culture. We examined Her4 expression in OS diagnostic biopsies and determined the correlation with metastasis free survival. Tumors with high Her4 expression have higher probability of developing metastatic disease.
In this dissertation, we demonstrate that Her4 expression is induced by conditions that enrich stem-like cells and its expression correlates with the ability to upregulate various OS TIC markers. Therefore, Her4 may contribute to pathogenesis of OS by conferring a stem-like phenotype.
Keywords
osteosarcoma, stem-like phenotype, Her4, ERBB4