UC Irvine

ABSTRACT OF THE DISSERTATION

Elucidating the mechanism through which loss-of-function in RB1 mediates poor prognosis in osteosarcoma

By

Stephanie Cherie Wu

Doctor of Philosophy in Biological Sciences

University of California, Irvine, 2019

Assistant Professor Claudia A. Benavente, Chair

Osteosarcoma is the most common primary bone malignancy in children and adolescents. Among the various molecular mechanisms implicated in osteosarcomagenesis reviewed in Chapter 1, the RB/E2F pathway is of particular importance as virtually all cases of osteosarcoma display alterations in the this signaling pathway. Loss of function mutations at the retinoblastoma (RB1) gene are associated with increased mortality, metastasis and poor therapeutic outlook in osteosarcoma. However, the mechanism(s) through which loss of RB worsens clinical outcome remain to be elucidated. We examined the transcription factor E2F family members that are associated with increased malignancy in RB-null osteosarcoma tumors and found that loss of activator E2Fs, E2F1 and E2F3, significantly delays tumor progression and increases the overall survival of the Tp53/Rb1-deficient osteosarcoma mouse model (Chapter 2). We then studied the role of two critical downstream effectors of the RB/E2F pathway, HELLS and UHRF1. While both are upregulated and overexpressed in osteosarcoma, we observed that HELLS has limited effect on tumor proliferation and migration. Loss of Hells in osteosarcoma has no effect in tumor initiation and overall survival of osteosarcoma mice. This suggests that while HELLS may serve as a biomarker for tumorigenesis and for RB/E2F pathway status, it is unlikely to serve as a relevant target for therapeutics in osteosarcoma (Chapter 3). On the contrary, we showed that UHRF1 upregulation is critical in rendering osteosarcoma cells more aggressive. Using gain/loss-of-function assays to study the role of UHRF1 in osteosarcoma, we observed that UHRF1 is involved in promoting cell proliferation, migration, and invasion. Loss of Uhrf1 in genetically engineered osteosarcoma model lengthens the life span of mice bearing osteosarcoma and decreases pulmonary metastases. These are described in chapter 4, where we discuss how UHRF1 mediates its tumor-promoting functions in osteosarcoma, and provide evidence supporting UHRF1 targeting as a novel therapeutic option. Taken together, this study illuminates new mechanistic insights of RB-mediated poor prognosis that could improve current therapeutic strategies of osteosarcoma, as well as other malignancies harboring RB/E2F pathway inactivation. Following our discovery, chapter 5 documents our proposed methods in attempting to provide a more comprehensive mechanistic understanding of the RB-UHRF1 network.