Parallel Investigation of Cultured Spheroids and Organoids as Models to Study Epithelial Ovarian Cancer Pathogenesis

Tomas, Emily J.

Parallel Investigation of Cultured Spheroids and Organoids as Models to Study Epithelial Ovarian Cancer Pathogenesis

Files

tomas_emily_PHD_2026_thesis (1).pdf (36.86 MB)

Date

2026-02-12

Authors

Tomas, Emily J.

Abstract

Epithelial ovarian cancer (EOC) is a devastating gynaecological disease that affects many women in the developed world. The most common subtype of high-grade serous carcinoma (HGSC) has genetic abnormalities, such as loss of function TP53 mutations and homologous recombination repair deficiency. HGSC uniquely metastasizes through multicellular spheroids, where tumour cells exfoliate into the peritoneal cavity, naturally aggregate together within the malignant ascites fluid, and subsequently reattach to surfaces to establish secondary lesions. For this reason, HGSC cells are highly adaptable during metastasis and transition between proliferative and dormant states through a process defined as cancer dormancy. Several intracellular signalling pathways, including bioenergetic stress, cell cycle control, transforming growth factor beta (TGFβ), and epithelial-mesenchymal transition (EMT), have been implicated in supporting this dormant phenotype. Elucidating these mechanisms is critical for understanding the basis of chemoresistance and recurrence in advanced HGSC. We suspected that HGSC cells possess the capacity to shift between dormant and proliferative states depending on environmental cues. However, the precise molecular signals governing this transition remain poorly understood. In this thesis, we integrated three-dimensional in vitro culture model systems, suspension spheroids and tumour-like organoids, to accurately portray the cell biology of HGSC for therapeutic and mechanistic research. First, PARP inhibitor (PARPi) sensitivity alone and in combination with the chemotherapeutic, carboplatin, varied across patient-derived HGSC cell lines, yet one line showed sequencing of PARPi before carboplatin enhanced cytotoxicity. This indicates that spheroids and organoids could be used to determine new treatment strategies for HGSC. Also, we determined that spheroids have increased bioenergetic stress and TGFβ signalling to be in a dormancy state, with Snail being a critical protein in cell survival and metastatic capacity; organoids have increased cell proliferation with elevated G2/M checkpoint regulation for survival. Next, EMT was independently regulated from TGFβ signalling, as our spheroids and organoids display a hybrid EMT phenotype expressing both epithelial and mesenchymal markers. Therefore, the results of this thesis provided new insights into cancer dormancy mechanisms involved within spheroids and organoids, and the potential of key proteins to be exploited as novel treatment options for HGSC patients.

Summary for Lay Audience

Epithelial ovarian cancer (EOC) is a deadly disease affecting women, where most are diagnosed at a late stage making treatment more difficult. Unlike other cancers, EOC spreads in an unusual way; cancer cells break off from the main ovarian tumour, float in abdominal fluid, gather together into small clusters called spheroids, and then attach to new places to form additional tumours. During this spread, cells are extremely adaptable by switching between active growth and a dormant state through a process known as “cancer dormancy”. Several internal cell signals, such as those involved in stress responses, energy use, and cell growth, are known to help cancer cells enter and maintain this dormant state. Understanding these signals is essential because dormancy is closely linked to chemotherapy resistance and cancer recurrence. We believed that EOC cells can shift back and forth between dormancy and growth depending on the environment around them. However, the exact signals controlling this switch are not well understood. In this thesis, we used advanced three-dimensional cell culture models to accurately study EOC cell biology. The “spheroid” model was used to study how these cells spread suspended in a fluid, while the “organoid” model was used to observe how cells behave in a tumour-like state. Our results showed that spheroids and organoids respond differently to treatment with chemotherapy and targeted agents, but can be used test new treatment strategies for patients. In terms of behaviour, spheroids experience increased response to stress, and rely on a protein called Snail for their survival and ability to spread. In contrast, organoids have higher levels of cell division and increased activity of proteins that control the cell cycle, such as CDK1. Across both models, we also found that the cells showed a hybrid state, meaning they adopt mixed phenotypic features that help them adapt during metastasis. Together, these findings reveal new details about how EOC cells enter dormancy and survive treatment. By uncovering these mechanisms, this work may help guide future strategies to target dormant cancer cells and improve treatment options for EOC patients.

Keywords

high-grade serous ovarian cancer, spheroids, organoids, cancer dormancy, epithelial ovarian cancer, PARP inhibitors, G2/M checkpoint, TGF-beta signaling, epithelial-mesenchymal transition