Molecular and functional analysis of Mucinous Ovarian Carcinoma

Abstract

Mucinous ovarian carcinoma (MOC) is a rare subtype of ovarian cancer (OC) distinct from all other subtypes. MOC represents 3% of OC cases; however, it responds very poorly to conventional chemotherapies contributing to the poor outcome of OC. Moreover, there is a lack of effective therapies for MOC since treatments with platinum-based chemotherapy or PARP-inhibitors are unlikely to be effective. The recent description of the molecular landscape of MOC, including contributions by work in this thesis, allowed the identification of novel therapies that target the most recurrent genetic features of the disease. Nevertheless, there is still the need to develop accurate pre-clinical models in order to test the efficacy of these new therapies.

This thesis evaluates the transcription factor ELF3 as a new MOC candidate driver gene. We performed a comprehensive analysis of 210 human mucinous ovarian tumours and identified that ELF3 was mutated in 8.2% of our sample cohort. To assess whether ELF3 acts as a tumour suppressor or oncogene in MOC and how its mutated function is expressed in cancer, we evaluated the ability of MOC cells to proliferate, migrate and invade, by introducing truncating mutations using CRISPR-Cas9 technology. It was found that ELF3 could possibly act as a tumour suppressor in MOC; however, its role might depend on coexisting mutations in the cells.

This thesis also describes a novel 3D patient-derived therapeutic approach to pre-test the effectiveness of drugs, as monotherapy or in combination, before exposing patients to ineffective therapies or side effects. A mucinous ovarian tumour 3D platform that enables the correlation of patient genetic data with drug testing was established in the first result chapter of this thesis. MOC organoid development was based on the MOC genetic profile and characteristics as well as optimization that adapted human colorectal and pancreatic organoid culture systems. It was found that Wnt-3a is essential for mucinous ovarian tumour organoid long-term derivation. Moreover, MOC organoids faithfully recapitulate MOC primary tumours at the genomic level and retain histological characteristics of their corresponding primary tumours.

The development of MOC organoids and 3D spheroids from cell lines together with the description of the genetic profile of MOC allowed the development and testing of a panel of novel therapies that targets the most frequent mutations found in MOC tumours, including a mutant TP53 activator, a Wnt inhibitor, RAF/EGFR/RAS/CDK4 and CDK6 inhibitors, and two antitumour compounds for HER2-overexpressing tumour cells (APR-246, OMP-18R5, BGB-283, BKM120, PD-0332991, PLX4032, Trastuzumab and Lapatinib). The organoids and spheroids showed different responses to the individual therapies tested, depending on the individual genetic features of the models. Spheroids showed sensitivity to PD-0332991, Trastuzumab, PLX4032 , and BKM-120 . MOC organoids also showed different responses: MOC organoid MOCOR-16 showed sensitivity to APR-246, and resistance to Trastuzumab; however, MOCOR-38 was sensitive to Trastuzumab but resistant to APR-246. This chapter also shows the first attempt of drug synergy scoring using automatic robots to create 3D MOC models. Synergistic combinations were identified for one of the spheroid lines when APR-246 and BKM-120 were combined. Interestingly, the spheroids were resistant to APR-246 in monotherapy. If validated independently, these results could potentially represent a more effective treatment for MOC. Antagonistic interactions were also found, with the strongest antagonism occurring between PLX4032 and BGB-283.

The findings of this study substantially increase our understanding of mucinous ovarian tumours together with potential personalized therapeutic targets to improve the survival outcomes for women with this deadly disease.