Sir Peter MacCallum Department of Oncology - Theses
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ItemIdentifying heterogenous mechanisms of drug resistance in high grade serous ovarian cancer( 2023-05)Ovarian, fallopian tube and primary peritoneal cancer is the sixth most common cause of cancer death for women in Australia. A diagnosis of ovarian cancer is life-altering with a five-year survival of only 45.7%. While clinical factors such as disease stage are important for prognosis and response to therapy, the underlying genomic, transcriptomic and immune characteristics of the tumour are also key determinants of outcome. High grade serous ovarian cancer (HGSC) is the most common epithelial ovarian cancer subtype, and is characterised by frequent germline, somatic and epigenetic aberrations which result in homologous recombination DNA repair (HR) deficiency. Aneuploidy and frequent somatic copy number aberrations (SCNA) are common features of the genomic landscape of HGSC. Despite often initially responding to therapy, acquired resistance is extremely common, arising through a variety of genomic, transcriptomic, adaptive and microenvironmental mechanisms. Owing to the scarcity of biopsies or surgery in relapsed disease, the frequency and distribution of these acquired resistance mechanisms is only partially known. In particular, there is a paucity of information about resistance mechanisms in late-stage disease following the multiple lines of therapy that a typical patient with HGSC receives. This thesis addressed these important gaps in knowledge by examining resistance mechanisms in a cohort of women with HR deficient HGSC who underwent a research autopsy, providing unique insight into the architecture of end stage disease. Importantly, resistance mechanisms were frequently subclonal, often with more than one mechanism detected within a single metastatic site. Evidence of convergent evolution was also observed, including of reversion mutations and BRCA1-specific mechanisms of HR restoration. An increased frequency of whole genome duplication, an important event in cancer evolution, was noted in the end-stage samples compared to what is known in primary tumours. Following this observation, the transcriptomic differences between tumours with and without WGD were assessed. Examination of the transcriptomic differences in whole genome duplication revealed downregulation of CIITA and other MHC-II genes, highlighting a potential means of immune escape and a potential therapeutic vulnerability. This thesis also examined mechanisms of resistance specifically occurring following receipt of poly-ADP ribose polymerase (PARP) inhibitor therapy, and the natural history and evolution of reversion mutations in HR deficient HGSC, showing that they are in fact not frequently present outside of a progressing or resistant cohort. The findings of this thesis add to the current knowledge of resistance to therapy, demonstrate the need for accessible genomic testing in the clinical management of HGSC and indicate that further exploration of resistance should focus on transcriptomics, epigenetics, and the tumour microenvironment.
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ItemReducing the morbidity and mortality of ovarian cancer through prevention( 2022)Ovarian cancer (OC) remains a major cause of morbidity and mortality in women. With the current absence of effective screening and early detection methods, prevention of OC through risk-reducing salpingo-oophorectomy remains the most effective means of reducing OC morbidity and mortality. However, a significant challenge lies in identifying the women at greatest risk of OC and enabling their access to risk reducing interventions before cancer occurs. This study aimed to explore pathways to facilitate cancer prevention in at risk individuals that are not currently available in the clinical setting. In Chapter 3 of this thesis, I describe the TRACEBACK study - a novel approach to OC prevention through finding previously unrecognised carriers of pathogenic germline variants (PVs) among women with high grade non-mucinous ovarian cancer (HGNMOC) who had missed the opportunity for genetic testing. I piloted three methods of ascertainment and evaluated sequenced DNA of tissue samples from over 1000 women with HGNMOC, including some under a waiver of consent. The findings were returned to the women (probands) if they were alive, or if deceased the results were returned to a family member. PVs were detected in 119 probands and to date, 56 results have been returned. Although logistically and ethically complex, I have demonstrated the feasibility of this approach to find previously unknown carriers of PVs. The success of TRACEBACK as a cancer prevention initiative hinges on the subsequent uptake of cascade genetic testing and use of risk reducing strategies by family members. Therefore, the acceptability and value of TRACEBACK to probands and their family members was explored in Chapter 4 of this thesis, through qualitative interviews performed as part of the TRACEBACK.Evaluation study. In this pilot study, eight interviews were performed. While participants were shocked by the TRACEBACK contact irrespective of their method of ascertainment or consent process, all were unanimously positive in their perceptions of the value of the project and were appreciative and grateful that a project of this type was being conducted. Chapter 5 of this thesis explores the utility of polygenic risk scores (PRS) in characterising OC risk and response to treatment. I demonstrated that a one unit increment in PRS increases the risk of high grade serous ovarian cancer (HGSOC) by 38% (OR 1.38, p value <0.001). This study gives further validation to the role that PRS is likely to have in OC clinical risk prediction models in the future. Interestingly, this study revealed that a high PRS may be negatively impacting OC treatment response. My research has demonstrated that identification of at-risk individuals through the TRACEBACK study and examination of PRS are likely to facilitate OC prevention, and provide important steps toward reducing the morbidity and mortality of OC. The genetic testing of samples from deceased patients provides a novel model for risk reduction in cancers where there is a substantial frequency of pathogenic germline variants, including triple negative breast cancer.
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ItemCyclin E1 as a therapeutic target in high grade serous ovarian cancer( 2017)The central theme of this thesis is developing therapeutic strategies to selectively target CCNE1 amplified high grade serous ovarian cancer (HGSC). Patients with CCNE1 amplified HGSC represent a key unmet clinical need given that they are associated with primary treatment resistance and poor clinical outcome. Novel therapeutic strategies are urgently required in order to provide these patients with additional treatment options. Using short interfering RNA and short hairpin RNA, I demonstrated selective sensitivity of CCNE1 amplified HGSC to CDK2 gene suppression. However, I did not demonstrate similar amplicon dependent sensitivity to dinaciclib, a potent small molecule inhibitor of multiple CDKs. In order to identify drug combinations that would synergise with dinaciclib, I performed a high throughput compound screen in CCNE1 amplified HGSC cell lines. I identified a combination of dinaciclib and MK-2206, an AKT inhibitor, that was selectively synergistic in in vitro and in vivo models of CCNE1 amplified HGSC. CCNE1 and AKT2 were noted to be co-amplified in primary HGSC samples, and a number of genes in the AKT pathway were found to be required in CCNE1 amplified HGSC cell lines. Furthermore, over-expression of cyclin E1 and AKT isoforms resulted in uncontrolled growth characteristics in TP53-mutant fallopian tube secretory cells, the proposed cell of origin for HGSC. Taken together, these findings suggest that co-operative interaction between CCNE1 and the AKT pathway in HGSC may be exploited therapeutically. I also explored the potential mechanisms of resistance to CDK inhibitors by generating cell lines resistant to dinaciclib. Dinaciclib in combination with multiple BH3-mimetic compounds was noted to be synergistic in CDK-inhibitor resistant cell lines. Upregulation of multiple anti-apoptotic genes was observed in resistant cell lines compared to parental sensitive cell lines, suggesting that this is a potential mechanism of resistance to CDK inhibitors. Targeting homologous recombination (HR) may also be a therapeutic option in CCNE1 amplified HGSC. Proteasome inhibitors such as bortezomib have been shown to be indirect inhibitors of HR, and I showed that CCNE1 amplified cell lines were highly sensitive to bortezomib and MLN9708, a second generation proteasome inhibitor. Potential synergistic combinations with bortezomib were identified in a high throughput compound screen, including a number of HDAC inhibitors, suggesting a possible class effect.