Surgery (RMH) - Theses

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    Hormonally induced defects of DNA damage repair genes: an oncogenic driver of prostate cancer
    McCoy, Patrick ( 2018)
    Patients with defects in the mismatch repair pathway, driven by either MSH2 or MSH6 loss, experience a significant increase in the incidence of prostate cancer, while germline mismatch repair defects in either MLH1 or PMS2, exhibit no such increase. This PhD project demonstrates that androgen-receptor activation, a known driver of prostate cancer, can disrupt the MSH2 gene in prostate cancer model systems through the induction of structural variations. Prostate tumours from two contrasting risk cohorts were screened to confirm loss of MSH2 protein expression in a small number of patients. Surprisingly, it was also found that a small but significant fraction of high-risk cases exhibited reduced expression of MSH2 without complete loss. Stratifying a large independent TCGA prostate cancer cohort for MSH2 expression levels revealed that patients whose tumours exhibited either complete loss or aberrant levels of MSH2 had significantly worse survival outcomes and accelerated clinical progression. In contrast, aberrant MSH2 levels had no impact on clinical survival in colorectal cancer. This PhD project also demonstrates that reduced expression of MSH2 can be explained by androgen-induced microRNA regulatory mechanisms. Here, it is demonstrated that miR-21 and miR-141 may both target the MSH2 gene leading to reduced MSH2 protein staining and both microRNAs were seen to be upregulated in prostate cancer patients with reduced MSH2 levels. Interestingly, this thesis also shows that miR-21 and miR-141 are both regulated by androgens, implicating this mechanism as a second androgen driven method of MSH2 downregulation. Importantly this PhD project also found that aberrant MSH2 expression in prostate tumours does not induce the same enhanced immune cell mobilisation seen in colorectal tumours suggesting that the prostate is an immune privileged site. This is contrary to the findings of other studies and may warrant a re-evaluation of whether MSH2 deficient prostate cancers are likely to benefit from immunotherapies. To further investigate if the prostate tumour microenvironment is indeed in an immunosuppressive state, a detailed investigation of the transcriptomic profile of the cells of the tumour microenvironment was also conducted. This resulted in not only the discovery of immunosuppressive signatures in tumour infiltrating T-cells but also significant transcriptomic alterations in other cancer associated pathways such as osteogenesis, cell migration, epithelial mesenchymal transition (EMT), hormone signalling and angiogenesis throughout the tumour microenvironment compared to the cellular make-up of benign prostatic tissue. The data presented in this thesis constitute a significant contribution to the current understanding of how defects in the mismatch repair gene MSH2 may affect prostate cancer severity. Additionally, these studies demonstrate multiple mechanisms through which the prostate tumour microenvironment may enhance prostate cancer progression.
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    Investigating the evolution of structural variation in cancer
    Cmero, Marek ( 2017)
    Cancers arise from single progenitor cells that acquire mutations, eventually dividing into mixed populations with distinct genotypes. These populations can be estimated by identifying common mutational profiles, using computational techniques applied to sequencing data from tumour tissue samples. Existing methods have largely focused on single nucleotide variants (SNVs), despite growing evidence of the importance of structural variation (SV) as drivers in certain subtypes of cancer. While some approaches use copy-number aberrant SVs, no method has incorporated balanced rearrangements. To address this, I developed a Bayesian inference approach for estimating SV cancer cell fraction called SVclone. I validated SVclone using in silico mixtures of real samples in known proportions and found that clonal deconvolution using SV breakpoints can yield comparable results to SNV-based clustering. I then applied the method to 2,778 whole-genomes across 39 distinct tumour types, uncovering a subclonal copy-number neutral rearrangement phenotype with decreased overall survival. This clinically relevant finding could not have been found using existing methods. To further expand the methodology, and demonstrate its application to low data quality contexts, I developed a novel statistical approach to test for clonal differences in high-variance, formalin-fixed, paraffin-embedded (FFPE) samples. Together with variant curation strategies to minimise FFPE artefact, I applied the approach to longitudinal samples from a cohort of neo-adjuvant treated prostate cancer patients to investigate whether clonal differences can be inferred in highly noisy data. This thesis demonstrates that characterising the evolution of structural variation, particularly balanced rearrangements, results in clinically relevant insights. Identifying the patterns and dynamics of structural variation in the context of tumour evolution will ultimately help improve understanding of common pathways of tumour progression. Through this knowledge, cancers driven by SVs will have clearer prognoses and clinical treatment decisions will ultimately be improved, leading to better patient outcomes.
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    Integrated genomics in prostate cancer: defining the lethal phenotype
    Hong, Matthew Kok-Hao ( 2014)
    Prostate cancer is the second leading cause of cancer death in Australian men. However, it has a wide spectrum of biological aggressiveness from those that lead indolent courses to cases with rapidly lethal propensity. The inability to confidently identify cases with lethal disposition at an early clinical stage has led to widespread overtreatment and an urgent need for more discriminating biomarkers. Recent advances in genomics techniques have presented powerful new tools with which to study cancer. Simultaneous analysis can be undertaken of millions of genetic markers and even entire cancer cell genomes and this has transformed the landscape for molecular discovery. Methods in computational biology have similarly evolved to meet the challenges of deriving meaningful conclusions from the huge quantities of data produced in genomics experiments. This thesis aimed to combine genomics and the question of lethality in prostate cancer. Prostate cancer lethality is intertwined with the development of metastasis, which usually heralds the onset of a trajectory towards prostate cancer-specific death, and it is around this central theme that the majority of the work is based. As the acquisition of clinical tissue samples for prostate cancer research has previously been based on radical prostatectomy programs for primary cancer samples and rapid autopsy studies for metastases, a significant gap exists along the spectrum from localised disease to end-stage treatment-resistant metastatic disease. I therefore developed a program to obtain metastatic prostate cancer tissue from live patients, and demonstrated the feasibility of percutaneous image-guided biopsy of osseous prostate cancer metastasis in a way that yields nucleic acids suitable for genomics applications. The remaining work follows lethality along the clinical spectrum of disease from distant metastatic disease to locoregional lymph node metastases to localised disease, and finally to the biopsy setting. Samples of primary and metastatic prostate cancer from two patients were interrogated through genomic sequencing, with observation of the subclonal origin of metastases and the dynamic shift in subclonal populations within a metastasis in response to therapy. The relationships between lymph node metastases to different primary cancer foci within the prostate gland were then explored in three patients with high-risk prostate cancer. The final two chapters demonstrate an approach towards clinically relevant biomarker discovery by using integrated genomics to characterise molecular differences between high-risk prostate cancer and indolent disease based on both primary cancer tissue and surrounding benign tissue. Biomarkers discovered based on patient cohorts selected on clinical outcome may be exploited for therapy or clinically relevant prognostic tools. For genomics research in prostate cancer, one requires precision in clinical informatics and patient selection, careful tissue preparation and integrated use of genomic platforms. In this thesis I show how this might be achieved in different patient groups representing different aspects of the disease spectrum, and how this can be used to unravel the molecular underpinnings of lethality in prostate cancer.
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    Targeting steroid signalling pathways in hormone dependent cancer
    Fankhauser, Matthew James ( 2011)
    A sex hormone binding decoy protein made by fusing the ligand binding domain of the androgen receptor to an immunoglobulin Fc region or an elastin like polypeptide (ELP) was designed to bind circulating testosterone and other androgens, due to the high affinity of the androgen receptor ligand binding domain for the male sex hormones. Multiple cell based in vitro assays were designed and monitored for both growth and the biomarker, prostate specific antigen (PSA), with data confirming the decoy protein was able to inhibit the growth of hormone dependent prostate cancer cell lines and the androgen dependent expression of PSA. In vivo models were able to show the binding decoy protein was able to reduce free testosterone levels within a mouse, with further in vivo subcutaneous and orthotopic xenograft models of various prostate cancer stages demonstrated the ability of the binding decoy protein to reduce the growth and PSA expression of prostate tumours. Prostate cancer relies on circulating male hormones, in particular testosterone, to drive growth and proliferation of prostate tumour cells. This reliance on testosterone led to the development of novel therapeutics targeted at decreasing available testosterone, collectively termed Androgen Deprivation Therapy (ADT) and upon treatment resulted in decreased tumour growth. ADT is routinely used for treatment of prostate cancer; however the failure of current therapies to completely remove circulating testosterone and other androgens has been linked with the continued hormone dependence associated with the more aggressive form of disease, Castrate Resistant Prostate Cancer (CRPC). While the mechanisms involved in the progression of prostate cancer to the castrate resistant form of disease remain undetermined, the continued reliance on circulating hormones means new novel treatment options for the complete removal of circulating hormones could result in better outcomes and an increased life expectancy for prostate cancer patients. The establishment of a human trial demonstrated the potential for advanced prostate cancer specimens to produce testosterone and dihydrotestosterone (DHT) from an earlier precursor, androstenedione. The early data obtained from the human trial will be used to expand the study to obtain statistically significant numbers to help determine the mechanisms involved in the progression of prostate cancer to the castrate resistant phase. Due to the successful implementation of the decoy binding protein in the prostate cancer preclinical setting, preliminary in vivo orthotopic xenograft models were able to demonstrate the potential for the application of the drug therapy to be utilised in other hormone dependent cancers such as breast and ovarian. A modified version of the binding decoy protein was constructed with the ligand binding domain of the estrogen receptor for the use in estrogen driven diseases. The data obtained in this thesis has resulted in the submission of three separate patents.