Surgery (RMH) - Theses
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ItemImpact of polyacrylamide hydrogel (Bulkamid®) in the management of stress urinary incontinence in women( 2021)Stress urinary incontinence (SUI) is a highly prevalent condition among women, with a significant impact on quality of life. Although mid-urethral sling (MUS) is widely considered the reference standard treatment following failure of conservative measures, increasing concern over the risk of mesh morbidity has resulted in it falling out of favour. Instead, more women are choosing to undergo urethral bulking agent treatment as a minimally invasive alternative, despite a lower efficacy rate to open surgery. Despite urethral bulking agents being a well-established treatment in women with SUI, there is a paucity of data to guide its use in clinical practice. Currently marketed urethral bulking agents include polyacrylamide hydrogel (Bulkamid) polydimethylsiloxane (Macroplastique), carbon-coated zirconium oxide (Durasphere), calcium hydroxylapatite (Coaptite) and polymerising polydimethylsiloxane silicone gel (Urolastic). Clinical data comparing the outcomes of these agents are limited. This thesis assesses and compares all available urethral bulking agents used in the treatment of SUI in women. Variable mean success rates of 30%-80% are reported in the short-term. Better long-term success rates were found with Bulkamid, Coaptite and Macroplastique on qualitative review. The majority of urethral bulking agents are reported to be safe, with less frequent adverse events such as urinary tract infection, temporary acute urinary retention and de novo urgency reported. More significant complications such as migration into lymph nodes and erosion have also been reported yet are rare. Despite the common use of polyacrylamide hydrogel urethral bulking agent injections since its introduction in 2006, long-term clinical data are limited. This thesis demonstrates the long-term outcomes of polyacrylamide hydrogel (Bulkamid) transurethral injections in an Australian cohort of women with SUI performed by a single surgeon. 21% of women did not respond to primary polyacrylamide hydrogel (Bulkamid) treatment and proceeded to alternative anti-incontinence surgery. As opposed to the common perception of urethral bulking agents being a short-term therapy, requiring frequent repeat injection, 53% of women at 7-8 years post initial injection self-reported a successful outcome. Polyacrylamide hydrogel (Bulkamid) injection was associated with benefits on other important patient-reported outcomes such as urinary incontinence-related symptom distress and life impact. Short-term adverse events were infrequent and mild and there was no serious long term adverse event. Knowledge of factors associated with superior outcomes in women treated with urethral bulking agents for stress urinary incontinence remains limited yet could help clinicians better select and counsel patients on expected outcomes. This thesis explores factors associated with polyacrylamide hydrogel (Bulkamid) treatment success in women with SUI, and demonstrates that women with type 3 urethral hypermobility, a well-supported urethra, were more likely to report treatment success than women with non-type 3 urethral hypermobility before treatment. Poor bladder compliance before treatment was associated with higher urinary symptom distress, and higher severity and frequency of urinary incontinence post-treatment. Older age was associated with higher levels of self-reported urinary frequency and severity post-treatment. Finally, the severity of pre-treatment incontinence impact was associated with worse incontinence impact post-treatment. Findings from this thesis will assist clinicians in the selection of urethral bulking agents. It will also assist clinicians in the selection of patients most likely to benefit from polyacrylamide hydrogel (Bulkamid) urethral bulking agents injection treatment, and in the counselling of expected long-term efficacy and safety outcomes.
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ItemFocused ultrasound as an adjunct to clinical evaluation for patients admitted to general medicine units( 2021)General internal medicine physicians have started to incorporate point-of-care ultrasound (POCUS) into their clinical practices. Limited data is available on POCUS use in internal medicine. This thesis aimed to evaluate the clinical impact of adding POCUS to the initial assessment of patients hospitalised in internal medicine units through three main research projects. First, a systematic review was conducted to investigate POCUS' clinical impact on hospitalised internal medicine patients. Five previous studies have addressed this question differing in their design, intervention, and outcomes reported. Two observational studies described the influence of POCUS on the diagnosis formulation. POCUS use changed in the principal diagnosis and added relevant new diagnoses occurred in up to 18% and 24 % of the cases, respectively. Impact on the management plan was reported in 37% to 52% of the participants as a composite outcome including change in medications, additional testing, change in prognosis or change in discharge time. Two randomised controlled trials (RCTs) addressed the effect of POCUS on the length of hospital stay. One study reported no difference between the groups, and the other study found a reduction of one day using serial lung ultrasound in patients admitted with heart failure. These studies were assessed as having moderate to severe risk of bias, which highlights the need for high-quality studies investigating the effect of POCUS on clinical outcomes. Subsequently, an RCT was conducted at the Royal Melbourne Hospital, Victoria, Australia that tested the impact of adding a multiorgan POCUS exam to the initial assessment of cardiopulmonary admissions on the length of hospital stay, clinical decision-making process, readmissions and health costs. Two hundred fifty participants were enrolled and randomised to intervention or control group. The intervention was a POCUS exam of the heart, lungs, and lower extremities (2-point venous compression) performed in the first 24 hours of admission to the unit. POCUS identified new pathology in 70% and changed the primary diagnosis in 28 %, medical treatment in 28%, and imaging tests in 60% of the subjects. However, there was no significant difference between the POCUS and control groups in the hospital length of stay, (POCUS 113 hours vs. control 125 hours, p=0.53), readmission rates (POCUS 16 % vs. control 12%, p=0.43) and total hospital costs ($7.8K vs. $7.9K, p=0.79). Finally, this thesis reports a prospective observational study assessing the feasibility and effectiveness of a heart and lung POCUS training program delivered to internal medicine physicians. The study identified the potential barriers of implementing POCUS' training programs in Australian hospitals. Moreover, it showed that a combination of electronic learning material, ultrasound simulators and supervised clinical rounds effectively improved participant's knowledge, image acquisition and interpretation skills. Overall, this thesis has generated substantial data on the impact of using POCUS on the clinical decision-making process performed by the treating physician and on patient's outcome, such as the length of hospital stays. Moreover, it has explored a POCUS training program for general internal medicine physicians in Australian hospitals.
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ItemEvaluating the role of invadopodia in glioma invasion and response to therapeutics( 2021)Glioblastoma (GBM) is the most prevalent and aggressive form of glioma, and is associated with an extremely poor prognosis, with a low median patient survival time of just 15 months post-diagnosis with the current therapeutic approach known as the Stupp protocol, consisting of surgical resection, followed by radiotherapy (RT) and concomitant chemotherapy with temozolomide (TMZ). A significant contributing factor that impacts the survival of GBM patients is the highly infiltrative nature of GBM cells, which prevents complete tumour resection and also limits the capacity of targeted therapies to effectively reach the infiltrating tumour cells. Consequently, these tumours can exhibit high rates of recurrence, appearing within months following the completion of the first round of treatment and can also demonstrate minimal response to further rounds of RT/TMZ treatment. Evidence suggests that the efficacy of current therapeutic approach may be compromised by an enhanced invasive phenotype that is displayed by the GBM cells that survive the current treatment protocol (Wild-Bode, Weller et al. 2001, Cordes, Hansmeier et al. 2003, Hegedus, Zach et al. 2004, Trog, Fountoulakis et al. 2006, Trog, Yeghiazaryan et al. 2006, Steinle, Palme et al. 2011). The targeting of the enhanced invasive abilities exhibited by RT/TMZ treated GBM cells could provide a potential therapeutic approach for improving patient outcome, however the mechanisms utilised by invasive GBM cells following the current treatment are not well understood. As GBM cells have been shown to form actin-rich membrane protrusions known as invadopodia that can facilitate invasion by degrading the surrounding ECM via highly localised proteolytic activity (Stylli, Kaye et al. 2008, Mao, Whitehead et al. 2017, Petropoulos, Guichet et al. 2018), it is possible that the enhanced invasive capabilities of GBM cells post- RT/TMZ treatment may be mediated by invadopodia. In this thesis, the role of invadopodia in GBM cell invasion and response to RT/TMZ treatment was investigated. Using clinically relevant doses of RT and TMZ, it was demonstrated that the enhanced invasive capabilities of GBM cells post-RT/TMZ treatment may be attributed to an increase in invadopodia formation and activity. The role of intracellular communication between GBM cells via small extracellular vesicles (sEVs) was also investigated, highlighting the ability of GBM cell line secreted sEVs to transfer a pro-invadopodia phenotype to recipient GBM cells, as well as their potential to facilitate an enhanced pro-invadopodia phenotype following RT/TMZ treatment. Demonstrating the potential to dualistically target invadopodia activity and sEV secretion to overcome RT/TMZ-induced GBM invasion, the addition of the microtubule-targeting agent Vinorelbine Tartrate (VT) alongside RT/TMZ reduced the enhanced secretion of sEVs, in accordance with previous data from our laboratory showing VT also reduces invadopodia activity in GBM cells surviving RT/TMZ (Whitehead, Nguyen et al. 2018). Lastly, GBM cell lines and their corresponding secreted sEVs were subjected to comprehensive proteomic profiling to identify proteins that may facilitate invadopodia formation and activity following exposure to RT/TMZ treatment, thereby contributing to enhanced GBM invasion. Collectively, this work highlights the contributing role of invadopodia and sEVs to the pro-invasive abilities of GBM cells, and provides insight into the dysregulated proteomic landscape of GBM cells and sEVs following exposure to RT/TMZ treatment that may contribute to enhanced invasive capacity, which may ultimately assist in the development of novel adjuvant therapeutic strategies to improve the clinical efficacy of RT and TMZ treatment.
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ItemUnderstanding and overcoming resistance to epidermal growth factor receptor therapy( 2020)Colorectal cancer (CRC) is the fourth most common cancer diagnosed in Australia. Current standard treatment includes surgery, chemotherapy, and targeted therapy. Cetuximab is often used as part of the clinical management of unselected patients until a subset of patients were found to harbor KRAS mutations that conferred intrinsic resistance to Cetuximab. In addition, some patients are resistant to Cetuximab despite having wild-type KRAS. Using RNA-sequencing data and differential expression analysis, we discovered five potential biomarkers for predicting resistance to Cetuximab in CRC with wild-type KRAS. After generating 3 CRC models of acquired resistance to Cetuximab, we also employed proteomics analysis to determine potential biomarkers of acquired Cetuximab resistance in CRC cells with wild-type KRAS. In addition, we generated pre-clinical data for the repurposing of Carfilzomib (CFZ) as a novel drug to overcome Cetuximab resistance in metastatic colorectal cancer patients. In conclusion, this study provides a Cetuximab resistant model that can be used for further studies coupled with possible resistance mechanisms as well as a novel drug to overcome Cetuximab resistance.
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ItemColorectal Cancer resection outcomes using administrative data( 2020)Background Colorectal cancer is the second leading cause of cancer death in Australia. If found early, a patient can undergo potentially curative surgery. Even in the third of patients that do recur, there can be palliation or even cure with further surgery with or without chemotherapy. Although surgery remains a vital tool in treating colorectal cancer, it is not without complications. The principal studies of this thesis used administrative data to report on short- and long-term outcomes following resection for colorectal cancer in the state of Victoria over a ten-year period. Methods Administrative data are collected on all patients admitted to Australian Hospitals. Patient demographics, co-morbidities, type of operation, post-operative complications, histopathology and some staging information are recorded. Trained coders review clinical notes and then assign alphanumeric codes to these data based on the International Classification of Diseases Tenth Revision, Australian Modification (ICD-10-AM). These codes were developed for the purpose of billing and therefore may not be focused on reporting data in a clinically relevant fashion. We have previously shown that the use of algorithms of code combinations can increase the accuracy of this data source for clinical research1. This thesis added laparoscopic detail to these coding algorithms. These algorithms were then applied to a central repository of administrative data in Victoria, to report on short-term outcomes following resection for colorectal cancer comparing regional to metropolitan hospitals. Results were adjusted for potential confounding variables using a multivariable logistic regression analysis. This data source was then linked to death data to report on overall long-term survival following colorectal cancer surgery, comparing regional to metropolitan hospitals. Survival results were presented as a rate, adjusted for potential confounders using a multivariable Cox regression analysis. Results These studies found strong evidence for lower odds of prolonged length of stay (OR 0.53, 95% CI 0.48 – 0.58, p=<0.001) and inpatient mortality (OR 0.67, 95%CI 0.49 – 0.91, p=0.01) in inner regional hospital compared with metropolitan hospitals. For outer regional hospitals, there was strong evidence of decreased odds of prolonged LOS (OR 0.64, 95%CI 0.52 – 0.77, p=<0.001) and return to theatre (OR 0.67, 95%CI 0.47 – 0.95, p=0.03). There was no difference in overall survival comparing colorectal cancer resection patients from inner or outer regional hospitals to metropolitan ((HR 1.02, 95%CI 0.95 – 1.09, p=0.59) and (HR 0.97, 95%CI 0.85 – 1.11, p=0.68) respectively). Conclusion These studies demonstrated the strength of administrative data with validated algorithms and data linkage in reporting on outcomes following colorectal cancer resection. This methodology resulted to two of the largest and most detailed studies concerning colorectal cancer resection in Australia. Importantly, they validated current practices in Victoria by revealing similar outcomes in regional and metropolitan centres after resection for colorectal cancer.
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ItemGlioblastoma: Treatment Stagnation and Cellular and Molecular Mechanisms( 2020)Glioblastoma, a WHO grade IV primary brain tumour, remains as one of the most aggressive forms of human cancer. Despite intensive research efforts into understanding the key drivers of tumour progression, few therapeutic advances have been made, with the current standard of care (the Stupp protocol) remaining unchanged for 15 years. The overall improvement to glioblastoma survival in the real-world population has been attributed to the use of the Stupp protocol, yet evidence suggests that survival outcomes were already significantly improving in the years prior to the introduction of this standard of care questioning the overall veracity of this claim. Using the Surveillance, Epidemiology and End Results (SEER) registry data we analysed the survival outcomes for real-world glioblastoma patients diagnosed from 2000 – 2016. Our findings show a consistent incremental survival improvement that preceded the introduction of the Stupp protocol and continued to increase at the same rate till 2009, stagnating afterwards. Significantly, however, this survival improvement is short-term for patients, with no survival improvement observed in patients surviving more than 2 years. Additionally, with the exception of complete tumour resection, all treatment modalities did not improve survival beyond 2 years for glioblastoma. These findings highlight the clinical stagnation of glioblastoma treatment and highlight the inability of current treatments to target the underlying causes of tumour progression. Following the introduction of the Stupp protocol attempts to develop new treatment options have universally been disappointing with a close to 0% success rate for over 1000 phase II and above clinical trials. Conflictingly, many of the therapeutic agents tested have shown promising results in preclinical trials. Current preclinical models, however, test therapies against the primary tumour, which does not recapitulate the biology or targets of tumour recurrence. We therefore developed a highly sensitive luciferase-based glioblastoma mouse model capable of single cell detection in mouse tissue. Analysis of mouse brain tissue implanted with luciferase-labelled human glioblastoma U87MG or MU20 tumours revealed the presence of tumour cells ubiquitously spread across the supratentorial regions of the brain, and distally located from the primary tumour. These tumour cells were observed as single cells in U87MG implanted mice and clusters in MU20 glioblastoma cells. Remarkably, U87MG tumours did not exhibit invasive margins and were contained within an expansive growth phenotype, suggesting invasion-independent dissemination. Our model is consistent with reports of glioblastoma as a systemic brain disease and is capable of sensitive detection of disseminated tumour cells, a model of recurrence potential. Furthermore, this model can be utilised to investigate new mechanisms of glioblastoma infiltration. Targeting aberrant angiogenesis in glioblastoma has been the major focus for glioblastoma treatment since the Stupp protocol. Yet after over a decade of basic research and clinical trials, antiangiogenic inhibitors have failed to translate into improved patient outcome. The discovery of abnormalities in the tumour vasculature suggest that there may be alternate mechanisms driving tumour progression. Vasculogenic mimicry has been observed in glioblastoma and presents as a novel aspect of tumour biology, yet the mechanisms and functional relevance of these structures remain unknown. Our study has confirmed the ability of some glioblastoma cell lines to undergo endothelialisation, forming lattice structures similar to endothelial cells when seeded onto Matrigel in vitro. One lattice forming cell line, U87MG, was also found incorporate into the tumour vasculature in an in vivo orthotopic mouse model. This behaviour was found to be regulated by an expanded TGF-beta-ALK1-Smad1/5 signalling pathway. In vivo inhibition of the Smad1/5 signalling pathway via intracranial treatment with Ad-Smad6 resulted in reduced endothelialisation in the tumour vasculature and inhibited whole brain infiltration in the U87MG mouse model. Since U87MG xenograft tumours are non-invasive, these results suggest that endothelialisation may lead to haematogenous dissemination and distal brain infiltration, providing a novel mechanism for glioblastoma progression.
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ItemCirculating miRNAs as a novel biomarkers and intercellular regulators in glioma( 2018)Glioma is the most common intracranial malignant cancer despite the rarity. Glioblastoma (GBM, grade IV) has very dismal outcomes where fewer than 20% of patients survive beyond 5 years. Low Grade Glioma (LGG, grade II), on the other hand, frequently occurs in a younger population with a variable outcome that depends on their individual genetic alterations. A tissue-based biopsy requires an invasive operation for every glioma patient and it is the gold standard diagnosis. In monitoring the patients for recurrence, MRI has the evitable limitation that it could not differentiate pseudo-progression after irradiation treatment for glioma patients. Therefore, a non-invasive biomarker, in addition to tissue biopsy and MRI, is urgently needed to provide more precise cancer care and management with glioma patients. In fact, cancer cells including glioma cells, release microRNA (miRNA) into the microenvironment and peripheral circulation, which allows physicians and scientists the ability to detect the disease status by a blood test. In addition, microRNA (miRNA) displays accuracy in diagnosing and monitoring other cancer patients by analysis of miRNA abundance. To investigate the diagnostic capacity of circulating miRNA in glioma, a meta-analysis of previous relevant studies was performed. Circulating miRNA provided an over 90% accuracy of diagnosing glioma patients from healthy controls. A bioinformatic analysis revealed seven IDH1 mutation-associated miRNAs and these miRNA signatures were down-regulated in IDH1 mutant glioblastoma due to the hypermethylation in their promoter areas. Our proof-of-concept cohort recruited 91 glioma patients and 17 healthy controls and new circulating miRNAs were identified with remarkable diagnostic capacity. However, no circulating miRNA showed an association with IDH1 mutation in our cohort. As most circulating miRNAs were packaged and delivered by extracellular vesicles (EVs), serial cell experiments were conducted to examine the function of these miRNA-enriched EVs. Glioma Stem Cells-derived EVs significantly promoted glioma cell proliferation, cell migration and radiation resistance. This effect was largely caused by the degradation of PTEN and activation of AKT in the recipient cells through miRNAs. Collectively, these works established circulating miRNA as a clinical biomarker where it provides a complementary tool for glioma management. These results also formed the basis of future research into the role of EVs in glioma and highlighted the potential therapeutic target of EVs to glioma.
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ItemThe role of receptor tyrosine kinases in mediating glioblastoma resistance to radiotherapy and temozolomide( 2020)Glioblastoma is the most common and aggressive form of malignant glioma. Currently, despite treatment with surgery followed by radiotherapy and the chemotherapeutic agent temozolomide (TMZ), mean patient survival time is approximately 12 months and the 5-year survival rate is close to 0%. A key factor for the dismal prognosis is tumour recurrence post-treatment which is largely due to: 1) the infiltrative nature of glioblastoma rendering complete resection impossible and 2) glioblastoma cell resistance to radio-chemotherapy. In this thesis we aimed to investigate the cellular mechanisms of receptor tyrosine kinases in conferring resistance to therapy. We first performed a literature search and found that almost all studies that advocated for the utility of targeting RTKs in overcoming treatment resistance did not employ both therapeutic agents comprising standard therapy – radiotherapy and TMZ. We next generated an in vitro glioblastoma resistant model via short-term treatment with radiotherapy and TMZ and found that these cells had down-regulated RTK activity in addition to down-regulated protein and gene expression of the commonly altered and studied epidermal growth factor receptor (EGFR) and MET receptor. After generating an in vitro glioblastoma recurrent model via long-term treatment we demonstrated that the surviving sub-population of cells also displayed down-regulated EGFR and MET expression compared to treatment naive cells. Furthermore, we also showed that the resistant cell population already pre-exists within the parental population which suggests the possibility of pre-emptively targeting the inherently resistant population. Interestingly, we also observed differential microRNA expression in radiotherapy- and TMZ-treated cells and, specifically, found that miR-221 confers resistance to glioblastoma cells and is capable of down-regulating EGFR expression. We validated this relationship in a human cohort of 105 primary and 36 recurrent glioblastoma patients, showing a significant inverse relationship between miR-221 and EGFR. Consistently, we showed that high miR-221 and low-EGFR expression at recurrence is associated with a poorer prognosis. Lastly, we investigated the relevance of epithelial to mesenchymal transition markers after observing that migration rates were maintained in resistant cells despite low EGFR and MET. Both N-Cadherin and CD44 were found to be highly expressed in treatment-resistant cells and the down-regulation of AKT activity with wortmannin led to reduced levels of EMT markers, suggesting that AKT is a regulator of key EMT transcription factors that are specific to N-Cadherin and CD44. The thesis gains it significance by providing an explanation to the failure of RTK inhibitors in the glioblastoma clinic by suggesting that standard radio-chemotherapy down-regulates RTK activity and expression, thereby diminishing any theorised benefit of targeting RTKs. Furthermore, the thesis advocates for microRNAs to be crucial regulators of therapy resistance, potential biomarkers and targetable molecules for the clinic.
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ItemThe clinical significance of cyclin E1 deregulation in high grade serous ovarian cancer and basal like breast cancer( 2019)High grade serous ovarian cancer (HGSOC) and basal like breast cancer (BLBC) are genomically unstable and aggressive cancers that frequently co-occur and share common molecular features. Of these molecular characteristics are P53 inactivation occurring in almost all cases of HGSOC and BLBC, BRCA1/2 inactivation reported in more than 50% of both cancers and CCNE1 amplification reported in up to 30% and 8% of HGSOC and BLBC respectively. Both HGSOC and BLBC are currently grouped in many clinical trials to test new drugs or drug combination, for instance, PARP inhibitors in the context of BRCA1/BRCA2 mutation. We asked whether deregulated cyclin E1 (CCNE1 amplification and/or its encoding protein, cyclin E1, overexpression) is an additional biomarker that can potentially be used to group patients with both diseases for therapeutic purposes. We studied two well characterised cohorts of 262 HGSOC and 222 familial breast cancer (BLBC enriched) samples of formalin fixed paraffin embedded sections. HGSOC and the BLBC enriched cohort were from patients enrolled in the Australian ovarian (AOCS) and the Kathleen Cuningham Foundation Consortium for research into Familial Breast cancer (KConFab) respectively. Using automated tissue based assay and an in situ hybridization probe that spans 19q12 locus harbouring CCNE1, we assessed the level of CCNE1 amplification. We also assessed the expression of cyclin E1 and a cyclin E1 degradation associated protein FBXW7 and a cyclin E1 deubiquitinase, USP28, by immunohistochemistry, as possible drivers of high cyclin E1 expression in amplified and non-amplified cyclin E1hi subsets. We also assessed the expression of URI1 in our HGSOC cohort. URI1 is a protein encoded by the URI1 gene which co-localise with CCNE1 on 19q12 locus. In HGSOC, we identified seemingly two separate subsets of cyclin E1hi tumors that have different pathological and biological characteristics as well as different clinical outcomes. These are the amplified/cyclin E1hi group that had amplification and high expression of cyclin E1, low expression of FBXW7, higher genomic instability, intact BRCA1/2 and worse outcome. The other is the non-amplified/cyclin E1hi tumors that typically had high expression of cyclin E1 in the absence of amplification, high USP28 expression, lower genomic instability, more prevalent BRCA1/2 loss and more favorable outcome compared to the amplified group. Next we assessed cyclin E1 deregulation in the overlapping groups BRCA1 mutant breast cancer and BLBC. Both subtypes had significantly higher expression of cyclin E1 and amplification compared to other breast cancer types. However, the intensity of cyclin E1 expression and level of 19q12 amplification were lower in BLBC compared to those observed in HGSOC. Moreover, in BRCA1 mutant breast cancer and BLBC patients, only high expression of cyclin E1 was associated with lower overall survival while amplification did not seem to impact outcome. These observations were further supported by our meta-analysis that included our cohorts as well as other published datasets. In the meta-analysis, both CCNE1 amplification and cyclin E1 expression were found to be adverse prognostic factors in HGSOC while only high expression was associated with worse survival in BLBC patients. In fact, both amplified and non-amplified cyclin E1hi BLBC subsets shared almost all cyclin E1 deregulation associated features as well as many features with the non-amplified cyclin E1hi HGSOC subset. Of these are the prevalence of high expression of the cyclin E1 deubiquitinase, USP28, BRCA inactivation, the lower genomic instability and cyclin E1hi linked adverse outcome. In order to provide better therapeutic options for cyclin E1hi BRCA1 mutant breast cancer/BLBC patients, we sought to further assess mechanisms behind the co-occurrence of cyclin E1 overexpression and BRCA1 inactivation. Using the KConFab cohort we have found that BRCA1 loss correlated with decreased phosphorylation of cyclin E1, on Threonine 62, assessed by immunohistochemistry. We also showed by in vitro analysis that BRCA1 loss in cell lines led to cell cycle specific stabilisation of cyclin E1 by reducing cyclin E1 T62 phosphorylation. Conversely, BRCA1 overexpression increased T62 phosphorylation. Overexpression of cyclin E1 with an inactivated T62 site, to mimic loss of phosphorylation, increased cyclin E1 stability and resistance to Paclitaxel. These findings suggest that BRCA1 regulates cyclin E1 stability in breast cancer cells via regulating T62 phosphorylation. We next assessed a combination therapy that target cyclin E1 and BRCA1 inactivation using CDK2 and PARP inhibitors. CDK inhibitors are suggested to induce DNA damage and therefore we hypothesised that CDK2 inhibition would enhance sensitivity of BRCA1 deficient cells to PARP inhibition. Our finding is that CDK2 inhibition induced DNA damage and synergised with the PARP inhibitor Rucaparib in BRCA1 mutated cell lines. Combination treatment of xenograft are in progress but the preliminary data is supportive of our hypothesis. Our results propose a new therapeutic strategy for BRCA1-mutant breast cancer/BLBC by combining CDK2 and PARP inhibitors to enhance synthetic lethality. As this group shares similarities with non-amplified cyclin E1hi HGSOC subset, we suggest that this combination is likely to be effective in the comparable HGSOC subset.
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ItemThe Postoperative Quality of Recovery Score: validation of its cognitive domain and feasibility analysis of its use in an interventional trial and in providing individualised real-time recovery data( 2019)“Knowledge is power. Information is liberating.” Kofi Atta Annan (1998). Modern postoperative recovery assessment has progressed from that which was focused purely on the physiological restitution in the immediate postoperative period to one that is multidimensional, individualised, dichotomised and provided in real time. The Postoperative Quality of Recovery Score (PostopQRS) is an extensively validated multidimensional recovery assessment tool that has been developed de-novo for assessment of recovery in postoperative patients, and has been widely adopted as an outcome measure in observational trials. It is unique in its ability to provide individualised recovery information to each patient, and in its assessment of a patient’s postoperative recovey in relation to their own unique preoperative baseline performance. Through its digital interface, it has the potential to provide individualized, contemporaneous recovery data to each patient, thus keeping patients informed of their own recovery throughout their postoperative journey, and thus potentially improving their ultimate postoperative outcome. Prior to this thesis, the PostopQRS’ cognitive domain, whilst based on widely accepted neurocognitive tests, was yet to demonstrate clinical face validity in its assessment of cognitive recovery, and was currently unable to assess cognitive recovery in patients who score low on preoperative cognitive baseline testing. Simmilarly, the PostopQRS had been widely adopted as an outcome measure in observational trials but was yet to be implemented in interventional studies. Furthermore, the utility of the PostopQRS in providing real-time recovery assessment was yet to be fully explored. The EchoNOF-I pilot study demonstrated the clinical utility of the PostopQRS as an outcome measure in the interventional trial setting, and demonstrated feasibility with providing fractured neck of femur surgery patients with focused point of care ultrasound. This study highlighted the need for there to be a validated method with which to measure cognitive recovery in patients with low cognitive baseline PostopQRS scoring. Head-to-head comparison of patient cognitive performance on both the PostopQRS and formal neurocognitive test battery was performed, both at preoperative baseline and during postoperative recovery. This demonstrated clinical face validity in defining patients as having low, as opposed to normal, PostopQRS baseline performance, and demonstrated face validity in the proposed method with which to score cognitive recovery in thse patients with low cognitive baseline scoring. The RTR-I pilot study was ground-breaking as it was the first study in which the PostopQRS was used as the intervention itself, and which demonstrated the clinical feasibility of the PostopQRS in providing patients with individualised real-time recovery information throughout their recovery journey. This study has the potential to revolutionise recovery assessment from one that has been traditionally research focused, to one that has direct clinical applications that may ultimately improve patient outcomes postoperatively. Work from this thesis has formed the basis for three additional multicentre randomised control trials (RTR-II, ECHONOF –II and ECHONOF –III), one MACH (Translational Research Projects 2019), one ANZCA (ANZCA Projects 2019) and two NHMRC grant applications (Project Grants 2019, MRFF Keeping Australians Out of Hospital 2019). The modified scoring system with which to measure cognitive recovery in patients with low cognitive baseline PostopQRS scoring has been adopted into the PostopQRS.com website. What is truly exciting is the future post-doctoral exploration of the potential for the PostopQRS to provide individualised real-time recovery information to both patient and health care provider, and through this, ultimately improve patient outcomes.