Surgery (RMH) - Theses
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ItemDeconstructing the brain tumour microenvironment using multimodal analysis( 2023-06)Gliomas are a type of astrocytoma and are the most prevalent type of primary brain cancer, with the most aggressive form being glioblastoma (GBM), with a median survival of only 15 months. Rapid tumour cell invasion and progression is a significant challenge for patients and their oncologists and neurosurgeons, reducing treatment efficacy and inevitably leading to tumour recurrence. Cancer cells thrive by responding and adapting to cellular and non-cellular cues in the tumour microenvironment, including the extracellular matrix (ECM). However, little is known about ECM composition in brain tumours and how the ECM evolves during disease progression, and the impact of the ECM on immune cell localisation, cancer cell signalling and the functional activity of tumour cells. The PI3K and MAPK signalling pathways are typically dysregulated in GBM, and can activate the downstream transcription factor, CREB, which has been reported to regulate GBM malignancy. By integrating multiplex immunohistochemistry, histopathological staining, and spatial tissue analysis, as well as in vitro 3D GBM models, I investigated ECM composition in low- and high-grade glioma, and the spatial relationship between neoplastic cells, immune cells and the ECM in GBM tissue. My results demonstrated a grade-dependent increase in ECM deposition and an upregulation of type I and type IV collagen mRNA expression, which is associated with poor survival in patients with GBM. GBM cells and vascular cells were identified as key contributors of ECM protein deposition in GBM. Spatial analysis demonstrated that T-cells were predominantly located in perivascular niches in ECM-rich regions, while macrophages exhibited more efficient infiltration into tumour cell-rich regions. Extensive tissue remodelling contributes to cellular compartmentalisation in the tumour microenvironment and this compartmentalisation correlates with PI3K, MAPK and CREB activity, and histopathological hallmarks, including angiogenesis, tumour cell density and cell invasion. Inhibiting the PI3K and MAPK signalling pathways reduced 3D cell invasion and also facilitated a shift in the ECM composition, from a more fibrotic to a less fibrotic state. Taken together, the results suggest that the accumulation of ECM plays an important role in GBM progression, affecting both immune cell distribution and cancer cell signalling. These findings suggest that targeting the PI3K and MAPK pathways to ‘normalise’ the ECM could serve to enhance the efficacy of existing and novel therapies for GBM.
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ItemExosomes mediate TGF-β signaling activity and promote breast cancer progression( 2023-03)Metastasis is the main cause of cancer-related deaths. Transforming growth factor-beta (TGF-beta) promotes metastasis by inducing epithelial-mesenchymal transition (EMT). Further, increased TGF-beta signaling is associated with reduced cancer patient survival. Nonetheless, the precise mechanisms underlying TGF-beta signaling activity amplification remain ill-characterized. Also, while on-target anti-TGF-beta inhibitors ameliorate metastasis in pre-clinical cancer models, cancer clinical trials show inconsistent results, highlighting a gap in the understanding of TGF-beta biology. Since our group has shown that cancer cell-exosomes (small extracellular vesicles/sEVs) increase TGF-beta signaling levels in vitro, sEV-induced TGF-beta signaling amplification could underlie metastasis. Analysis of breast cancer datasets revealed a neglectable mutation frequency in exosome-related genes, while their transcriptional levels were inversely associated with cancer patient survival and directly correlated with scores for TGF-beta signaling and EMT levels. In vitro, breast cancer cells were challenged with TGF-beta signaling inhibitors with out without recombinant human (rh)TGF-beta1 or cancer cell-sEVs to investigate alterations by immunofluorescence staining, western blot, scratch assay, and in transwell inserts. Compared with rhTGF-beta1-treated cell cultures, exosomes enhanced breast cancer cell EMT, migration and invasion, which was abrogated by TGF-beta signaling inhibitors. In vivo, exosomes amplified the TGF-beta signaling activity in breast cancer orthotopic xenografts, as quantified by In Vivo Imaging System (IVIS). Moreover, luciferase assay demonstrated that sEV-treatment increased the number of circulating tumor cells, multi-organ metastases, and tumor self-seeding. These results confirm previous findings from our group, further establishing a role for exosomes in driving TGF-beta signaling amplification and breast cancer progression. In addition to cancer cell-exosomes, cancer-associated fibroblast (CAF)-exosomes also contribute to cancer progression. Yet, CAFs are often overlooked in pre-clinical models. Here, we characterize human breast CAF-sEVs and compare their impact on breast cancer cells with that induced by rhTGF-beta1. Treatment with exogenous CAF-sEVs amplifies TGF-beta signaling activity in cancer cells in vitro and in vivo, as quantified by luciferase assay and IVIS, respectively. Accordingly, CAF-sEVs increase cancer cell aggressiveness in vitro and promote cancer progression in vivo. Likewise, CAFs induce TGF-beta signaling amplification in co-culture with poorly invasive MCF7 cells, promoting metastasis and tumor self-seeding. Conversely, genetically and pharmacologically targeting exosome trafficking and TGF-beta signaling reduce CAF-promoted effects. Thus, CAF-exosomes are critical for CAF-induced TGF-beta signaling amplification and breast cancer progression. Considering the role of exosomes, drugs targeting exosome trafficking could reduce TGF-beta-induced metastasis. In vitro, challenging with DMA to decrease exosome secretion reduces TGF-beta signaling levels in highly invasive MDA231 cells in a dose dependent manner. Further, combining DMA and SB431542 (TGF-beta type I receptor kinase inhibitor) at suboptimal doses potentiates this effect and inhibits MDA231 cell migration. Similar effects were observed in vivo where DMA reduces TGF-beta signaling levels and cancer progression, potentializing SB431542 activity when combined at suboptimal doses. In addition to characterizing the relevance of exosomes to TGF-beta signaling amplification, our study establishes a new approach to impair metastasis by simultaneously inhibiting exosome trafficking and TGF-beta signaling. We envision that future research may build on our findings, evaluating the translation of this therapeutic strategy for the treatment of cancer patients.
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ItemThe role of IL-11 signaling in glioblastoma progression( 2022)Glioblastoma is the most common and lethal brain tumour in adults with a mean survival rate of only 12-15 months with current treatment. The microenvironment of a tumour is becoming increasingly more important to current research, with many findings suggesting that the transcription factors driving oncogenic processes are more often due to cytokine stimulation than gene mutation. There have been multiple signalling molecules and corresponding receptors identified as key role-players in the development of glioblastoma, its severity and ability to evade treatment. Cytokines are molecules that initiate and mediate a range of cellular activities essential to the homeostasis of a heathy person but also to tumour growth, invasion and survival. This includes the critical growth factors and cytokines that activate signalling pathways controlling many pro-oncogenic cellular functions. The interleukin-11 (IL-11) cytokine has become increasingly recognised as a driver of the pathogenesis of a wide range of cancers, however, very little is known regarding its role in glioblastoma. Considering this, we hypothesized that IL-11 would contribute to glioblastoma cell viability, migration, invasion and overall tumour progression. We initially identified that IL-11 and its receptor (IL-11RA) inversely correlate with tumour grade and glioblastoma survival. To study the role of IL-11 in glioblastoma, we next determined the expression of endogenous IL-11RA in a range of cell lines and transfected those expressing very little of the gene with the IL-11RA (cell lines #20 and #28). Proteomic analysis was conducted to reveal changes in protein expression after transfection. A large number of proteins involved in proliferation, migration and invasion were seen to be upregulated in the IL-11RA transfected cells. Indeed, the IL-11RA transfected cells displayed significantly greater growth, migration and invasion in proliferation, wound healing, transwell and spheroid invasion assays. This was reversed with IL-11RA knockdown. The proteomic analysis also highlighted the upregulation of proteins involved in metabolism, particularly glutaminolysis and inhibition of apoptosis. Metabolomic analysis revealed the IL-11RA transfected cells displayed increased levels of glutamine oxidation, as well as increased proliferation and survival of these cells in conditions of depleted glucose or glutamine. Similarly, IL-11RA transfected cells displayed no significant difference in invasion rate in the presence or absence of glucose, when glutamine was available. Alternatively, blocking both glucose and glutamine metabolism with a number of drugs significantly reduced the proliferation, migration and invasion of these cells. Our findings suggest that the IL-11RA transfected cells are able to utilise alternative metabolites such as glutamine, in the absence of glucose, in order to proliferate, migrate and survive. Overall, the results of this thesis suggest that the IL-11RA plays an important role in proliferation, migration, invasion, survival and metabolism in glioblastoma.
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ItemCirculating Biomarkers in Glioma( 2022)The identification of accurate circulating biomarkers, otherwise termed “liquid biopsy,” is a major goal in oncology research due to the potential clinical applications in population screening, tumour monitoring and delivery of individualised treatments resulting from tumour genotyping. Recently, large datasets have been made available due to advances in genome sequencing and mass spectrometry, however the difficulty remains in translating these findings to clinically meaningful applications. Gliomas are the most common primary brain cancer and its most aggressive form, glioblastoma (GBM), is rapidly and uniformly lethal.1 Circulating biomarkers have the potential to aid in a number of challenges that are faced in managing these patients. Despite increased efforts in biomarker discovery, currently there is no validated liquid biopsy for glioma. In this thesis, three candidate biomarkers are investigated for use in several clinical applications. Firstly, the small non-coding RNA sequence microRNA, is shown to be able to monitor for glioma progression, assess tumour burden and improve prognostic predictions. Following this, reliable detection of circulating tumour DNA (ctDNA) in plasma of patients is demonstrated using highly sensitive next generation sequencing and PCR techniques. The potential of ctDNA as a diagnostic adjunct for complete molecular tumour characterisation is shown, as well as the ability of ctDNA to monitor an individual tumour’s genomic evolution including identifying early in the blood key gene mutations associated with chemoresistance. Finally, a pilot study is presented using spiral microfluidics, confirming the presence of circulating tumour cells in the blood of glioma patients as a method to increase tumour DNA concentration.
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ItemIdentifying the Risks and Drivers of Aggression in Prostate Cancer( 2022)Prostate cancer is the most commonly diagnosed cancer and the second commonest cause of death in Australian men. The natural history of localised prostate cancer is markedly variable. Most individuals will lead a biologically indolent course where it may never be clinically significant while an unpredictable minority of individuals, approximately 10%, will rapidly progress to metastases and eventually succumb to the disease. Currently there are no adequate methods in determining the difference between an indolent or aggressive course of the disease at an early stage; therefore, leading to significant over-treatment of indolent disease as well as under-treatment of aggressive disease. The identification of potential drivers of aggression in prostate cancer are explored through the compilation of published papers in an integrated thesis by investigating the impacts of obesity, velocity of biochemical recurrence, aggressive ductal adenocarcinoma variant, as well as the molecular comparison between primary and matched metastatic tumours. Obesity is linked with more aggressive prostate cancer and higher rates of disease recurrence post treatment. It is unclear if this is due to specific tumour-promoting effects of obesity or diagnostic bias. The effect of obesity on the accuracy of pre-treatment risk categorisation was determined, and mediation analysis was used to identify the contribution of biologic versus non-biologic mechanisms to the observed increased risk of biochemical recurrence. The analysis included 1587 patients. Despite similar rates of adverse pathological features at prostatectomy, biochemical recurrence rates were significantly higher in very obese patients, which persisted after adjustment for stage, grade and PSA. Tumour volume however correlated significantly with BMI (p = 0.004), and the difference in predicted and observed ‘tumour attributable’ PSA (Delta-PSA) in very obese patients was greater than three times higher than that of healthy patients (p = 0.0067). Regression analysis indicated that the effect of BMI on tumour volume was fully mediated indirectly by its effect on PSA. Inclusion of this diagnostic error as a covariate in the survival analysis attenuated the effect of BMI on recurrence. Being very obese suppresses tumour-associated PSA resulting in a diagnostic bias that is responsible for errors in risk classification, and potentially contributes to a delay in initial presentation. Radical prostatectomy is one of the preferred treatment modalities for localised prostate cancer. Although the majority of patients experience long term disease control, depending on the pre-treatment clinical characteristics of the cohort under study, up to a third of men will develop disease recurrence. The most common manifestation of disease recurrence is a detectable serum PSA in the postoperative period. This thesis characterises the pattern of late disease recurrence in the largest contemporary cohort of localised prostate cancer patients treated with radical prostatectomies in the active surveillance era. Total of 2312 patients were included in the final analysis with up to 12 years of follow up data. The average patient had clinically localised prostate cancer, an elevated PSA, and ISUP grade group 2 on biopsy. 88.7% of patients had ISUP grade group 2 or higher at prostatectomy. A subgroup of 446 patients had undetectable PSA levels at 5 years after prostatectomy; 11.7% of them progressed to experience biochemical recurrence. In this subgroup, late recurrers had significantly higher-grade tumours on ISUP and Gleason sum (p < 0.001 and p = 0.001, respectively), higher rates of extraprostatic extension (p = 0.022), and larger tumour volumes (p = 0.032). Logistic regression demonstrated that prostatectomy ISUP grade group was a significant predictor (OR 2.14, 95% CI 1.43-3.20, p < 0.001). Additionally, the timing of recurrence resides on a continuum of risk and that the potential concept of dormant micrometastatic involvement requires further research and evaluation. Ductal adenocarcinoma is an uncommon prostate cancer variant. Previous studies suggest that ductal variant histology may be associated with worse clinical outcomes, but these are difficult to interpret. Prostatectomy patients with ductal variant histology from two institutional databases were identified and compared to an independent acinar adenocarcinoma cohort. A total of 202 ductal adenocarcinoma and 2037 acinar adenocarcinoma cases were analysed. Deep whole exome sequencing was performed in selected cases (n = 8). Survival analysis after matching demonstrated that patients with ductal variant histology had shorter salvage-free survival (8.1 vs 22.0 months, p = 0.03) and metastasis-free survival (6.7 vs 78.6 months, p < 0.0001). Ductal variant histology was consistently associated with RB1 loss, as well as copy number gains in TAP1, SLC4A2 and EHHADH. The presence of any ductal variant adenocarcinoma at the time of prostatectomy portends a worse clinical outcome than pure acinar cancers, with significantly shorter times to the initiation of salvage therapies and the onset of metastatic disease. The exploration of the impacts of obesity, velocity of biochemical recurrence, aggressive ductal adenocarcinoma variant, as well as the molecular comparison between primary and matched metastatic tumours to provide insights into potential drivers of aggression in prostate cancer.
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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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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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