Uncontrolled cell growth may lead to pathological conditions such as cancer. During the progression of cancer, cancer cells stimulate endothelial cells for angiogenesis to support their growth and migration. Previous studies suggest that Astragalus membranaceus, of which the dried root [Astragali Radix] is used as a traditional Chinese medicine, and its bioactive components, astragalus saponins (AST), astragaloside IV (AS IV) and isoflavonoid calycosin, inhibit cancer growth. The present study aimed to examine whether or not these components inhibit the growth and/or metastasis of colon cancer cells and/or angiogenesis of endothelial cells, and to determine the possible mechanisms involved. The growth of HCT 116 colon cancer cells and human umbilical vein endothelial cells (HUVEC) after 72 hours incubation with AST (1 to 25 μg/ml), AS IV (0.5 to 100 μM) or calycosin (10 to 200 μM) were detected with thiazolyl blue tetrazolium bromide assay. Wound healing migration and tube formation assays were used to examine the metastatic and angiogenic potential of HCT 116 cells and HUVEC. Moreover, the expressions of apoptotic [B-cell lymphoma 2 and procaspase-3] and metastasis/angiogenesis-related proteins [matrix metalloproteinase (MMP)-2, MMP-9 and vascular endothelial growth factor (VEGF)] were measured with Western immunoblotting. To investigate the potential mechanism(s) through which astragalus components affect the proliferation and/or migration of HCT 116 cells and HUVEC, the activities of mitogen-activated protein (MAP) kinases [extracellular signal-regulated kinase 1 and 2 (ERK1/2), p38 MAP kinase (p38) and c-Jun amino-terminal kinases] were studied by measuring the expressions of their phosphorylated and total proteins with Western immunoblotting. Calycosin (200 μM) inhibited the growth of HCT 116 cells without affecting that of HUVEC. While it inhibited the migration of both cell types, it stimulated tube formation only in HUVEC. In HCT 116 cells, calycosin downregulated the expressions of procaspase-3, VEGF, MMP-2 and MMP-9 proteins, inhibited ERK1/2 but activated p38. These effects of calycosin were not observed in HUVEC. Neither AST nor AS IV had any significant effects on the parameters studied in HCT 116 cells. AST also showed no effect in HUVEC; AS IV, at 100 μM, appeared to increase the number of tube formation by HUVEC. In conclusion, the present findings suggest that AST has no significant effect on both cancer and endothelial cells while AS IV may promote angiogenesis without any direct action in colon cancer cells. In colon cancer cells, calycosin induces apoptosis, possibly through activation of caspase-3 and p38, and inhibits metastasis, possibly by downregulating MMP-2 and MMP-9, and inhibiting ERK1/2. However, in endothelial cells, the effect of calycosin is not conclusive as it promotes tube formation but inhibits migration. These findings provide the pharmacological basis for the use of Astragali Radix in the treatment of colon cancer, and the scientific evidence for a therapeutic potential of calycosin in the management of this disorder. Further studies are needed to verify the effect of calycosin on endothelial cells. In order to better mimic the clinical situation, the interaction between cancer and endothelial cells [for example, tumor-induced angiogenesis] needs to be taken into consideration. / published_or_final_version / Pharmacology and Pharmacy / Master / Master of Philosophy

The majority of sporadic colorectal cancer (CRC) cases affects individuals over the age of 50, but about 10% of cases occur in young adults under 50 in Hong Kong. Apart from germline mutation of the DNA mismatch repair genes that predisposes to early-onset CRC with a high-level of microsatellite instability (MSI-H), it is unknown if the mechanisms that give rise to CRC in other young adults differ from those in older individuals. In an effort to understand the genetic and epigenetic basis of early and late-onset CRC outside the Lynch Syndrome setting, we performed a detailed characterization of 36 MSI-H and 198 non-MSI-H tumours from patients of varying ages. This characterization was based primarily on the presence of the CpG island methylator phenotype (CIMP), as measured by the level of DNA methylation; and presence of genetic instability, as measured by DNA copy number aberrations, as well as mutations in BRAF, KRAS, or TP53. Our findings revealed that early (≤50) and late-onset (>50) CRCs have different genetic and epigenetic features. In non-MSI-H cancers, CIMP-H was associated with early-onset, while CIMP-L and KRAS mutation was associated with late-onset. However, in MSI-H tumours, late-onset disease was associated with CIMP-H and BRAF mutation. In addition, promoter methylation of MLH1 in early-onset MSI-H patients had a higher frequency of occurring in the germline that was locus specific, whereas nearly all late-onset MSI-H patients showed somatic regional methylation at the MLH1 locus, as well as regional methylation on other chromosomes. This is the first study to show regional methylation at chromosome 9p21 and 7p14, which encompass the CIMP markers P16 and ELMO1, respectively. We also observed an association between regional methylation and CIMP-H, but in MSI-H cases it was linked with late-onset, whereas in non-MSI-H cases it was irrespective of age. This suggests that mechanisms of methylation seeding and spreading may be different in early and late-onset disease. Moreover, CIMP-H non-MSI-H cases had significantly worse prognosis (p=0.021 for overall survival, p=0.004 for disease-free survival) and poor response to chemotherapy compared to CIMP-L or CIMP-Neg cases. Lastly, a methylation score assigned to non-MSI-H patients based on the degree of methylation of known CIMP markers was a significant prognostic factor of disease-free survival (p=0.004), and patients with a high methylation score showed a poor response to chemotherapy. Thus, our results suggest that different genetic and epigenetic mechanisms may drive tumourigenesis in early and late-onset disease. Although further research will be needed to elucidate the exact nature of these mechanisms, our findings should help to improve current classification of CRC patients with a goal towards personalized treatment. / published_or_final_version / Pathology / Doctoral / Doctor of Philosophy

Substantial studies have investigated homogeneity of psychological distress level among cancer patients by using cross-sectional and longitudinal study design. Nonetheless, as proposed by Bonnano (2004), heterogeneity characteristics of psychological distress following stressful event could not be neglected and he further suggested that the majority of individuals were resilient in response to stressful events. To test this postulation, recent studies employed growth mixture modelling method to examine the heterogeneity characteristics of psychological distress trajectory among cancer patients. Furthermore, identifying relevant factors differentiate the psychological distress trajectories is an integral part for developing effective interventions for cancer patients in dealing with illness demands. However, only a few studies have examined these issues among Chinese colorectal cancer patients, a second most common cancer in Hong Kong. Therefore, it is of important need to address this knowledge gap. This study had two major aims: 1) to explore the patterns of psychological distress among Chinese patients with colorectal cancer from shortly after diagnosis but before surgery (i.e. 1-day prior operation) to 1-year post-surgery and to testify Bonnano’s theory on resilience; 2) to identify the effects of cancer-related intrusive thoughts, physical symptom intrusiveness and dispositional optimism on differentiating psychological distress trajectories. A total of 246 Chinese patients with colorectal cancer were recruited for the current study. Altogether, 5 consecutive face-to-face interviews were conducted on one day prior to surgery (baseline), 1-, 4-, 8- and 12-month post-surgery (T2-T5). Patients’ psychological distress (i.e. anxiety and depression), physical symptom intrusiveness, cancer-related intrusive thoughts, dispositional optimism, demographic and medical information were assessed by a standardised questionnaire with valid and reliable psychometric instruments. Growth mixture modelling was used to estimate and specify the psychological distress trajectories. Multinomial logistic regression was adopted to assess the proposed factors in relation to differentiate the trajectory patterns. Growth mixture modelling suggested three distinct trajectories were identified for both anxiety and depression model. The majority of patients with colorectal cancer were identified as resilient (i.e. maintaining low and stable distress level across time) for both models (anxiety: 82.3%, depression: 82.7%). Additionally, for anxiety trajectory model, the remaining 12.3% and 5.4% of patients were classified as moderately-low anxiety group (i.e. maintaining moderate to low distress level) and increasing anxiety group (i.e. increased from moderate level of distress at initial to subsequently high distress level) respectively. For depression trajectory model, the remaining 12.6% and 4.7% of the patients were grouped as delayed depression (i.e. delayed level of distress over time) and recovery depression (i.e. recovered from high distress level to low across time). Multinomial logistic regression showed that cancer-related negative intrusive thoughts, physical symptom intrusiveness and dispositional optimism were significant factors to differentiate anxiety and depression trajectories respectively. This study highlighted the heterogeneous feature of psychological distress among Chinese patients with colorectal cancer. Physical symptom intrusiveness, cancer-related negative intrusive thoughts and dispositional optimism played important role on predicting cancer patient’s psychological distress respectively. Nonetheless, further investigations are much needed to clarify the underlying mechanism. / published_or_final_version / Public Health / Master / Master of Philosophy

Cancer stem cells are a subpopulation of cells needed for cancer initiation and progression. Previous works have revealed CD26-expressing colorectal cancer (CRC) stem cells are not only endowed with tumor-initiating properties, but also capable of conferring metastasis. However, whether the CD26 molecule plays role in metastasis and the underlying mechanism by which CD26 may mediate metastasis remain unclear. This study aims to reveal the biology and the molecular characteristics of the CD26-expressing CRC stem cells. Here, by the gene manipulation experiment, we showed that CD26 molecule is a functional marker that confers metastasis as transient and stable knock-down of the CD26 molecule in the CRC stem cells resulted in reduced wound healing, migration and invasion abilities in vitro and the capability to generate metastatic liver nodules in vivo, respectively. With the use of genome-wide expression array and immuno-blotting analysis, Smad-dependent TGF-β signaling, orchestrated by the SMAD2, SMAD3 and SMAD4 molecules, was up-regulated and activated in the CD26 expressing colorectal CSCs. In addition, expressions of the SMAD2 and SMAD3 molecules were found to be positively correlated with the CD26 molecule in clinical samples by qPCR and immunohistochemistry studies. Furthermore, no metastasis through EMT could be achieved once the Smad-dependent TGF-β signaling was down-regulated in the CD26 expressing CRC stem cells, which suggested that Smad-dependent TGF-β signaling was necessary for CD26-expressing CRC stem cells to induce metastasis. Finally, our result showed that the Smad-dependent TGF-β signaling was regulated by the CD26 molecule possibly through the down-regulation of CAV1 protein. To conclude, our findings have not only revealed the functional role of CD26 molecule, but have also unveiled a linkage between the CD26 molecule and Smad-dependent TGF-β signaling. Further study of this connection may introduce a novel mechanism, through which CRC metastasis can be induced by this functional CD26 marker of CRC stem cells. / published_or_final_version / Surgery / Doctoral / Doctor of Philosophy

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published_or_final_version / abstract / toc / Medicine / Master / Doctor of Medicine

published_or_final_version / abstract / toc / Anatomy / Master / Master of Philosophy

published_or_final_version / abstract / toc / Pharmacology / Doctoral / Doctor of Philosophy

published_or_final_version / Anatomy / Master / Master of Philosophy