published_or_final_version / Pathology / Master / Master of Medical Sciences

published_or_final_version / Pathology / Master / Master of Medical Sciences

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

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

Errata pages inserted behind leaf 293. / Includes bibliography (leaves 255-293). / xv, 293, [60] leaves, [33] leaves of plates : ill. (some col.) ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Aims to determine the molecular mechanism by which expression of Fas is lost in colorectal tumours and investigates the effects of re-introducing Fas into colon cancer cells. An animal study was undertaken in addition to the studies of colorectal cancer, to investigate the role of Fas signalling in hormone-dependent involution of the prostate gland. / Thesis (Ph.D.)--University of Adelaide, Dept. of Surgery, 1998?

Errata pages inserted behind leaf 293. / Includes bibliography (leaves 255-293). / xv, 293, [60] leaves, [33] leaves of plates : ill. (some col.) ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Aims to determine the molecular mechanism by which expression of Fas is lost in colorectal tumours and investigates the effects of re-introducing Fas into colon cancer cells. An animal study was undertaken in addition to the studies of colorectal cancer, to investigate the role of Fas signalling in hormone-dependent involution of the prostate gland. / Thesis (Ph.D.)--University of Adelaide, Dept. of Surgery, 1998?

published_or_final_version / Pathology / Doctoral / Doctor of Philosophy

published_or_final_version / Pathology / Doctoral / Doctor of Philosophy

published_or_final_version / abstract / Pathology / Master / Master of Philosophy

Eukaryotic translation elongation factor-2 (EEF2) is regulated through phosphorylation by a specific kinase known as eukaryotic elongation factor-2 kinase (EEF2K), leading to translational down regulation. Currently, it has been reported that EEF2K could promote the autophagic survival in breast and glioblastoma cell lines. However, the precise function of EEF2K in cancer as well as the related mechanism is still poorly understood. Colorectal cancer is the third common malignant disease worldwide and more than half of the patients with colorectal cancer require chemotherapy after surgery. However, de novo or acquired resistance to the agents is common. Discovery of novel targets for the chemotherapeutic intervention or treatment of colorectal cancer is highly warranted. In this study, the role of EEF2K as well as the underlying mechanism involved was evaluated in HT-29 and HCT-116 human colon cancer cells. Contrary to the reported autophagy-promoting activity of EEF2K in certain cancer cells, EEF2K is shown to negatively regulate autophagy in colon cancer cells as indicated by the increase of LC3-II levels, the accumulation of LC3 dots per cell, and the promotion of autophagic flux in EEF2K silenced cells. Moreover, the silencing of EEF2K promotes the cell viability, clonogenicity, cell proliferation and cell size in colon cancer cells. The silencing of BECN1 and ATG7 significantly reduce silencing of EEF2K induced LC3-II accumulation and cell survival. However, autophagy induced by EEF2K silencing does not potentiate the anticancer efficacy of the AKT inhibitor MK-2206. In addition, EEF2K overexpression decreases the cell survival and potentiates the antitumor efficacy of oxaliplatin. Autophagy induced by silencing of EEF2K is attributed to induction of protein synthesis, which results in ATP consumption and then actives AMPK-ULK1 pathway. This process appears independent of the suppression of MTOR activity and ROS generation. Silencing of AMPK or ULK1 significantly decreases EEF2K silencing-induced autophagy as well as cell survival in colon cancer cells. In conclusion, EEF2K negatively regulates autophagic survival through the AMPK-ULK1 pathway in colon cancer cells. This study provide useful information in understanding the role of EEF2K in colon cancer cells and suggests that upregulation of EEF2K activity may be developed a novel approach for the treatment of human colon cancer. / 真核延伸因子2激酶 (EEF2K) 通過磷酸化修飾真核延伸因子2 (EEF2) 來調控其活性，進而下調蛋白質翻譯延伸的速度。目前，有研究表明在乳腺癌和多形性膠質母細胞瘤中，EEF2K能夠誘導細胞自噬，並且這種類型的細胞自噬有助於細胞生存。然而，對於EEF2K在腫瘤中的精確作用以及它所涉及的分子機理仍然知之甚少，有待於進一步的研究。結直腸癌是全球第三大惡性腫瘤疾病，約有半數以上的患者需要手術後進行化學藥物治療。然而，患者對目前已有藥物的耐藥性十分普遍，因此，研發新的化學藥物靶點或者新的治療方法十分必要。在本課題研究中，EEF2K的功能及其所涉及的分子機理在人結腸癌細胞系HT-29和HCT-116上進行了闡釋。與在某些特定種類腫瘤細胞中EEF2K能夠誘導細胞自噬產生的現象相反，在EEF2K表達下調的人結腸癌細胞中，細胞自噬標記物LC3-II表達上升， 細胞中LC3斑點的聚集增多，並且細胞自噬流增強的現象，都表明EEF2K在這類腫瘤細胞中負調控細胞自噬。在結腸癌細胞中，EEF2K表達下調能夠增強細胞的活力，單細胞克隆的形成，細胞增殖以及細胞大小。此外，沈默BECN1和ATG7基因的表達都能夠減少EEF2K下調引發的LC3-II積累以及細胞增殖。然而，降低EEF2K表達所引發的細胞自噬並不能夠增強AKT抑制劑MK-2206抗腫瘤的效果。EEF2K的過表達能夠減少細胞增殖並且加強oxaliplatin的抗腫瘤藥效。沈默EEF2K引發的細胞自噬是通過誘導蛋白質的合成，導致ATP的消耗進而激活AMPK-ULK1細胞通路，與MTOR活性的抑制及ROS的產生無關。在結腸癌細胞中，降低AMPK或者ULK1的表達能夠消除EEF2K沈默所引起的LC3-II表達升高，細胞中LC3斑點聚集增多以及細胞增殖加強等現象。綜上所述，在人結腸癌細胞中，沈默EEF2K基因表達能夠通過激活AMPK-ULK1細胞通路，誘導有助於細胞存活的自噬現象產生。本課題研究對理解EEF2K在結腸癌細胞中的功能提供了有用的信息並且表明增強EEF2K的活性可以作為一種潛在的新的治療人結腸癌的方法。 / Liu, Xiaoyu. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2014. / Includes bibliographical references (leaves 116-131). / Abstracts also in Chinese. / Title from PDF title page (viewed on 16, November, 2016). / Detailed summary in vernacular field only.