No description available.

Mature T-cell leukemias represent rare, but increasingly recognized diseases of which, compared to their B-cell counterparts, comparatively little is established on their pathogenesis, diagnosis, and treatment. These leukemic post-thymic T-cell neoplasms range from the spectrum of chronic, sometimes debilitating disorders such as T-large granular lymphocytic leukemia (T-LGLL), and related leukemias such as NKLGLL, to more aggressive malignancies such as T- prolymphocytic leukemia (T-PLL). In this series, entitled ‘Pathogenesis, Treatment, and Future Directions for Rare T-cell Leukemias’ we review the current state of the science of these important T-cell neoplasms to inform on their treatment, diagnosis, and pathophysiology.

京都大学 / 新制・課程博士 / 博士(医学) / 甲第23803号 / 医博第4849号 / 新制||医||1058(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 村川 泰裕, 教授 滝田 順子, 教授 小川 誠司 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

Acute myeloid leukemia (AML) is a highly lethal malignancy characterized by unchecked expansion of immature myeloid blasts. While certain genetic and cytogenetic aberrations have been associated with chemotherapy response and disease risk, clinical outcomes remain heterogeneous. AML harbors relatively few somatic mutations compared to other cancers, however, it shows marked enrichment for epigenetic regulator alterations, and has been shown to harbor DNA methylation defects. My focus has been to dissect these epigenetic defects using high-throughput DNA methylation data. I first characterized two genome-wide hypermethylation signatures in AML: AML-CpG island methylator phenotype (A-CIMP+), and IDH-associated CIMP (I-CIMP+). While I-CIMP+ leukemias showed significant enrichments for mutations in IDH1 or IDH2, A-CIMP+ cases were mutation independent, and were best defined by their epigenetic defects, and associated transcriptomic changes. Importantly, A-CIMP+ leukemias had relatively favorable clinical outcomes, while I-CIMP+ patients did not. I next sought to characterize epigenetic defects involving demethylation of normally methylated genomic regions. I identified two distinct demethylator phenotypes (DMPs): DMP.1+ and DMP.2+. DMP.1+ AML was largely defined by mutations in DNMT3A, FLT3, and NPM1, while DMP.2+ leukemias harbored favorable-risk genomic rearrangements and a distinct gene expression profile. Both DMPs also carried prognostic information in AML; DMP.1+ cases had poor outcomes, while DMP.2+ patients tended to have favorable survival. Using both CIMP and DMP signatures, I then built an integrated epigenetic model for AML prognosis I termed MethylScore. The MethylScore algorithm was prognostic independent of age and cytogenetic risk in multivariate Cox regression models, suggesting that DNA methylation defects may augment existing clinical tools for risk stratification, and/or treatment selection. Finally, I explored whether DNA methylation signatures and genetic mutations could serve as biomarkers of response to epigenetic therapy, and found that DNA hypermethylation correlated with poor overall survival, and a gene mutation profile was associated with lack of complete remission after treatment with a DNA methylation inhibitor. These data provide evidence of distinct epigenetic signatures in AML that define transcriptionally, genetically, and clinically distinct populations that should be evaluated in future translational/clinical studies. / Biomedical Sciences

The identification and understanding of early drivers in malignancy is crucial to revert preleukemic events and prevent leukemic relapse. Del(20q) is one of the most common primary cytogenetic abnormalities found in preleukemic malignancies from myeloproliferative neoplasms to myelodysplastic syndrome (MDS). Previous studies have identified a “common retained region” within 20q11.21 that is often amplified in a subset of MDS patients. PLAGL2 is one of the 4 genes identified to be within the minimally conserved amplified region. Indeed, in previously published datasets of MDS hematopoietic stem and progenitor cells (HSPCs) transcriptome, PLAGL2 is significantly elevated in del(20q) patients compared to healthy controls. However, we have found that its level is also higher in HSPCs of cytogenetically normal MDS patients with low blasts. Given these findings, we sought to define the role of PLAGL2 as a potential early driver of myeloid malignancies. Results In healthy cord blood (CB) HSPCs, PLAGL2 overexpression enhanced proliferation ex vivo, better maintained stemness and decreased apoptosis. Colony formation assays also identified increased output of the erythroid lineage. Xenotransplanted CB CD34+ HSPCs overexpressing PLAGL2 exhibited increased engraftment competitiveness and led to splenomegaly with signs of hypercellularity after 20 weeks, features consistent with clinical observations of hematological malignancy. Grafts derived from PLAGL2 overexpressing cells reproducibly maintained a significantly larger CD34+ HSPC compartment. Intriguingly we also identified that ~50% of PLAGL2-overexpressing grafts exhibited a significant erythroid (CD71+/CD235a+) component where none was observed in the control group. This unique finding of aberrant erythropoiesis is reminiscent of clinical observations in patients with 20q11.21 amplification, where a high proportion of erythroblasts in the marrow and in some cases progression to erythroleukemia was noted. To evaluate the progression of PLAGL2-overexpressing grafts, further secondary transplantations were carried out and showed the persistence of only immature erythroid progenitors (CD71+/CD235a-) coupled with a near complete absence of lymphopoiesis in the same grafts. Together, our data strongly suggests ectopic levels of PLAGL2 can independently drive the expansion of human HSPCs and enforce features of myeloid malignancy. To uncover the molecular mechanism underlying PLAGL2 function, we performed RNA-seq and CUT&RUN in human CB CD34+ HSPCs overexpressing PLAGL2. Gene set enrichment analysis of the transcriptome and over-representation analysis of bound genes both identified signatures consistent with LSCs. We compared these findings with identically-derived omics profiles of HSPCs overexpressing PLAG1, a closely related family member that our lab has identified as a potent expander of HSCs ex vivo but not capable of promoting malignant features. We found a strong common feature in the downregulation of ribosomal components and translation machinery, then functionally validated reduced protein synthesis in PLAGL2 overexpressing HSPCs through OP-Puro assays. We have shown dampened mRNA translation to be one of the mechanisms by which PLAG1 enhances stemness and survival of HSCs, one that potentially extends to PLAGL2 as well. However, we also identified discordant signatures, notably PLAGL2's unique capacity to reduce mitochondrial translation, a pathway associated with ineffective erythropoiesis and MDS and one potential pathway by which PLAGL2 can enforce malignant phenotypes. Finally, to investigate the potential of PLAGL2 as a therapeutic target in MDS and AML, we performed shRNA knockdown in MDSL, a human MDS cell line, and primary human AML. In vitro competitive assays with MDSL showed steady dropout of PLAGL2 depleted cells. Similarly, depletion of PLAGL2 in primary AML was also able to attenuate colony formation and engraftment in vivo, highlighting the therapeutic potential of PLAGL2 inhibition throughout myeloid malignancies. Conclusion We have identified PLAGL2's potential as an early independent driver of myeloid malignancy and aberrant erythroid differentiation. An understanding of PLAGL2 and its downstream mechanisms will not only further our understanding on the development of early myeloid malignancies but also potentially provide another avenue to treat or prevent leukemia before it manifests. / Thesis / Doctor of Philosophy (PhD)

No description available.

The improving prognosis for pediatric leukemia patients requires that involved professionals increase attention to the emotional adjustment of these children. This study was designed to determine (a) how mothers of leukemia patients perceived their children's personalities in order to identify any specific emotional difficulties which these children may experience and (b) if their perceptions differed from either mothers of cystic fibrosis and diabetes patients or mothers of healthy children. Subjects included 24 mothers in each of three groups: leukemia, other illness, and healthy. Children in both illness groups received higher scores than healthy children on Adjustment, Achievement, Somatic Concern, Depression, Psychosis, and Social Skills scales as measured by the Personality Inventory for Children; however, only the leukemic children were rated higher in areas of Anxiety and Withdrawal. Implications for treatment and future research are discussed.

Ng, Ka Yan. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2010. / Includes bibliographical references (leaves 118-131). / Abstracts in English and Chinese. / Thesis Abstract --- p.i / 論文摘要 --- p.vi / Acknowledgements --- p.x / Abbreviations --- p.xii / Thesis Content --- p.xv / List of Figures --- p.xix / List of Tables --- p.xxi / Chapter Chapter 1 --- Introduction --- p.1 / Chapter 1.1 --- Haematopoiesis --- p.1 / Chapter 1.2 --- Leukemia --- p.2 / Chapter 1.3 --- Childhood B-cell precursor acute lymphoblastic leukemia (BCP-ALL) --- p.3 / Chapter 1.3.1 --- Epidemiology --- p.4 / Chapter 1.3.2 --- Causes and risk factors --- p.4 / Chapter 1.3.3 --- Clinical features --- p.6 / Chapter 1.3.4 --- Morphology --- p.6 / Chapter 1.4 --- Classification of BCP-ALL --- p.7 / Chapter 1.4.1 --- Immunophenotyping --- p.7 / Chapter 1.4.2 --- Cytogenetics and molecular genetics --- p.9 / Chapter 1.5 --- Prognostic factors --- p.13 / Chapter 1.6 --- Current treatments of BCP-ALL --- p.15 / Chapter Chapter 2 --- Literature Review --- p.18 / Chapter 2.1 --- Cytogenetics abnormalities in BCP-ALL --- p.18 / Chapter 2.1.1 --- Chromosomal translocation --- p.18 / Chapter 2.1.2 --- Aneuploidy --- p.21 / Chapter 2.2 --- Epigenetic aberrations --- p.21 / Chapter 2.2.1 --- DNA methylation --- p.22 / Chapter 2.2.2 --- Mechanism of DNA Methylation in Transcription Repression --- p.23 / Chapter 2.3 --- DNA Methylation in Normal Haematopoiesis --- p.25 / Chapter 2.4 --- DNA Methylation in Haematological Malignancies --- p.26 / Chapter 2.4.1 --- DNA methylation in ALL --- p.26 / Chapter 2.4.2 --- DNA methylation in BCP-ALL --- p.29 / Chapter 2.5 --- Angiogenesis in pathogenesis of acute leukemias --- p.30 / Chapter 2.6 --- Thrombospondin-1 (TSP-1) --- p.32 / Chapter 2.6.1 --- Structure of TSP-1 --- p.33 / Chapter 2.6.2 --- The role of TSP-1 in tumorigenesis --- p.34 / Chapter 2.6.3 --- TSP-1 mediates the activation of TGFβ --- p.36 / Chapter 2.6.4 --- TSP-1 mediates TGFβ-induced Apoptosis --- p.37 / Chapter 2.6.5 --- Association of TGFβ with normal haematopoiesis and haematological malignancies progression --- p.40 / Chapter 2.6.6 --- TSP-1 Induced Apoptosis via its Receptor CD36 --- p.42 / Chapter 2.6.7 --- THBS1 promoter hypermethylation and its association with tumorigenesis --- p.43 / Chapter 2.6.8 --- Effect of THBS1 aberrant methylation on TGFp --- p.45 / Chapter Chapter 3 --- Rationale of Study --- p.47 / Chapter Chapter 4 --- Materials and Methods --- p.52 / Chapter 4.1 --- Patient sample --- p.52 / Chapter 4.2 --- Cell lines --- p.52 / Chapter 4.3 --- Mononuclear cells isolation --- p.53 / Chapter 4.4 --- THBS1 promoter hypermethylation analysis --- p.54 / Chapter 4.4.1 --- DNA extraction from mononuclear cells and cell lines --- p.54 / Chapter 4.4.2 --- Bisulfite conversion --- p.55 / Chapter 4.4.3 --- Methylation specific PCR (MSP) --- p.55 / Chapter 4.5 --- Quantification of THBS1 mRNA expression --- p.57 / Chapter 4.5.1 --- RNA extraction --- p.57 / Chapter 4.5.2 --- Reverse transcription PCR --- p.58 / Chapter 4.5.3 --- Real-time RT-PCR --- p.58 / Chapter 4.6 --- Determination of plasma TSP-1 level --- p.59 / Chapter 4.7 --- TSP-1 treatment --- p.60 / Chapter 4.8 --- Flow cytometry analysis --- p.60 / Chapter 4.8.1 --- Annexin-V analysis --- p.60 / Chapter 4.8.2 --- Cell fixation --- p.61 / Chapter 4.8.3 --- Analysis of Caspase-3 activation --- p.62 / Chapter 4.8.4 --- "Analysis of TGFβ downstream pathway activation: Phosphorylation of Smad2/3, JNK and p38" --- p.62 / Chapter 4.9 --- Determination ofTGF-β expression --- p.63 / Chapter 4.10 --- Statistical analysis methods --- p.64 / Chapter Chapter 5 --- Results / Chapter 5.1 --- THBS1 methylation statuses in BCP-ALL patients and cell lines --- p.66 / Chapter 5.2 --- Correlation of THBS1 methylation statuses and clinico- pathological features in BCP-ALL patients --- p.68 / Chapter 5.3 --- Association of THBS1 methylation and THBS1 mRNA expression --- p.69 / Chapter 5.4 --- Effect of TSP-1 treatment on apoptosis level of BCP-ALL cells --- p.72 / Chapter 5.4.1 --- Annexin-V assay --- p.72 / Chapter 5.4.2 --- Caspase-3 activation assay --- p.75 / Chapter 5.5 --- THBS1 methylation and activation of secreted TGFβ --- p.78 / Chapter 5.6 --- Effect of TSP-1 treatment on activation of TGFβ --- p.80 / Chapter 5.7 --- The involvement ofTGFβ activation in TSP-1 induced apoptosis in BCP-ALL --- p.82 / Chapter 5.8 --- The association of TGFβ signaling pathway activities with THBS1 methylation --- p.86 / Chapter Chapter 6 --- Discussion --- p.91 / Chapter 6.1 --- THBS-1 promoter hypermethylation in BCP-ALL cell lines and patients: Correlation with expression and clinico-pathological profile --- p.93 / Chapter 6.1.1 --- THBS1 promoter hypermethylation status in childhood BCP-ALL --- p.93 / Chapter 6.1.2 --- THBS1 methylation as prognostic markers --- p.94 / Chapter 6.1.3 --- "Association of THBS1 methylation status, mRNA expression and TSP-1 protein expression in childhood BCP-ALL" --- p.96 / Chapter 6.2 --- Study of the correlation of TSP-1 induced apoptosis with the THBS1 promoter methylation status --- p.99 / Chapter 6.3 --- Elucidation of the molecular mechanisms of TSP-1 induced apoptosis: study of the involvement of TGFβ activation --- p.103 / Chapter 6.3.1 --- Latent TGFβ activation by TSP-1 in BCP-ALL and association with THBS1 methylation status --- p.103 / Chapter 6.3.2 --- TSP-1 induced cell death through activation ofTGFβ --- p.105 / Chapter 6.3.3 --- TSP-1 induced apoptotic signals via TGFβ signaling pathway --- p.107 / Chapter 6.4 --- Limitation of study --- p.113 / Chapter 6.5 --- Future studies --- p.114 / Chapter 6.5.1 --- Continuation study in TSP-1 induced TGFβ-mediated pathways --- p.114 / Chapter 6.5.2 --- Microarray analysis --- p.115 / Chapter 6.6 --- TSP-1 in treatment of childhood BCP-ALL --- p.115 / Chapter Chapter 7 --- Conclusion --- p.117 / Reference --- p.118

Chung, Po Yin. / Thesis submitted in: December 2006. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2007. / Includes bibliographical references (leaves 128-155). / Abstracts in English and Chinese. / Thesis Abstract --- p.i / 論文摘要 --- p.iv / Acknowledgements --- p.vi / Abbreviations --- p.vii / Thesis Content --- p.xi / List of Figures --- p.xv / List of Tables --- p.xvii / Chapter Chapter 1 --- Introduction --- p.1 / Chapter 1.1. --- Normal Hematopoiesis --- p.1 / Chapter 1.2. --- Hematological Malignancy and the Aberrant Development of Blood Cells --- p.2 / Chapter 1.3. --- Leukemia and Its Classification --- p.3 / Chapter 1.4. --- Childhood Acute Lymphoblastic Leukemia (ALL) --- p.5 / Chapter 1.4.1. --- Epidem iology --- p.5 / Chapter 1.4.2. --- Causes and Risk Factors --- p.6 / Chapter 1.4.3. --- Molecular Pathophysiology --- p.7 / Chapter 1.4.4. --- Clinical Presentation --- p.9 / Chapter 1.4.5. --- Classification --- p.10 / Chapter 1.4.5.1. --- Immunophenotyping --- p.10 / Chapter 1.4.5.2. --- French-American-British (FAB) Classification --- p.12 / Chapter 1.4.6. --- Diagnosis and Prognosis --- p.14 / Chapter 1.4.6.1. --- Morphological and Cytochemical Analysis --- p.15 / Chapter 1.4.6.2. --- Cytogenetic and Molecular Genetic Characterizations --- p.16 / Chapter 1.4.7. --- Treatment --- p.19 / Chapter 1.5. --- Overview of Epigenetics --- p.21 / Chapter 1.6. --- Concepts ofDNA Methylation --- p.23 / Chapter 1.6.1. --- CpG Islands --- p.23 / Chapter 1.6.2 --- Mechanisms of DNA Methylation --- p.24 / Chapter 1.6.3 --- Physiological Roles of DNA Methylation --- p.28 / Chapter 1.6.4 --- Initiation of Aberrant DNA Methylation --- p.30 / Chapter 1.7. --- DNA Methylation in Tumorigenesis --- p.31 / Chapter 1.7.1. --- Regional Hypermethylation --- p.33 / Chapter 1.7.2 --- Global and Regional Hypomethylation --- p.34 / Chapter 1.7.3 --- Microatellite Instability and Oncogeneic Mutation --- p.35 / Chapter Chapter 2 --- Literature Review --- p.37 / Chapter 2.1. --- Aberrant DNA Methylation in Childhood ALL --- p.37 / Chapter 2.1.1. --- Cell Cycle --- p.39 / Chapter 2.1.2. --- Apoptosis --- p.41 / Chapter 2.1.3. --- Tissue Invasion and Metastasis --- p.42 / Chapter 2.1.4. --- Transcription Factors and Metabolic Enzymes --- p.44 / Chapter 2.1.5. --- Putative Tumor Suppressor Genes --- p.44 / Chapter 2.1.6. --- Chromosome Instability --- p.46 / Chapter 2.2. --- Methodologies in DNA Methylation Analysis --- p.50 / Chapter 2.2.1. --- Principle of Methylation-sensitive Arbitrarily Primed PCR (MS-AP PCR) --- p.50 / Chapter 2.2.2. --- Combined Bisulfite Restriction Analysis (COBRA) --- p.53 / Chapter 2.2.3. --- Cloned Bisulfite Sequencing --- p.55 / Chapter 2.2.4. --- Experimental Use of Demethylating Agents --- p.55 / Chapter Chapter 3 --- Background of Research --- p.58 / Chapter 3.1. --- Current Methylation Studies in Childhood ALL --- p.58 / Chapter 3.2. --- Objectives of Research --- p.60 / Chapter 3.3. --- Study Approach and Experimental Design --- p.61 / Chapter Chapter 4 --- Materials and Methods --- p.63 / Chapter 4.1. --- Clinical Samples and ALL Cell Lines --- p.63 / Chapter 4.1.1. --- Clinical Samples from Pediatric Patients with ALL and Normal Healthy Donors --- p.63 / Chapter 4.1.2. --- ALL Cell Lines --- p.63 / Chapter 4.2. --- Genomic DNA Isolation from Clinical Samples and Cell Lines --- p.64 / Chapter 4.2.1. --- Ficoll Gradient Centrifugation --- p.64 / Chapter 4.2.2. --- DNA Extraction --- p.64 / Chapter 4.3. --- MS-AP PCR --- p.65 / Chapter 4.3.1. --- Methylation-sensitive Restriction Enzyme Digestion of Genomic DNA --- p.65 / Chapter 4.3.2. --- Arbitrarily Primed Polymerase Chain Reaction --- p.66 / Chapter 4.3.3. --- Isolation of Differentially Methylated DNA Fragments --- p.69 / Chapter 4.4. --- Cloning of Differentially Methylated DNA Fragments --- p.70 / Chapter 4.4.1. --- TA Cloning --- p.70 / Chapter 4.4.2. --- Screening of Positive Clones --- p.71 / Chapter 4.4.3. --- Preparation of Plasmid DNA by Alkaline Lysis Method --- p.72 / Chapter 4.5. --- DNA Sequence Analysis of Differentially Methylated DNA Fragments --- p.72 / Chapter 4.5.1. --- Dye-terminator Cycle Sequencing --- p.72 / Chapter 4.5.2. --- CpG islands Analysis of Differentially Methylated Sequences --- p.73 / Chapter 4.6. --- DNA Methylation Analysis --- p.74 / Chapter 4.6.1. --- Sodium Bisulfite Modification --- p.74 / Chapter 4.6.2. --- Combined Bisulfite Restriction Analysis --- p.75 / Chapter 4.6.3. --- Cloned Bisulfite Genomic Sequencing --- p.76 / Chapter 4.7 --- Gene Expression Study --- p.76 / Chapter 4.7.1. --- RNA Extraction from Clinical Samples and ALL Cell Lines --- p.76 / Chapter 4.1.2. --- Reverse Transcription PCR --- p.77 / Chapter 4.7.3. --- Semi-quantitative RT-PCR --- p.78 / Chapter 4.7.4. --- 5-aza-2 '-deoxycytidine Demethylation Treatment --- p.79 / Chapter Chapter 5 --- Results --- p.80 / Chapter 5.1. --- Generation of DNA Methylation Pattern by MS-AP PCR --- p.80 / Chapter 5.1.1. --- Differential Methylation Patterns of Childhood ALL --- p.84 / Chapter 5.1.2. --- Methylation Patterns of B and T lineages Childhood ALL --- p.86 / Chapter 5.2. --- UCSC BLAT Analysis of Differential Methylated DNA Sequences / Chapter 5.3. --- Identification of Candidate Gene --- p.89 / Chapter 5.4. --- Fibrillin 2 --- p.90 / Chapter 5.4.1. --- FBN2 CpG Islands: UCSC BLAT Search Analysis --- p.90 / Chapter 5.4.2. --- Verification ofFBN2 by ALL Cell Lines --- p.91 / Chapter 5.4.2.1. --- Semi-quantitative RT-PCR --- p.91 / Chapter 5.4.2.2. --- COBRA --- p.92 / Chapter 5.4.2.3. --- Cloned Bisulfite Sequencing --- p.94 / Chapter 5.4.2.4. --- Demethylation Treatment Resorted FBN2 mRNA Expression in ALL Cell Lines --- p.98 / Chapter 5.4.3. --- Studies ofFBN2 in Childhood ALL --- p.99 / Chapter 5.4.3.1. --- Methylation Analysis --- p.99 / Chapter 5.4.3.2. --- Semi-quantitative RT-PCR --- p.105 / Chapter Chapter 6 --- Discussion --- p.107 / Chapter 6.1. --- Genome-wide Screening Approach: MS-AP PCR --- p.107 / Chapter 6.2. --- Sample Selection in this Study --- p.109 / Chapter 6.2.1. --- MS-AP PCR --- p.109 / Chapter 6.2.2. --- Methylation Studies --- p.109 / Chapter 6.2.3. --- Studies in ALL Cell Lines --- p.110 / Chapter 6.3. --- Methylation Patterns in Childhood ALL --- p.111 / Chapter 6.4. --- Candidate Genes Selection Strategies in MS-AP PCR --- p.112 / Chapter 6.5. --- Fibrillin 2: mRNA Expression and Methylation Studies --- p.113 / Chapter 6.5.1 --- ALL Cell Lines --- p.113 / Chapter 6.5.2 --- Childhood ALL --- p.113 / Chapter 6.5.2.1 --- mRNA Expression and Methylation Studies --- p.113 / Chapter 6.5.2.2 --- Statistical Analysis --- p.115 / Chapter 6.5.3. --- Possible Roles of FBN2 in Leukemogenesis --- p.116 / Chapter 6.6. --- Clinical Application of FBN2 Aberrant Methylation --- p.119 / Chapter 6.6.1. --- Tumor Markers --- p.119 / Chapter 6.6.2. --- Use of Demethylating Drugs in Chemotherapy --- p.121 / Chapter 6.7. --- Limitations of Methylation Studies --- p.122 / Chapter 6.7.1. --- MS-AP PCR --- p.122 / Chapter 6.7.2. --- Techniques Used in Methylation Study --- p.122 / Chapter 6.7.3. --- Problems in Methylation Study --- p.123 / Chapter 6.8. --- Future Studies --- p.125 / Chapter Chapter 7 --- Conclusion --- p.127 / References --- p.128 / Appendix --- p.155

In this study, a novel compound was isolated and purified from the solid culture medium (potato dextrose agar) of shiitake 1358 strain through series of methods, such as ethanol precipitation, macroporous resin column separation, semi-preparative high performance liquid chromatography separation and preparative thin-layer chromatography separation. Analyzing spectra from fourier transform infra-red spectroscopy, gas chromatography-mass spectrometry, 1-dimension and 2-dimension nuclear magnetic resonance, the chemical structure of the novel compound was determined and named as 4-amino-5,6-dihydrobenzo[d]oxonine-2,7(1H,4H)-dione. It could inhibit the proliferation of HL-60 leukemia cells significantly and with an IC50 of 1.56 mug/ml (7.123 mumol/L) in the 72-hour treatment. From the results, it is suggested that this compound could activate the G2 phase checkpoint control of the cell cycle to arrest the cell cycle in G2 phase. In addition, it could suppress the replicative DNA synthesis to inhibit the proliferation of HL-60 leukemia cells. The more important is that this compound can induce the apoptosis of HL-60 leukemia cells significantly through intrinsic and extrinsic apoptotic pathways. The compound could induce intrinsic and extrinsic apoptosis through the regulation of the apoptosis-related proteins, such as Fas ligand, Bax, Bcl-2, Caspase 8, Caspase 9, and Caspase 3. For intrinsic pathway, the compound might upregulate Bax, downregulated Bcl-2, activated the Caspase 9, subsequently activated Capase 3, and ultimately led to cell death. For extrinsic pathway, the compound upregulated the Fas ligand, cleaved and activated Procaspase 8 to active Caspase 8, further cleaved and activated Procaspase 3 to active Caspase 3 to commit the cells to apoptosis. / Leukemia is a malignant cancer that involves the bone marrow and blood circulation systems. Leukemia results in the uncontrolled growth of abnormal (leukemic) white blood cells and may also invade other organs, including the liver, spleen, lymph nodes, testes, and brain. In 2007, about 44,240 new cases of leukemia were diagnosed and 21,790 patients died from all types of leukemias in USA. / Shiitake was first cultivated in China more than 800 years ago. It is the second most commonly cultivated edible mushrooms in the world nowadays. For a long time, shiitake has been valued for its unique taste and flavor and as a medicinal invigorant. According to ancient Chinese medicinal theory, consumption of shiitake was in favor of long life and good health. In China and Japan, shiitake has been used as both a food and a medicinal herb for thousands of years. It is the source of several well-studied preparations with proven pharmacological properties, especially the polysaccharide lentinan. Currently, most researches concentrate on the anticancer activities of the extracts from the fruiting body of shiitake, especially polysaccharides. Report about the anti-cancer effects of other components from the shiitake mushroom is scarce. The objectives of this investigations were: (1) to study the anticancer activities of brownish substances obtained during the solid medium culture of shiitake on specific cancer cell unes, especially HL60 cancer cell line; (2) to isolate and characterize the active compound(s) in the brown mushroom exudates; and (3) to propose the possible mechanism of actions, especially the function of the bcl-2 family genes and proteins. / by Guo, Yuming. / Adviser: Chung Hale Yin. / Source: Dissertation Abstracts International, Volume: 70-06, Section: B, page: 3314. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2008. / Includes bibliographical references (leaves 188-199). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts in English and Chinese. / School code: 1307.