2011-08-012024-05-13https://scholars.lib.ntu.edu.tw/handle/123456789/645394摘要：背景：骨髓化生不良症侯群(簡稱MDS)的特徵為周邊血常表現出全血球減少，造血細胞呈現化生不良(dysplasia)及血球分化異常的現象。部分病人的疾病會轉化為急性骨髓性白血病(AML)，因此過去又稱做白血病前期疾病(preleukemia)。MDS根據法美英(FAB)或世界衛生組織(WHO)的國際分類，可細分為數個亞型，臨床上則大致將其劃分為低危險及高危險兩群。目前對MDS的病因學及其轉化為AML的機轉尚不清楚，一般認為可能是造血細胞的基因或表觀基因體(epigenetics)異常所造成。我們過去的研究發現MDS病人會有染色體異常，N-RAS、K-RAS、AML1/RUNX1等基因突變，及抑癌基因p15、SOCS1的甲基化，但許多問題還有待解決。DNMT3A為DNA methyltransferase (DNMT)，可以造成DNA的甲基化，具有修飾表觀基因的作用。最近研究發現有相當高比例的AML病人，其白血病細胞內有DNMT3A基因的突變，且為一個獨立會影響病人預後的危險因子。這個突變在MDS尚未有正式的論文發表，僅在今年12月的美國血液學會年會中有一篇摘要，顯示8%的MDS病人有DNMT3A的突變；由於病人數不多，無法探討DNMT3A突變是否為獨立的預後因子，這個突變與染色體異常及其他基因突變間的交互作用亦完全未提。除了DNMT3A外，最近有幾個與表觀基因修飾有關的基因突變陸續在MDS被發現，如TET2, EZH2及ASXL1等；TET2被認為會使DNA demethylation；EZH2為histone H3K27 methyltransferase上的catalytic subunit；ASXL1被認為可以調控histone methylation；另外IDH1及IDH2的突變會影響TET2的功能，也發現會間接造成DNA methylation的增加。由於表觀基因體的異常，如DNA甲基化，Histone的修飾，在MDS扮演很重要的角色，且由我們過去的研究發現東西方的MDS有許多差異，因此在此計畫中，我們將探討此地DNMT3A基因突變在MDS發生及進展中的角色，它與TET2, EZH2, ASXL1, IDH1, IDH2突變及染色體與其他基因異常（如RAS, RUNX1, FLT3等）的關聯性，及這些基因突變對病人預後單獨的影響或幾個突變同時存在時的影響。在系列的追踪檢查中，也可以知道基因突變在疾病進展時的角色。研究方法：我們將對約300個MDS病人在診斷時及後續的追踪過程中，檢測DNMT3A及其他基因包括TET2, EZH2, ASXL1, IDH1, IDH2, RUNX1, RAS, FLT3等的突變，探討DNMT3A突變與臨床表現及病人預後間的關聯性，與其他基因的交互作用及影響性。在體外，將對MDS細胞株調控正常或突變種DNMT3A的表現，對於會經常同時出現在同一個病人的兩個或數個基因的異常，也將檢測這些基因突變同時在細胞株表現時對細胞生長及藥物反應的影響。預期成果：這項研究將有助於了解DNMT3A及其他與表觀基因調控相關之基因的突變在MDS所扮演的角色及臨床重要性。由所得結果可以將MDS病人做更好的risk-stratification，也有助於日後以表觀基因異常做為標靶治療的參考。<br> Abstract: Background：Myelodysplastic syndrome (MDS) is a clonal disorder characterized by cytopenia in peripheral blood (PB), and dysplastic changes and impairment of differentiation in hematopoietic cells. In some patients, the disease may transform to acute myeloid leukemia (AML); that is why it was called preleukemia in the past. MDS can be classified into several subtypes, according to French-America-British (FAB) or World Health Organization (WHO) criteria. Clinically, MDS patients are usually separated into high-risk and low-risk groups. The pathogenesis of MDS and its progression to AML remain unclear. It is suggested that MDS may be arisen from genetic or epigenetic aberrations of hematopoietics cells. We have found some recurrent chromosomal abnormalities, gene mutations, such as N-RAS, K-RAS and AML1/RUNX1, and aberrant methylation of suppressor genes, such as p15 and SOCS1, in MDS. However, a lot of questions remain to be answered. DNMT3A, a DNA methyltransferase, transfers methyl groups onto CpG dinucleotides leading to DNA methylation. Recently, DNMT3A mutation was detected in AML patients and was found to be an independent factor for poor prognosis. There has been no report concerning DNMT3A mutation in MDS in literature till now. Only one abstract was presented in the Annual Meeting of American Society of Hematology in Dec 2010, in which 8% of MDS patients were found to have DNMT3A mutation. Because the number of patients studied is limited, it is not known whether DNMT3A mutation is an independent prognostic factor. The interaction of DNMT3A mutation with cytogenetic abnormalities and other gene mutations remains unclear, too. In addition to DNMT3A, mutations of other genes, including TET2, EZH2 and ASXL1 which are associated with epigenetic modifications, were reported in MDS. TET2 plays a role in DNA demethylation; EZH2 encodes the catalytic subunit of polycomb repressive complex2 (PRC2), a highly conserved histone H3K27 methyltransferase; and ASXL1 is suggested to involve the regulation of histone methylation. Furthermore, mutations of IDH1 and IDH2 result in hypermethylation phenotype by disruption of TET2 enzyme activity. Since abnormalities in epigenetic modifications, including DNA methylation, histone acetylation and methylation, play important roles in the pathogenesis of MDS, and MDS in Taiwan is quite different form that in the West, we will explore the roles of DNMT3A mutation in the development and progression of MDS in this area, its interaction with other genetic alterations, especially those related to epigenetic deregulation, and the prognostic implication, either alone or in combination with other gene mutations.Methods：Mutations of DNMT3A and other genes, including TET2, EZH2, ASXL1, IDH1, IDH2, RUNX1, RAS and FLT3 etc., will be performed in about 300 MDS patients at diagnosis and during subsequent follow-up. The interaction of these gene mutations and clinical implications will be explored. In vitro manipulation of MDS cell lines by gene transfection or gene knock-down will be performed to find out the effect of DNMT3A mutation alone or in combination with other gene mutations in cell growth and response to drugs.Prospective results：From this study, the roles of DNMT3A mutation and other gene mutations related to epigenetic deregulation and their clinical implications in MDS will be known. The results may also help develop treatment to reverse aberrant epigenetic modification.骨髓化生不良症候群DNMT3A基因突變Myelodysplastic SyndromeDNMT3A mutation