2010-07-012024-05-13https://scholars.lib.ntu.edu.tw/handle/123456789/645954摘要：背景有多種遺傳變異被發現並證明在急性骨髓性白血病的癌化過程扮演重要角色。這些變異，依照其對血液細胞所造成的影響，可區分為兩類:一類會造成細胞出現不受控制的存活或分裂，另一類會阻斷血液細胞正常的分化。目前認為血液細胞同時帶有兩類變異時，就會癌化為急性骨髓性白血病。然而，這些遺傳變異間的交互作用與它們造成血液細胞癌化的詳細機制仍然未知。近年有報告發現兩個孤兒受體Nr4A1 及Nr4A3 與急性骨髓性白血病的癌化相關，在動物實驗中剔除此兩個基因會導致急性骨髓性白血病的發生。此兩個孤兒受體的功能過去並不完全清楚，僅有部分研究指出Nr4A1 會促進細胞凋亡。若將此發現連結至前述雙重變異導致白血病癌化的理論上，Nr4A1 下降會減少血液細胞凋亡而增加存活及分裂的能力，因此Nr4A3 表現量下降在血液細胞癌化的過程中可能扮演阻斷血液細胞分化的角色。初步探索在我們實驗室的初步研究中發現，利用化學物質引導k562 細胞株分化的同時，Nr4A3 的表現亦有明顯上升，另外臨床上使用全反式A 酸治療急性前骨髓性白血病病患誘導其白血病芽細胞分化的同時，Nr4A3 的表現亦隨之上揚。這些觀察與我們的假說相符合。實驗假說1 Nr4A3 表現下降影響血液細胞的分化而促進血液細胞的癌化2 Nr4A3 表現的程度對病人有臨床意義及特徵3 提高Nr4A3 的表現，可以加強藥物治療骨髓性白血病的效果方法1 利用化學藥物誘導細胞株分化的模型，分析在細胞分化的過程中Nr4A3 表現量的變化，是否跟分化現象相關。同時利用siRNA 阻斷Nr4A3 表現，觀察分化的誘導是否因此受到影響。2 採用過去急性骨髓性白血病病患的檢體，量化其Nr4A3 表現，進而分析跟病人臨床表現及預後的關聯。同時分析其他骨髓性血液疾病的病人檢體(慢性骨髓型白血病，骨髓化生不良症候群等)，測試此現象是否同時在其他血液疾病亦扮演角色。3 篩選一些曾被研究具有誘導骨髓性白血病細胞分化之藥物，檢測其效果是否與Nr4A3 表現量上升相關。利用細胞株，調控Nr4A3 的表現，探求其在急性骨髓性白血病藥物敏感度的影響。並利用動物模式，調升Nr4A3 表現及合併(傳統、標靶)藥物，尋求做為急性骨髓性白血病治療標的之可行性。<br> Abstract: BackgroundA lot of genetic aberrations have been found to contribute to the leukemogensis ofacute myeloid leukemia (AML). These genetic aberrations are classified into twogroups: the class I aberrations leading to survival or proliferation benefit of myeloidblood cells, and the class II aberrations blocking the differentiation of blood cells. It isthought that a myeloid progenitor cell transforms to AML when it harbors bothgenetic mutations. However, the interactions between these genetic aberrations, aswell as the definite steps by which a genetic aberration leads to leukemogenesis,remain to be clarified. Recently two orphan receptors Nr4A1 and Nr4A3 are reportedto play important roles in myeloid leukemogenesis as knocking-out of both genesresulted in acute myeloid leukemia in mouse models. The functions of two orphanreceptors Nr4A1 and Nr4A3 have not been well defined, although there are a fewstudies demonstrating that Nr4A1contributed to cell apoptosis. According to thetwo-hit theory mentioned above, if Nr4A1 down-regulation is able to block myeloidcells apoptosis and promote cell survival, Nr4A3 down-regulation is thus supposed tobe able to block the differentiation of myeloid cells during myeloid leukemogenesis.Preliminary ExplorationIn our study, K562 cell line differentiation induced by a chemical compound phorbol12-myristate 13-acetate (PMA) was accompanied with up-regulation of Nr4A3expression. In another clinical observation, Nr4A3 expression was also up-regulatedafter treatment with all-trans retinoic acid (ATRA) to induce primary leukemic celldifferentiation in a patient of acute promyelocytic leukemia (APL). These findingswere compatible with our hypothesis that Nr4A3 is able to induce myeloid leukemiccell differentiation.Experimental Hypothesis1 Nr4A3 down-regulation contributes to AML leukemogenesis by interference ofmyeloid differentiation.2 The expression levels of Nr4A3 in AML patients correlate to clinical feature andbears prognostic significance.3 Induction of the expression of Nr4A3 can overcome drug resistance of AML cellsand further translate to clinical therapeutic strategy to improve the patients’ outcome.Methods1 Using the cell-line models for differentiation-induction by chemical agents, wetest if the expression levels of Nr4A3 are associated with the differentiationphenotypes. Using small interfering RNA (siRNA) strategy, we observe if thedifferentiation phenotypes are interfered after knock-down of the Nr4A3 expression.2 We will quantify the Nr4A3 expression levels in archived AML specimens tocorrelate with the ptients’ clinical presentations, treatment outcomes and prognosis.We will also test the Nr4A3 expression levels in other myeloid blood diseases(chronic myeloid leukemia, myelodysplastic syndrome, etc.) to explore the scale ofthe phenomenon in other blood diseases.3 We will screen the available chemical compounds that has been reported to inducemyeloid leukemia cell differentiation, checking if they possess the ability of induceNr4A3 expression. In cell lines, we are going to test if up-regulation ordown-regulation of the Nr4A3 expressions have anything to do with the drugsensitivity or resistance. In animal model, to combine therapeutic agents andup-regulation of Nr4A3 expression will be evaluated in order to establish the rationalethat Nr4A3 could be a novel therapeutic target in AML patients.