2011-08-012024-05-13https://scholars.lib.ntu.edu.tw/handle/123456789/645530摘要：Cordycepin，或被稱為3-deoxyadenosine，是“冬蟲夏草”中的重要活性萃取物。它的結構類似於adenosine，因而具有抑制mRNA 合成過程中重要的polyadenylatepolymerase 的功能，從而可阻斷細胞內的mRNA 及蛋白質生成而造成細胞死亡。另一方面，cordycepin 又可活化細胞表面的adenosine 受器，從而影響細胞的功能。在以往文獻中，cordycepin 被認為可以造成多種癌細胞的凋亡，但它對急性骨髓性白血病細胞的效果及可能的機轉卻未被詳細探討。在我們的初步試驗中，cordycepin 可在4 小時內引發U937 人類急性骨髓性白血病細胞株的凋亡，同時會導致細胞內β-catenin 蛋白量的下降。這個效應會因為蛋白體抑制劑(proteasome inhibitors)的存在或是glycogensynthase kinase(GSK)-3β 抑制劑的存在而消失。我們因而合理的假定cordycepin 是經由活化GSK-3β 造成β-catenin 蛋白的磷酸化增加進而使其分解速度增加而誘發細胞凋亡。另一方面，由於Wnt/β-catenin 訊息傳遞路徑的活化在白血病幹細胞的生長及去分化中扮演重要角色，我們也假設cordycepin 對白血病幹細胞具有毒性而能做為清除白血病幹細胞的重要藥物。根據這些假設，我們在本計畫中將設定並執行下列目標：(1).探討cordycepin 在人類急性骨髓性白血病細胞株中影響Wnt/β-catenin 訊息傳遞路徑的可能控制機轉及其透過Wnt/β-catenin 訊息傳遞路徑對下游分子及細胞功能的影響。(2).在急性骨髓性白血病病患檢體中驗證上述影響。(3). 進一步探討cordycepin 對急性骨髓性白血病細胞的轉譯體(transcriptome)及蛋白體(proteome)的影響。(4).在體外培養系統及小鼠異種造血幹細胞移植系統中檢驗cordycepin 對急性骨髓性白血病幹細胞及正常造血幹細胞的影響及可能機轉。預期計劃完成後，我們將對cordycepin 對急性骨髓性白血病的治療效果及可能機轉有一個完整的實驗室及臨床前評估及了解。由於目前為止沒有任何一個臨床藥物可以針對Wnt/β-catenin 訊息傳遞路徑加以抑制，這個計畫會讓我們更了解cordycepin 作為Wnt/β-catenin 訊息傳遞路徑的效果。一方面由於這些cordycepin 的藥理作用特殊性，更由於cordycepin 的毒性考慮會因為其為一種長期被使用的中藥的萃取物而大為降低，相信cordycepin 用於第一或二期人體試驗治療急性骨髓性白血病的可能性會由於本計畫的研究成果而大為提高。<br> Abstract: Cordycepin, also now known as 3-deoxyadenosine, is a major active ingredient in theextract of “Dong Chong Xia Cao”. As an adenosine analogue, cordycepin could suppress theactivities of polyadenylate polymerase (PAP), then terminate mRNA synthesis prematurelyand finally result in cell death. On the other way, cordycepin also could activate adenosinereceptor on cell membrane and affect cell functions. Cordycepin had been reported to induceapoptosis in various types of cancer, but detailed analysis of its effects on acute myeloidleukemia (AML) cells has not been illustrated. In our preliminary experiments, cordycepinwas found to inhibit the growth of the U937 human AML cell line as early as within 4 hours,and the amount of β-catenin was decreased at the same time. Proteasome inhibitors restoredthe amount of β-catenin, and inhibitors of glycogen synthase kinase(GSK)-3β had a similareffect. We then hypothesize that cordycepin will increase the activity of GSK-3β, whichresults in increased β-catenin phosphorylation and degradation and finally in cell death. Andsince the Wnt/β-catenin pathway is crucial to the survival of leukemic stem cells (LSCs), wecan also reasonably assumpt that cordycepin can be LSC-toxic and cordycepin treatmentwould benefit in eradicating LSCs. Based on our hypothesis, we will target to the followingaims in this project: (1). To illustrate the possible mechanisms about how cordycepin affectsthe signaling of Wnt/β-catenin pathway and also the downstream effects in AML cell lines. (2).To validate the phenomenon found in aim (1) in primary human AML cells. (3). To explorethe change of transcriptome and proteome of AML cells after exposure of cordycepin. (4). Toevaluate the toxicities of cordycepin on AML stem cells and normal hematopoietic stem cellsby in vitro colony-forming assay and in vivo SCID mice transplantation model. With theaccomplishment of this project, we will then have a throughout pre-clinical evaluation aboutusing cordycepin in treating acute myeloid leukemia and rander cordycepin as a inhibitor ofWnt/β-catenin signal, which has never been targeted before. With our data, human phase I/Iclinical trials can also be engaged soon, because cordycepin is an extract from a long usedherb drug and therefore toxicities would be a less concern.Cordycepin急性骨髓性白血病Beta-cateninGSK-3betaCordycepinAcute myeloid leukemiaBeta-cateninGSK-3beta