本文選題：新型合成藥物 + 組蛋白去乙�；敢种苿���； 參考：《北京協(xié)和醫(yī)學院》2015年博士論文

【摘要】：研究目的：開發(fā)具有生物活性高、毒性低等特點的新藥一直是腫瘤治療研究領域的熱點,其中分子合成藥物由于能夠克服傳統(tǒng)化療藥物的缺陷而備受關注。NL-101是一種連接組蛋白去乙�；敢种苿㏒AHA以及烷化劑苯達莫司汀的活性基團而合成的化合物。本文探討這種化合物在體外和體內對于白血病細胞的殺傷作用以及相關分子機制。研究方法：采用MTT法檢測NL-101、SAHA和苯達莫司汀對多種白血病細胞系增殖能力的影響,用SPSS軟件計算半數(shù)抑制劑量(ICso),用CompuSyn軟件計算SAHA和苯達莫司汀的藥物聯(lián)合指數(shù)(CI)；用流式細胞術分析藥物處理后細胞的周期和凋亡變化；Western blot檢測藥物處理后組蛋白H3乙�；�,以及蛋白凋亡和DNA損傷相關蛋白包括y-H2AX、PARP、caspase-3、Bax、Bcl-2和Bcl-xL的表達；Ficoll法分離病人原代細胞,瑞氏染色觀察細胞經藥物處理后的形態(tài)改變。體內實驗部分,通過逆轉錄病毒感染構建共表達AML1-ETO和C-KIT突變的移植白血病小鼠模型,給予NL-101、苯達莫司汀和SAHA注射治療,動態(tài)監(jiān)測小鼠體重以及體內白血病細胞浸潤情況,記錄各組小鼠生存期。研究結果：在體外實驗中：(1)苯達莫司汀和SAHA在白血病細胞中具有協(xié)同作用。(2)NL-101對多種白血病細胞系的生長有明顯的抑制作用,其中Kasumi-1和NB4細胞的敏感性最為顯著；NL-101對正常細胞系的作用較小,顯示出對腫瘤細胞作用的選擇性；NL-101的抗增殖能力顯著高于苯達莫司汀,與SAHA沒有顯著差異。(3)NL-101導致白血病細胞周期阻滯于S期,以及顯著的細胞凋亡。(4)NL-101的細胞毒性作用在分子水平上表現(xiàn)為激活依賴caspase-3的凋亡通路以及Bcl-2家族促凋亡蛋白；NL-101能增加DNA損傷標志蛋白y-H2AX和乙�；M蛋白H3的表達。(5)NL-101能誘導急性髓系白血病患者的原代細胞發(fā)生DNA損傷和細胞凋亡。在體內實驗中：(1)成功建立共表達AML1-ETO和C-KIT突變的移植白血病小鼠模型,造血器官中有大量原始細胞(c-Kit+、Gr1-、CD11b-)浸潤。(2)NL-101能抑制t(8；21)白血病細胞的增殖和浸潤,顯著延長白血病小鼠的生存期,具有比苯達莫司汀和SAHA更強的抗白血病活性。(3)NL-101對小鼠體重沒有顯著影響,小鼠的耐受性良好。研究結論：由SAHA和苯達莫司汀的活性基團合成的新型藥物NL-101,在體內及體外實驗中均具有抑制白血病細胞增殖以及誘導細胞凋亡的生物學效應。針對作用機制的研究顯示NL-101具有抑制組蛋白去乙�；负推茐腄NA結構的雙重活性。
[Abstract]:Objective: to develop new drugs with high bioactivity and low toxicity has been a hot topic in the field of tumor therapy. Among them, molecular synthetic drugs have attracted much attention because of their ability to overcome the defects of traditional chemotherapeutic drugs. NL-101 is a kind of compound synthesized by linking histone deacetylase inhibitor SAHA and the active group of benzamoastine. The cytotoxicity of this compound to leukemia cells in vitro and in vivo and its molecular mechanisms were investigated. Methods: the effects of NL-101 SAHA and bendamoastine on the proliferation of various leukemia cell lines were detected by MTT assay. SPSS software was used to calculate the dose of ICSU, CompuSyn software was used to calculate the combined index of SAHA and bentamoastine, and flow cytometry was used to analyze the changes of cell cycle and apoptosis after drug treatment. Western blot was used to detect the acetylation level of histone H3 after drug treatment. The expression of protein apoptosis and DNA damage related proteins, including the expression of y-H2AXPnPARPncaspase-3, Bcl-2 and Bcl-xL, was detected by Ficoll method. The morphological changes of the cells treated with drugs were observed by Rayleigh staining. In vivo experiment, the transplanted leukemia mice model which co-expressed AML1-ETO and C-KIT mutations was constructed by retrovirus infection. NL-101, bentamoastine and SAHA were injected into mice to dynamically monitor the body weight and infiltration of leukemia cells in vivo. The survival time of mice in each group was recorded. Results: in vitro, the effects of bendamolastine and SAHA on the growth of various leukemia cell lines were significantly inhibited. The sensitivity of Kasumi-1 and NB4 cells was the most significant. NL-101 had little effect on normal cell lines, which showed that the selective action of NL-101 on tumor cells was significantly higher than that of bentamoastine. There was no significant difference between NL-101 and SAHA. NL-101 resulted in cell cycle arrest in S phase. The cytotoxic effects of NL-101 on apoptosis were as follows: activation of caspase-3 dependent apoptotic pathway and increased expression of DNA damage marker protein y-H2AX and acetylated histone H3 in Bcl-2 family. It can induce DNA damage and apoptosis in primary cells of patients with acute myeloid leukemia. In vivo, the mouse model of leukemia transplanted with AML1-ETO and C-KIT mutations was successfully established. A large number of primitive cells, c-Kit Gr1-Gr1-NL-101, could inhibit the proliferation and infiltration of tn821) leukemia cells, and prolong the survival time of leukemia mice. There was no significant effect on body weight of mice with stronger antileukemia activity than bendamoastine and SAHA. The mice had good tolerance. Conclusion: NL-101, a novel drug synthesized from the active groups of SAHA and bentamoastine, has the biological effects of inhibiting the proliferation and inducing apoptosis of leukemia cells in vivo and in vitro. Studies on the mechanism of action showed that NL-101 had dual activities of inhibiting histone deacetylase and destroying the structure of DNA.
【學位授予單位】：北京協(xié)和醫(yī)學院
【學位級別】：博士
【學位授予年份】：2015
【分類號】：R733.7

【相似文獻】

相關期刊論文 前10條

1 任健;程卯生;;鹽酸苯達莫司汀(bendamustine hydrochloride)[J];中國藥物化學雜志;2008年05期

2 陳棟;宮平;;鹽酸苯達莫司汀[J];中國藥物化學雜志;2009年02期

3 吳蔚;劉智;程航;劉姍姍;余鵬;程澤能;;高效液相色譜熒光檢測法測定人血漿中苯達莫司汀及γ-羥基化苯達莫司汀濃度[J];腫瘤藥學;2011年01期

4 黃世杰;抗腫瘤藥苯達莫司汀[J];國外醫(yī)學.藥學分冊;2001年06期

5 王曉坤;;新型抗腫瘤藥物——苯達莫司汀[J];齊魯藥事;2009年09期

6 陳祥峰;楊奇珍;魏佳;;注射用鹽酸苯達莫司汀的制備及質量控制[J];中國藥房;2012年13期

7 陳磊;葉東;楊建楠;陳林;孫強;;鹽酸苯達莫司汀的合成工藝改進[J];中國藥師;2013年02期

8 劉珍仁;葉曉嵐;范國榮;;鹽酸苯達莫司汀臨床給藥過程中的降解產物及其結構確證[J];中國新藥與臨床雜志;2013年03期

9 高麗梅;汪燕翔;宋丹青;;鹽酸苯達莫司汀的合成[J];中國新藥雜志;2007年23期

10 宗在偉;杜有國;陳磊;楊建楠;;鹽酸苯達莫司汀的合成研究[J];海峽藥學;2011年11期

相關重要報紙文章 前2條

1 金景新　譯;歐洲推薦批準苯達莫司汀治療白血病[N];中國醫(yī)藥報;2010年

2 陸志城;鹽酸苯達莫司汀凍干粉針[N];醫(yī)藥經濟報;2009年

相關博士學位論文 前1條

1 喻靜;新型HDAC抑制劑—苯達莫司汀化合物NL-101的抗白血病作用及機制研究[D];北京協(xié)和醫(yī)學院;2015年

相關碩士學位論文 前5條

1 張勝菊;鹽酸苯達莫司汀及其衍生物的合成與表征[D];南昌大學;2010年

2 劉珍仁;鹽酸苯達莫司汀的質量分析[D];第二軍醫(yī)大學;2012年

3 穆楠;鹽酸苯達莫司汀的合成[D];河北工業(yè)大學;2013年

4 殷昕;伏立諾他和鹽酸苯達莫司汀的合成與結構修飾[D];華東理工大學;2012年

5 孔芳;苯達莫司�。˙endamustine）誘導小鼠乳腺癌細胞凋亡及其機制的研究[D];山東大學;2007年

，

本文編號：1939932