本文選題：靶向 + 聚合　； 參考：《浙江省醫(yī)學科學院》2015年碩士論文

【摘要】：目的：肝細胞肝癌(hepatocelluar carcinoma, HCC)是致死率很高的惡性腫瘤之一,目前的治療手段效果有限。新興的基因治療給腫瘤患者帶來了新的希望,可以通過組織特異性啟動子調(diào)控導入細胞的治療性基因靶向表達。然而,肝癌基因治療的靶點多集中于肝癌發(fā)生發(fā)展過程中的相關基因,只能對相應類型的腫瘤起效。在本研究中,我們選擇DNA復制環(huán)節(jié)這一所有細胞增殖的關鍵步驟作為治療靶點,來抑制腫瘤細胞生長。方法：本課題構建了包含HCC特異性甲胎蛋白(a-fetoprotein, AFP)增強子的重組AFP啟動子調(diào)控串聯(lián)連接的分別靶向DNA聚合酶α,δ,ε的人工microRNA和重組活化Caspase3基因的重組腺病毒Ad/AFP-Casp-AFP-amiR,同時構建僅有重組AFP啟動子的重組腺病毒Ad/AFP作為空白病毒對照。結果：體外實驗：重組腺病毒以MOI 50感染AFP強陽性的HCC細胞HepG2、AFP弱陽性的HCC細胞Hep3B和AFP陰性的正常肝細胞HL7702, Ad/AFP-Casp-AFP-amiR能夠抑制HepG2和Hep3B DNA聚合酶α,δ,ε的表達,阻滯細胞周期在G1期；誘發(fā)HepG2和Hep3B細胞凋亡、抑制細胞增殖。Ad/AFP-Casp-AFP-amiR對肝癌細胞的抑制效應與AFP的表達水平正相關。Ad/AFP-Casp-AFP-amiR對正常肝細胞HL7702沒有作用。體內(nèi)實驗：建立裸鼠Hep3B移植瘤模型,瘤內(nèi)注射Ad/AFP-Casp-AFP-amiR,未能觀察到明顯的抑瘤效應,但能夠誘發(fā)細胞凋亡。結論：Ad/AFP-Casp-AFP-amiR有望成為新的肝癌基因治療藥物,通過AFP啟動子特異性抑制AFP陽性的肝癌細胞DNA聚合酶α,δ,￡表達,阻遏肝癌細胞生長,誘發(fā)肝癌細胞凋亡。這一方法基于抑制細胞增殖的關鍵步驟DNA復制,不局限于某一種基因異常表達的HCC,為肝癌的基因治療提供了新的思路。
[Abstract]:Objective: hepatocellular carcinoma (HCC) is one of the malignant tumors with high mortality. The new gene therapy has brought new hope to tumor patients, which can regulate the target expression of therapeutic gene by tissue specific promoter. However, the target of gene therapy for liver cancer is mainly related to the occurrence and development of liver cancer, which can only work on the corresponding types of tumor. In this study, we selected DNA replication, a key step of cell proliferation, as a therapeutic target to inhibit the growth of tumor cells. Methods: in this study, the recombinant AFP promoter containing HCC specific alpha-fetoprotein (AFP) enhancer was constructed to regulate the tandem linkage of DNA polymerase 偽, 未, 蔚, and the recombinant adenovirus Ad-AFP-Casp-AFP-amiR was constructed, and the recombinant adenovirus Ad-AFP-Casp-AFP-amiR was constructed respectively, which was targeted at DNA polymerase 偽, 未, 蔚, and the recombinant adenovirus Ad-AFP-Casp-AFP-amiR, respectively. The recombinant adenovirus Ad/AFP with only recombinant AFP promoter served as a blank virus control. Results: in vitro, recombinant adenovirus infected AFP strongly positive HCC cell HepG2Afefefefefefefefefefefefefefefefefefefefefefe-positive HCC cells with MOI 50 could inhibit the expression of HepG2 and Hep3B DNA polymerase 偽, 未, 蔚 and arrest the cell cycle in G1 phase, and Ad/AFP-Casp-AFP-amiR could inhibit the expression of HepG2 and Hep3B DNA polymerase 偽, 未, 蔚 in normal hepatocytes. The apoptosis of HepG2 and Hep3B cells was induced, and the inhibitory effect of Ad-r-AFP-Casp-AFP-amiR on hepatoma cells was positively correlated with the expression level of AFP. Adr-AFP-Casp-AFP-amiR had no effect on HL7702 of normal hepatocytes. In vivo experiment: Hep3B transplanted tumor model was established in nude mice. Ad-AFP-Casp-AFP-amiR was injected intratumoral into the tumor. No obvious tumor inhibition effect was observed, but apoptosis was induced by the injection of Ad-AFP-Casp-AFP-amiR. Conclusion the AFP promoter can inhibit the expression of DNA polymerase 偽, 未 in AFP positive hepatoma cells, inhibit the growth of HCC cells and induce apoptosis of HCC cells. This method is based on the key step of inhibiting cell proliferation, and is not limited to the abnormal expression of DNA, which provides a new idea for gene therapy of hepatocellular carcinoma.
【學位授予單位】：浙江省醫(yī)學科學院
【學位級別】：碩士
【學位授予年份】：2015
【分類號】：R735.7;R450

【共引文獻】

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2 任小強;張建國;辛士永;程濤;;MicroRNA-218(miR-218)在腎透明細胞癌中的表達及臨床意義[J];中外醫(yī)療;2014年11期

3 楊穎卓;康鵬;高杰;徐春林;王詩美;吳霞;;肝特異性miR-122調(diào)控肝臟疾病的研究進展[J];臨床肝膽病雜志;2014年05期

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本文編號：1949696