【摘要】：宮頸癌是常見的婦科惡性腫瘤之一，占全世界婦女癌癥死亡率第二位，僅次于乳腺癌。在某些發(fā)展中國家宮頸癌死亡率甚至居首位。宮頸癌的傳統(tǒng)治療方法有手術(shù)、放療、化療等，但這些手段各有其局限性。研究表明，HPV是宮頸癌的主要致病因子，幾乎所有的宮頸癌發(fā)病都與高危型HPV感染有關(guān)。本課題以宮頸癌治療性疫苗為中心，開展了兩個方向的研究，即基于HPV16E6E7基因疫苗的抗腫瘤免疫反應(yīng)研究以及優(yōu)化突變?nèi)诤匣騂PV18E6E7的去致癌性檢測。 E6和E7蛋白是高危型HPV的主要致癌蛋白，E6蛋白通過結(jié)合并降解P53蛋白，，破壞正常的細(xì)胞周期調(diào)控，促使細(xì)胞永生化；E7蛋白則通過結(jié)合并降解pRb蛋白而使其喪失抑癌功能。E6和E7蛋白在正常組織中不表達(dá)，只表達(dá)于HPV感染組織中，因此E6和E7蛋白是制備宮頸癌治療性疫苗的理想靶抗原。 在疫苗的抗腫瘤免疫反應(yīng)研究中，以優(yōu)化突變?nèi)诤系腍PV16E6E7基因?yàn)榭乖�，�?gòu)建重組腺病毒疫苗和重組DNA疫苗，另外用表達(dá)野生型和突變型E6E7蛋白的重組腺病毒感染小鼠樹突狀細(xì)胞，得到致敏樹突狀細(xì)胞疫苗。用ELISPOT檢測疫苗免疫小鼠后誘導(dǎo)的細(xì)胞免疫反應(yīng)；通過腫瘤移植保護(hù)實(shí)驗(yàn)檢測疫苗的抗腫瘤效果。結(jié)果表明致敏樹突細(xì)胞引發(fā)了強(qiáng)烈的細(xì)胞免疫反應(yīng)，同時具有較好的抗腫瘤免疫效果。在優(yōu)化突變?nèi)诤匣騂PV18E6E7的去致癌性檢測中，本課題參照人類基因中的優(yōu)勢密碼子，對HPV18E6基因和E7基因進(jìn)行優(yōu)化合成，同時對E6基因和E7基因進(jìn)行定點(diǎn)突變和融合，以消除其致癌性。實(shí)驗(yàn)表明兩種優(yōu)化突變?nèi)诤系腍PV18E6E7基因失去了對NIH3T3細(xì)胞的轉(zhuǎn)化活性和對BALB/c裸鼠的致瘤作用，表明優(yōu)化融合突變基因是安全的。
[Abstract]:Cervical cancer is one of the most common gynecological malignancies, accounting for the second highest cancer mortality rate in the world, second only to breast cancer. In some developing countries, cervical cancer mortality even ranks first. The traditional treatment of cervical cancer includes surgery, radiotherapy, chemotherapy and so on, but each of these methods has its limitations. Studies have shown that HPV is the main cause of cervical cancer, almost all cervical cancer is associated with high-risk HPV infection. This paper focuses on the therapeutic vaccine for cervical cancer, and has carried out two research directions. That is, the study of anti-tumor immune response based on HPV16E6E7 gene vaccine and the optimized detection of carcinogenicity of mutant fusion gene HPV18E6E7. E6 and E7 proteins are the main carcinogenic proteins of high risk HPV. Disrupting normal cell cycle regulation, impelling cell immortalized E7 protein to lose its tumor suppressor function by binding and degrading PRB protein, E6 and E7 proteins were not expressed in normal tissues, but only in HPV infected tissues. Therefore, E6 and E7 proteins are ideal target antigens for the preparation of therapeutic vaccines for cervical cancer. The recombinant adenovirus vaccine and recombinant DNA vaccine were constructed by optimizing the fusion of HPV16E6E7 gene as antigen gene in the study of anti-tumor immune response of the vaccine. In addition, the mouse dendritic cells were infected with recombinant adenovirus expressing wild type and mutant E6E7 protein, and the sensitized dendritic cell vaccine was obtained. Elisa pot was used to detect the cellular immune response induced by the vaccine, and the anti-tumor effect of the vaccine was tested by tumor transplantation protection test. The results showed that the sensitized dendritic cells induced a strong cellular immune response and had a good antitumor immune effect. In the detection of the decarcinogenicity of the mutant fusion gene HPV18E6E7, HPV18E6 gene and E7 gene were optimized and synthesized according to the dominant codon of human gene, and the E6 gene and E7 gene were mutated and fused. To eliminate its carcinogenicity. The results showed that the two optimized fusion genes of HPV18E6E7 lost the transformation activity of NIH3T3 cells and the tumorigenicity of BALB- / c nude mice, which indicated that the optimized fusion gene was safe.
【學(xué)位授予單位】：北京工業(yè)大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2014
【分類號】：R737.33

【引證文獻(xiàn)】

相關(guān)期刊論文 前2條

1 張玉;孫立新;;人乳頭瘤病毒E6/E7蛋白在子宮頸癌中的研究進(jìn)展[J];腫瘤研究與臨床;2016年08期

2 羅小蘭;毛熙光;;HPV16E7蛋白在宮頸癌中的作用研究進(jìn)展[J];現(xiàn)代醫(yī)藥衛(wèi)生;2016年04期

相關(guān)碩士學(xué)位論文 前1條

1 張玉;HPV E6癌蛋白檢測在宮頸癌篩查中的臨床應(yīng)用研究[D];山西醫(yī)科大學(xué);2016年

本文編號：2132460