本文選題：TNF-α + 拮抗分子��； 參考：《中國人民解放軍軍事醫(yī)學(xué)科學(xué)院》2005年博士論文

【摘要】：TNF-α的過量表達(dá)與很多疾病密切相關(guān),應(yīng)用TNF-α的拮抗劑部分阻斷這些疾病中的TNF-α水平,將成為一種有效的治療措施。已開發(fā)的TNF-α拮抗劑主要包括兩大類,即TNF受體與IgG的FC段形成的融合蛋白和抗TNF-α的單抗,國外已有成熟產(chǎn)品上市。開發(fā)具有自主知識(shí)產(chǎn)權(quán)的TNF-α拮抗分子具有重要的理論和實(shí)際意義。 本論文擬根據(jù)構(gòu)建的TNF-α與本實(shí)驗(yàn)室獲得的鼠源功能性單抗Z12相互作用的復(fù)合物模型合理設(shè)計(jì)得到的低親和力拮抗肽,借助建立的基于抗原—抗體相互作用結(jié)構(gòu)信息設(shè)計(jì)抗體模擬物的技術(shù)平臺(tái),創(chuàng)造性地提出了三種提高TNF-α拮抗分子生物學(xué)活性的技術(shù):(1)將設(shè)計(jì)的短肽通過Linker與人IgG1的FC段連接后構(gòu)建成的免疫黏附分子(短肽—Linker-FC);(2)利用人抗體重鏈可變區(qū)(VH)的框架結(jié)構(gòu)作為支架,同時(shí)展示三條短肽,設(shè)計(jì)成新型分子PTVH5;(3)優(yōu)化TNF-α與Z12的復(fù)合物模型,設(shè)計(jì)活性提高的短肽。具體研究結(jié)果如下: (1)針對(duì)設(shè)計(jì)的TNF-α拮抗肽(PT1～PT4),選擇抑制TNF-α細(xì)胞毒性效果最好的PT4,利用Linker將其與人IgG1的FC段連接,構(gòu)建成PT4-Linker-FC融合蛋白,提高了PT4熱力學(xué)穩(wěn)定性。所構(gòu)建的融合蛋白能夠競(jìng)爭(zhēng)抑制HRP標(biāo)記的Z12與TNF-α的結(jié)合,能夠明顯抑制TNF-α介導(dǎo)的細(xì)胞毒效應(yīng),其抑制活性比單獨(dú)的PT4有明顯提高。 (2)考察能否利用人抗體可變區(qū)的框架結(jié)構(gòu)作為支架,同時(shí)展示三個(gè)活性肽段。選擇已設(shè)計(jì)的功能肽PT2、PT3和PT4,分別替換人抗體IgG重鏈可變區(qū)的三個(gè)CDR區(qū),設(shè)計(jì)成抑制TNF-α的新型分子(命名為PTVH)。利用同源模建技術(shù)從
[Abstract]:The overexpression of TNF- 偽 is closely related to many diseases. The use of TNF- 偽 antagonist to partially block the level of TNF- 偽 in these diseases will be an effective treatment. The developed TNF- 偽 antagonists mainly include two categories: fusion protein formed by FC segment of TNF receptor and IgG and monoclonal antibody against TNF- 偽. It is of great theoretical and practical significance to develop TNF- 偽 antagonist molecules with independent intellectual property rights. In this paper, the low affinity antagonist peptide was designed according to the constructed complex model of TNF- 偽 interacting with mouse functional monoclonal antibody Z12. A technical platform for the design of antibody simulants based on the structure information of antigen-antibody interaction, Three techniques for enhancing the antagonistic molecular biological activity of TNF- 偽 were proposed in this paper: 1) Immunoadhesive molecules (short peptide-Linker-FCX _ 2) using the variable region of heavy chain of human antibody were constructed by connecting the designed short peptide with FC segment of human IgG1 through Linker. The frame structure acts as a scaffold, At the same time, three short peptides were designed to optimize the complex model of TNF- 偽 and Z12, and to design short peptides with improved activity. The specific findings are as follows: 1) for the designed TNF- 偽 antagonist peptide PT1, PT4 was selected to inhibit the cytotoxicity of TNF- 偽. The fusion protein of PT4-Linker-FC was constructed by ligating it with FC segment of human IgG1 by Linker, and the thermodynamics stability of PT4 was improved. The constructed fusion protein could competitively inhibit the binding of HRP labeled Z12 to TNF- 偽 and inhibit the cytotoxic effect mediated by TNF- 偽. The inhibitory activity of the fusion protein was significantly higher than that of PT4 alone. To investigate whether the frame structure of variable region of human antibody can be used as scaffold, and to display three active peptide segments at the same time. The designed functional peptides PT2, PT3 and PT4 were selected to replace the three CDR regions of the heavy chain variable region of human antibody IgG, respectively, and were designed as a novel molecule (named PTVHG) to inhibit TNF- 偽. Using the technology of homologous modeling
【學(xué)位授予單位】：中國人民解放軍軍事醫(yī)學(xué)科學(xué)院
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2005
【分類號(hào)】：R392

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