發(fā)布時(shí)間：2018-05-30 02:41

  本文選題：KN032 + 帕尼單抗　； 參考：《蘇州大學(xué)》2016年碩士論文

【摘要】：目的比較一種帕尼單抗生物類(lèi)似物KN032與帕尼單抗的生物活性以及對(duì)腫瘤生長(zhǎng)抑制作用的相似性,研究KN032的放射治療增敏作用及其機(jī)制。方法選擇EGFR高表達(dá)的表皮癌細(xì)胞株A431為研究對(duì)象,采用MTT方法檢測(cè)KN032和Panitumumab在體外對(duì)腫瘤生長(zhǎng)的抑制作用,并比較它們對(duì)腫瘤抑制的生物學(xué)相似性;采用流式細(xì)胞學(xué)技術(shù)檢測(cè)Panitumumab和KN032與A431細(xì)胞的結(jié)合能力,比較它們的相似性;建立A431細(xì)胞裸鼠荷瘤模型,隨機(jī)分為對(duì)照組、KN032 20μg組、KN032 200μg組、Panitumumab 20μg組、Panitumumab 200μg組五組,觀察兩種抗體在不同濃度時(shí)在體內(nèi)對(duì)腫瘤增殖的影響,并且比較兩種抗體抑制作用的相似性;通過(guò)克隆形成實(shí)驗(yàn)分析KN032對(duì)A431細(xì)胞的放療增敏作用,設(shè)單純照射、KN032+照射組、Panitumumab+照射組3組,采用免疫熒光方法檢測(cè)不同實(shí)驗(yàn)組細(xì)胞中γH2AX的表達(dá)。結(jié)果MTT結(jié)果顯示KN032和Panitumumab對(duì)A431細(xì)胞均有抑制作用,在4μg/mL的濃度范圍內(nèi),隨著藥物濃度的增加,抗體對(duì)細(xì)胞的抑制效果增強(qiáng),并且在各個(gè)相同的濃度下,KN032和Panitumumab對(duì)細(xì)胞的抑制作用相似;流式細(xì)胞學(xué)實(shí)驗(yàn)結(jié)果顯示KN032與A431細(xì)胞的結(jié)合能力與Panitumumab相似,KN032的EC50=0.21,Panitumumab的EC50=0.23;體內(nèi)實(shí)驗(yàn)顯示,KN032和Panitumumab對(duì)小鼠腫瘤生長(zhǎng)均有抑制作用,并且當(dāng)劑量為200μg時(shí),抗體對(duì)腫瘤生長(zhǎng)的抑制作用強(qiáng)于20μg劑量時(shí),并且在相同劑量時(shí)兩者對(duì)小鼠腫瘤生長(zhǎng)抑制作用相似;細(xì)胞克隆形成實(shí)驗(yàn)顯示,KN032和Panitumumab對(duì)A431細(xì)胞均具有放療增敏作用,并且兩種抗體的放療增敏作用效果相似;免疫熒光實(shí)驗(yàn)顯示,KN032和Panitumumab可以增加X(jué)射線照射后0.5和4h時(shí)的A431細(xì)胞核內(nèi)γH2AX的表達(dá)量(P0.05),在射線照射后12h時(shí),三組細(xì)胞核內(nèi)γH2AX表達(dá)量較射線照射后初期均明顯下降,但三組間的差異無(wú)統(tǒng)計(jì)學(xué)意義。結(jié)論KN032對(duì)腫瘤生長(zhǎng)有明顯的抑制作用,其生物活性與帕尼單抗相似。KN032能夠提高放射治療的敏感性,其增敏效果與Panitumumab相似,其作用機(jī)制可能與增強(qiáng)DNA雙鏈損傷、抑制損傷后DNA雙鏈斷裂修復(fù)有關(guān)。
[Abstract]:Objective to compare the biological activity of a Panimab biological analogue (KN032) with that of Pannimab (Pannimab) and its similarity to tumor growth inhibition, and to study the radiosensitizing effect of KN032 and its mechanism. Methods Epidermal cancer cell line A431 with high expression of EGFR was selected as the study object. The inhibitory effect of KN032 and Panitumumab on tumor growth was detected by MTT method in vitro, and the biological similarity of their inhibition to tumor was compared. Flow cytometry was used to detect the binding ability of Panitumumab and KN032 to A431 cells, and to compare their similarity, and to establish a tumor-bearing model of A431 cells in nude mice, which was randomly divided into five groups: control group (KN0320 渭 g), KN032 200 渭 g group, Panitumumab 20 渭 g group, Panitumumab 200 渭 g group. To observe the effects of two antibodies on tumor proliferation in vivo at different concentrations, and to compare the similarities of the inhibitory effects of the two antibodies, and to analyze the radiosensitization effect of KN032 on A431 cells by clone formation assay. The expression of 緯 H2AX in the cells of different experimental groups was detected by immunofluorescence method. Results MTT showed that both KN032 and Panitumumab could inhibit A431 cells. In the range of 4 渭 g/mL, the inhibitory effect of antibody on A431 cells was enhanced with the increase of drug concentration. The inhibitory effects of KN032 and Panitumumab on cells were similar at the same concentration. The results of flow cytology showed that the binding ability of KN032 to A431 cells was similar to that of Panitumumab, and that of EC50KN032 and Panitumumab EC500.23.The in vivo experiments showed that both KN032 and Panitumumab could inhibit the growth of mice tumor, and when the dose was 200 渭 g, The inhibitory effect of antibody on tumor growth was stronger than that at 20 渭 g dose, and at the same dose, both of them had similar inhibitory effects on tumor growth, and cell clone formation assay showed that both KN032 and Panitumumab could sensitize A431 cells by radiotherapy. Immunofluorescence assay showed that KN032 and Panitumumab could increase the expression of 緯 H2AX in the nucleus of A431 at 0.5 and 4 h after X-ray irradiation, and P0.05 at 12 h after irradiation, and the effect of the two antibodies on radiosensitization was similar, and the results showed that KN032 and Panitumumab could increase the expression of 緯 H2AX in the nucleus of A431 at 0.5 and 4 h after X-ray irradiation. The expression of 緯 H2AX in the nuclei of the three groups was significantly lower than that in the early days after irradiation, but there was no significant difference between the three groups. Conclusion KN032 has obvious inhibitory effect on tumor growth, and its biological activity is similar to that of Panimab. KN032 can increase the sensitivity of radiotherapy, its sensitizing effect is similar to that of Panitumumab, and its mechanism may be related to the enhancement of DNA double strand damage. Inhibition of DNA double strand break repair after injury is related.
【學(xué)位授予單位】：蘇州大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2016
【分類(lèi)號(hào)】：R73-36

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本文編號(hào)：1953570