帕尼单抗生物类似物对肿瘤的抑制及放疗增敏作用的研究

发布时间：2018-05-30 02:41

本文选题：KN032 + 帕尼单抗　；参考：《苏州大学》2016年硕士论文

【摘要】：目的比较一种帕尼单抗生物类似物KN032与帕尼单抗的生物活性以及对肿瘤生长抑制作用的相似性,研究KN032的放射治疗增敏作用及其机制。方法选择EGFR高表达的表皮癌细胞株A431为研究对象,采用MTT方法检测KN032和Panitumumab在体外对肿瘤生长的抑制作用,并比较它们对肿瘤抑制的生物学相似性;采用流式细胞学技术检测Panitumumab和KN032与A431细胞的结合能力,比较它们的相似性;建立A431细胞裸鼠荷瘤模型,随机分为对照组、KN032 20μg组、KN032 200μg组、Panitumumab 20μg组、Panitumumab 200μg组五组,观察两种抗体在不同浓度时在体内对肿瘤增殖的影响,并且比较两种抗体抑制作用的相似性;通过克隆形成实验分析KN032对A431细胞的放疗增敏作用,设单纯照射、KN032+照射组、Panitumumab+照射组3组,采用免疫荧光方法检测不同实验组细胞中γH2AX的表达。结果MTT结果显示KN032和Panitumumab对A431细胞均有抑制作用,在4μg/mL的浓度范围内,随着药物浓度的增加,抗体对细胞的抑制效果增强,并且在各个相同的浓度下,KN032和Panitumumab对细胞的抑制作用相似;流式细胞学实验结果显示KN032与A431细胞的结合能力与Panitumumab相似,KN032的EC50=0.21,Panitumumab的EC50=0.23;体内实验显示,KN032和Panitumumab对小鼠肿瘤生长均有抑制作用,并且当剂量为200μg时,抗体对肿瘤生长的抑制作用强于20μg剂量时,并且在相同剂量时两者对小鼠肿瘤生长抑制作用相似;细胞克隆形成实验显示,KN032和Panitumumab对A431细胞均具有放疗增敏作用,并且两种抗体的放疗增敏作用效果相似;免疫荧光实验显示,KN032和Panitumumab可以增加X射线照射后0.5和4h时的A431细胞核内γH2AX的表达量(P0.05),在射线照射后12h时,三组细胞核内γH2AX表达量较射线照射后初期均明显下降,但三组间的差异无统计学意义。结论KN032对肿瘤生长有明显的抑制作用,其生物活性与帕尼单抗相似。KN032能够提高放射治疗的敏感性,其增敏效果与Panitumumab相似,其作用机制可能与增强DNA双链损伤、抑制损伤后DNA双链断裂修复有关。
[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 纬 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 纬 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 纬 H2AX in the nucleus of A431 at 0.5 and 4 h after X-ray irradiation. The expression of 纬 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.
【学位授予单位】：苏州大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R73-36

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