CN114137215A - 一种结直肠癌诊断生物标志物mdk蛋白及其应用 - Google Patents
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Abstract
本发明公开了一种结直肠癌诊断生物标志物MDK蛋白及其应用。本发明通过免疫组织化学方法检测MDK蛋白在结直肠癌组织中的表达，结果显示MDK蛋白在结直肠癌中的表达是异常的，MDK蛋白在结直肠癌患者癌组织中的表达量显著高于癌旁组织。本发明提示MDK蛋白可以作为一种特异性的生物标志物在结直肠癌的辅助诊断中发挥重要作用。
Description
技术领域
本发明属于生物技术领域，具体涉及一种结直肠癌诊断生物标志物MDK蛋白及其应用。
背景技术
结直肠癌(colorectal cancer，CRC)是最常见的消化系统恶性肿瘤之一，也是目前致死率最高的肿瘤之一。世界卫生组织调查数据显示，每年约有120万新发结直肠癌病例，其中死亡病例约61万。随着我国经济的迅速发展，环境污染及饮食习惯改变问题的凸显，结直肠癌发病率亦呈逐年迅速上升的趋势。最新数据显示，我国2015年新发结直肠癌病例37.6万，死亡病例19.1万，发病率和死亡率居所有肿瘤的第5位。近年来，包括手术切除、放化疗及新辅助治疗在内的结直肠癌治疗手段都取得了显著进步，但其5年生存率仍徘徊在50％左右。因此，开发一种可以在结直肠癌诊断和治疗中具有应用价值的生物标志物，对结直肠癌的诊断和治疗具有重大的临床价值和意义。
中期因子(Midkne，MDK)是1988年由日本学者Kadomasu等发现的一种新型肝素结合生长因子(HBGF)，与多效生长因子(Pleiotrophin，PTN)共同构成新型肝素蛋白家族。MDK因最初被发现在妊娠中期小鼠胚胎肾脏组织中表达广泛而得名，但出生后表达水平则明显降低。研究发现MDK具有诱导细胞恶性转化、抗细胞凋亡、促血管生成等促进肿瘤生长和演进的作用，其在肿瘤领域的作用越来越引起研究者们的关注。MDK无论在结直肠癌组织中呈高表达，与结直肠癌的恶性程度及预后密切相关，有可能成为一种新型结直肠癌诊断的生物标志物。
本发明旨在利用MDK蛋白作为生物标志物可以用于结直肠癌的诊断，并以MDK为靶点设计治疗结直肠癌的新药。
发明内容
本发明所要解决的技术问题是提供一种可用于结直肠癌诊断的MDK蛋白生物标志物，MDK蛋白生物标志物与结直肠癌发病率相关，在结直肠癌组织中的表达高于癌旁组织，差异具有显著生物统计学意义。
本发明解决上述技术问题所采用的技术方案为：一种用于结直肠癌诊断的MDK蛋白生物标志物，所述MDK蛋白生物标志物可用于检测肿瘤患者组织中的MDK蛋白的表达。
一种可用于结直肠癌诊断的MDK蛋白生物标志物的应用，该结直肠癌诊断的MDK蛋白生物标志物可用于结直肠癌辅助诊断。
进一步地，
(1)结直肠癌患者癌组织与癌旁组织MDK蛋白表达的检测；评估其作为结直肠癌诊断生物标志物，用于结直肠癌辅助诊断的可能性；
(2)验证MDK蛋白表达与结直肠癌患者预后的相关性。
本发明的优点在于：本发明首次公开了可用于结直肠癌诊断的MDK蛋白生物标志物及其应用，研究表明MDK蛋白生物标志物在结直肠癌组织中呈高表达，与结直肠癌的发生呈正相关，并且参与调控结直肠癌的预后，是潜在的结直肠癌诊断的生物标志物。MDK蛋白生物标志物的开发，有望成为结直肠癌辅助诊断和靶向治疗的一种新手段。
附图说明
图1为MDK蛋白在结直肠癌组织及癌旁组织中表达水平。
图2为MDK蛋白在结直肠癌组织中表达的ROC曲线。
图3为MDK表达量与结直肠癌患者预后相关的生存曲线分析。
具体实施方式
以下结合附图实施例对本发明作进一步详细描述。
实施例：
免疫组织化学法检测MDK蛋白在结直肠癌组织及癌旁组织中表达水平
(1)样本收集
70例原发结直肠癌患者的手术切除癌组织和癌旁组织的组织芯片来源于上海国家生物芯片工程中心生物库(上海芯超生物科技有限公司)，组织芯片号为HColA180Su24。所有癌组织均术后病理证实为结直肠癌，全部结直肠癌患者术前均未行放、化疗，全部病例临床资料完整。
(2)免疫组织化学检测MDK蛋白的表达
将组织芯片从冰箱中取出在65℃下加热约1小时，以融化密封蜡。脱蜡过程：将载玻片在二甲苯中浸泡两次各10分钟，转移浸泡到无水乙醇中8分钟，再到95％乙醇中浸泡5分钟，最后在70％乙醇中浸泡10分钟。在室温下，用柠檬酸盐缓冲液(pH 6.0)进行抗原修复30分钟。载玻片与抗MDK兔单克隆抗体孵育1小时，然后与HRP标记的二抗抗体孵育1小时。组织芯片由病例组织专用扫描扫描系统扫描后进行分析。MDK抗体的染色强度评分为1(弱)，2(中)，3(强)，MDK蛋白表达阳性的百分比分数为5～100％。分析结果显示，MDK蛋白在结直肠癌组织中的表达显著高于癌旁组织(结果见图1)。
(3)MDK蛋白在结直肠癌中表达的ROC曲线绘制
根据结直肠癌患者癌组织和癌旁组织中MDK抗体染色的强度和阳性率计算出MDK蛋白表达的积分，以MDK蛋白表达的积分为依据，使用SPSS21.0软件计算MDK蛋白在结直肠癌中表达的ROC曲线，结果MDK蛋白在结直肠癌患者组织中的AUC为0.676(置信区间0.592-0.759)，敏感性为48.6％，特异性为85.7％(结果见图2)。
(4)结直肠癌组织中MDK表达与预后的相关性分析
根据TCGA数据库中MDK基因在结直肠癌组织中的表达情况，结合其患者的临床资料，对结直肠癌组织中MDK的表达与患者预后数据进行分析，结果MDK基因表达量越高，结直肠癌患者生存期越短，预后也越差(结果见图3)。
以上所述，仅为本发明较佳的具体实施方式，但本发明的保护范围并不局限于此，任何熟悉本技术领域的技术人员在本发明揭露的技术范围内，根据本发明的技术方案及其发明构思加以等同替换或改变，都应涵盖在本发明的保护范围之内。
Claims (3)
1.一种MDK蛋白作为生物标志物在结直肠癌诊断中的应用。
2.如权利要求1所述的结直肠癌生物标志物MDK蛋白的特异性抗体，其特征为：所述MDK蛋白的抗体为重组抗MDK兔单克隆抗体。
3.如权利要求2所述的结直肠癌生物标志物MDK蛋白的特异性兔单克隆抗体，其特征为：所述的检测MDK蛋白的产品包括用于检测结直肠癌及癌旁组织MDK蛋白表达的免疫组织化学相关试剂产品。
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