CN114414806A - 用于胆囊癌诊断的标志物及其应用 - Google Patents
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Abstract
本发明公开了一种用于胆囊癌诊断的标志物及其应用，所述标志物选自3‑磷酸甘油醛脱氢酶和人乙酰辅酶A脱氢酶中的至少一种。本发明是首次发现3‑磷酸甘油醛脱氢酶，特别是3‑磷酸甘油醛脱氢酶与人乙酰辅酶A脱氢酶的组合能够作为诊断胆囊癌的初步诊断标志物，具有较高的检测特异性。
Description
技术领域
本发明属于医学检测领域，具体涉及用于胆囊癌诊断的标志物及其应用，其用于构造诊断胆囊癌的试剂或试剂盒，属于医用检查检验服务领域，特别是体外诊断检测中的体外诊断用试剂、试纸及其配套设备与耗材领域。
背景技术
胆囊是人体中一个重要的消化器官，它不仅有储存、浓缩、排泄(收缩)功能，还有调节肝内外胆道压力的重要作用以及重要的分泌和免疫作用。胆囊癌(gallbladdercancer，简称GBC)是指发生于胆囊(包括胆囊底部、体部、颈部及胆囊管)的恶性肿瘤，占胆道恶性肿瘤的80％-95％。胆囊癌具有隐匿性、易侵袭转移、早期诊断难、预后差等特点。我国胆囊癌发病率占同期胆道疾病的0.4％-3.8％，居消化道肿瘤第6位。在过去的几十年中，胆囊癌治疗策略未取得实质性进展，且因缺乏典型症状，早期诊断率仅有19.1％，多数患者在被明确诊断时已处于中晚期阶段，有50％的患者被发现时已出现淋巴结转移。目前为止，手术切除仍然是胆囊癌患者的唯一治疗手段。然而手术切除对于晚期患者的治疗意义不大。胆囊癌对放疗、化疗等治疗措施均不敏感，故整体预后极差，胆囊癌患者5年总体生存率仅为5％。除此之外，术后易复发或者转移很大程度上决定了晚期胆囊癌的不良预后。胆囊癌的发病机制主要与多种基因以及环境因素相关，慢性胆道系统感染、特殊化学物质、重金属的环境暴露、饮食因素等，都与胆囊癌的形成有关。雌激素、胆固醇循环及沙门杆菌的感染，是造成胆囊癌在发展中国家高发病率的主要原因之一。尽管在胆囊癌分子生物学发病机制方面取得了显著进展，基本澄清了如PI3K-AKT-mTOR、Notch、ErbB、MAPK-ERK等信号通路及基因在胆囊癌中起到的作用，然而，上述研究尚不能改变胆囊癌的治疗与治愈途径，探究用于确诊胆囊癌的早期检测的生物标志物仍旧在胆囊癌治疗中具有至关重要的地位。
中国授权专利CN108931633B公开了胆囊癌诊断和预后判断标志物PIM1，还公开了胆囊癌诊断和预后判断的试剂盒，其可用于诊断、预测、检测或筛查人胆囊癌细胞扩散、胆囊癌临床分期或胆囊癌患者预后。该专利还公开了PIM1蛋白抑制剂在制备抗胆囊癌药物中的用途。该专利研究结果表明PIM1在临床实践中有非常重要的意义，有可能成为新一代的肿瘤生物诊断标志物和用于肿瘤治疗的分子靶标，开辟了以PIM1为主的胆囊癌疾病全新靶标的研究，也为研制抗肿瘤新药开辟新的途径。
中国专利申请CN111455049A公开了一种胆囊癌的诊断标志物，所述标志物为LncRNAMEG3和MALAT1。该专利申请通过生物信息学和现有的分子生物学的技术手段，从胆囊癌组织中筛选到用于胆囊癌诊断的标志物LncRNA MEG3和MALAT1(LncRNA MEG3在胆囊癌中低表达，MALAT1在胆囊癌中高表达)，为胆囊癌的治疗提供指导和帮助。
然而，暂未有数据公开上述标志物在胆囊癌诊断中的效果。如何发掘出更多能够用于胆囊癌针对的标志物，仍然是本领域技术人员亟待解决的问题之一。
发明内容
本发明的目的在于提供一种用于胆囊癌诊断的标志物，以解决现有技术中存在的问题。
3-磷酸甘油醛脱氢酶(Glyceraldehyde-3-phosphate dehydrogenase，GAPD)广布于人体各组织器官，已知可以催化3-磷酸甘油醛氧化(脱氢)和磷酸化，生成1，3-二磷酸甘油酸，是糖代谢的中心环节，并用于糖代谢有问题的患者的病情缓解及治疗。
人乙酰辅酶A脱氢酶已被商品化用于检测，但未见其在胆囊癌诊断检测中的应用。
不局限于任何理论，本发明人出人意料地发现，3-磷酸甘油醛脱氢酶和乙酰辅酶A脱氢酶在胆囊癌患者中的特征性存在，并由此完成了本发明。根据本发明的实施方式提供如下技术方案：
本发明一方面涉及一种用于胆囊癌诊断的标志物，所述标志物选自3-磷酸甘油醛脱氢酶和人乙酰辅酶A脱氢酶中的至少一种。
在本发明的一个优选实施方式中，所述标志物为3-磷酸甘油醛脱氢酶和人乙酰辅酶A脱氢酶的组合。
在本发明的一个优选实施方式中，所述3-磷酸甘油醛脱氢酶和所述人乙酰辅酶A脱氢酶是指血清中的3-磷酸甘油醛脱氢酶和人乙酰辅酶A脱氢酶。
本发明另一方面还涉及一种诊断胆囊癌的试剂盒，所述试剂盒包括检测人血清中3-磷酸甘油醛脱氢酶和人乙酰辅酶A脱氢酶浓度的试剂。
在本发明的一个优选实施方式中，所述检测人血清中3-磷酸甘油醛脱氢酶和人乙酰辅酶A脱氢酶浓度的试剂分别是指3-磷酸甘油醛脱氢酶ELISA试剂和人乙酰辅酶A脱氢酶ELISA试剂。
本发明又另一方面还涉及上述标志物和上述试剂盒在制备诊断胆囊癌的试剂中的应用。
有益效果
与现有技术相比，本发明至少实现了如下有益效果：
本发明是首次发现3-磷酸甘油醛脱氢酶，特别是3-磷酸甘油醛脱氢酶与人乙酰辅酶A脱氢酶的组合能够作为诊断胆囊癌的初步诊断标志物，具有较高的检测特异性，为早期诊断胆囊癌提供了快速、有效的新选择。
附图说明
图1为示出胆囊癌患者与健康人群的血清中3-磷酸甘油醛脱氢酶的浓度比较的图。
图2为示出胆囊癌患者与健康人群的血清中人乙酰辅酶A脱氢酶的浓度比较的图。
图3为示出3-磷酸甘油醛脱氢酶与人乙酰辅酶A脱氢酶组合作为胆囊癌诊断的ROC图。
具体实施方式
为了进一步理解本发明，下面将结合本发明实施例，对本发明实施例中的技术方案进行清楚、完整地描述，显然，所描述的实施例仅仅是本发明一部分实施例，而不是全部的实施例。基于本发明中的实施例，本领域普通技术人员在没有做出创造性劳动前提下所获得的所有其他实施例，都属于本发明保护的范围。
如无特殊说明，本发明实施例中所涉及的试剂均为市售产品，均可以通过商业渠道购买获得。
实施例1
样品采集
40位胆囊癌患者均来自河南省肿瘤医院，就诊时间为2020年6月至12月，患者年龄为40-50岁，平均年龄为46.8岁，男22例，女18例。另采集健康人群18例的全血样本，平均年龄46.3岁，男10例，女8例。
取每位受试者全血标本，室温放置2小时内3000g离心5分钟，吸取上清液转移至无菌EP管，分装后于-80℃保存至检测前。
试验方法与结果
采用ELISA试剂盒方法检测受试者血清中3-磷酸甘油醛脱氢酶与人乙酰辅酶A脱氢酶的含量。3-磷酸甘油醛脱氢酶ELISA检测试剂盒来自上海雅吉生物科技有限公司，其操作按照试剂盒说明书进行。人乙酰辅酶A脱氢酶ELISA试剂盒来自上海颖心实验室设备有限公司，其操作按照试剂盒说明书进行。
检测结果如图1和图2所示，结果为：健康组血清中3-磷酸甘油醛脱氢酶的平均相对浓度为100±27.4％，胆囊癌患者组血清中3-磷酸甘油醛脱氢酶的平均相对浓度为176.6±22.7％，健康组和胆囊癌患者组血清中3-磷酸甘油醛脱氢酶的浓度具有显著性差异。同样的，健康组血清中人乙酰辅酶A脱氢酶的平均相对浓度为100±22.3％，胆囊癌患者组血清中人乙酰辅酶A脱氢酶的平均相对浓度为150.6±21.8％，健康组和胆囊癌患者组血清中人乙酰辅酶A脱氢酶的浓度具有显著性差异。
上述实验结果表明，3-磷酸甘油醛脱氢酶与人乙酰辅酶A脱氢酶都显著高于健康人群，表明在胆囊癌患者中3-磷酸甘油醛脱氢酶与人乙酰辅酶A脱氢酶可以作为诊断标志物。
实施例2
另外选取2021年6-12月来河南省肿瘤医院就诊的胆囊癌患者的100例，基于血清中3-磷酸甘油醛脱氢酶与人乙酰辅酶A脱氢酶的含量确定AUC值，结果如下表1所示，其中3-磷酸甘油醛脱氢酶与人乙酰辅酶A脱氢酶组合作为诊断标志物的ROC曲线如图3所示。
表1：胆囊癌患者诊断AUC值
| 标志物 | AUC值 |
| 3-磷酸甘油醛脱氢酶 | 0.635 |
| 人乙酰辅酶A脱氢酶 | 0.574 |
| 3-磷酸甘油醛脱氢酶+人乙酰辅酶A脱氢酶 | 0.763 |
结果显示，3-磷酸甘油醛脱氢酶单独作为胆囊癌诊断标志物的AUC值为0.635，人乙酰辅酶A脱氢酶单独作为胆囊癌诊断标志物的AUC值为0.574。而3-磷酸甘油醛脱氢酶与人乙酰辅酶A脱氢酶组合作作为标志物进行联合诊断的AUC值为0.763。由此说明，3-磷酸甘油醛脱氢酶与人乙酰辅酶A脱氢酶组合作作为标志物应用于胆囊癌的诊断具有较高的准确性。
此外，应当理解，虽然本说明书按照实施方式加以描述，但并非每个实施方式仅包含一个独立的技术方案，说明书的这种叙述方式仅仅是为清楚起见，本领域技术人员应当将说明书作为一个整体，各实施例中的技术方案也可以经适当组合，形成本领域技术人员可以理解的其他实施方式。
以上所述仅为本发明的较佳实施例，并不用以限制本发明，凡在本发明的精神和原则之内，所作的任何修改、等同替换、改进等，均应包含在本发明的保护范围之内。
Claims (7)
1.一种用于胆囊癌诊断的标志物，其特征在于，所述标志物选自3-磷酸甘油醛脱氢酶和人乙酰辅酶A脱氢酶中的至少一种。
2.根据权利要求1所述的标志物，其特征在于，所述标志物为3-磷酸甘油醛脱氢酶和人乙酰辅酶A脱氢酶的组合。
3.根据权利要求1所述的标志物，其特征在于，所述3-磷酸甘油醛脱氢酶和所述人乙酰辅酶A脱氢酶是血清中的3-磷酸甘油醛脱氢酶和人乙酰辅酶A脱氢酶。
4.根据权利要求1-3中任意一项所述的标志物在制备诊断胆囊癌的试剂中的应用。
5.一种诊断胆囊癌的试剂盒，其特征在于，所述试剂盒包括检测血清中的3-磷酸甘油醛脱氢酶和人乙酰辅酶A脱氢酶浓度的试剂。
6.根据权利要求5所述的试剂盒，其特征在于，所述试剂分别是3-磷酸甘油醛脱氢酶ELISA试剂和人乙酰辅酶A脱氢酶ELISA试剂。
7.根据权利要求5或6所述的试剂盒在制备诊断胆囊癌的试剂中的应用。
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