CN111190008B - 血清mpo-dna在预测乳腺癌预后抑制乳腺癌远处器官转移中的应用 - Google Patents

血清mpo-dna在预测乳腺癌预后抑制乳腺癌远处器官转移中的应用 Download PDF

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CN111190008B
CN111190008B CN202010014581.XA CN202010014581A CN111190008B CN 111190008 B CN111190008 B CN 111190008B CN 202010014581 A CN202010014581 A CN 202010014581A CN 111190008 B CN111190008 B CN 111190008B
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宋尔卫
苏士成
杨林槟
陈嘉宁
张晓倩
李嘉倩
李和亮
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Abstract

本发明公开了血清MPO‑DNA复合物在预测乳腺癌预后或抑制乳腺癌远处器官转移的应用以及试剂盒，所述试剂盒包括用于检测乳腺癌患者中血清MPO‑DNA表达量的试剂；另外，MPO‑DNA抑制剂能抑制乳腺癌细胞的转移。本发明试剂盒辅助现有的预后诊断，以提高患者的生存率；本发明还提供一种新的乳腺癌治疗方案。

Description

血清MPO-DNA在预测乳腺癌预后抑制乳腺癌远处器官转移中 的应用
技术领域
本发明属于生物医药技术领域，尤其涉及血清MPO-DNA在预测乳腺癌预后抑制乳腺癌远处器官转移中的应用。
背景技术
2018年国家癌症中心发布的最新全国癌症数据统计显示，在168.9万新发女性肿瘤患者中，乳腺癌患者有27.9万，约占16.5％。乳腺癌已曾为女性发病率最高的肿瘤。而在中国女性恶性肿瘤死亡病因中，乳腺癌排名第5位，仅次于肺癌、胃癌、肝癌及结直肠癌。远处器官转移是乳腺癌致死的重要原因，因此，如何有效地预测乳腺癌患者预后，早期进行干预是提高患者生存疗效的关键。
目前，治疗乳腺癌以放化疗为主，联合内分泌治疗及靶向治疗是晚期乳腺癌患者的主要治疗手段，根据ER、PR、HER2等指标的表达情况，可将乳腺癌分为不同的亚型，不仅在预后上差别加大，还直接影响了治疗方案的选择，如 ER、PR均为阳性的患者，内分泌治疗必不可少；而HER2阳性的患者，赫赛汀可在一定程度上为患者保驾护航。
目前，尽管KI67、ER、PR、HER-2等指标可以一定程度上反应肿瘤的生长情况，特别是ER、PR、HER2还用于乳腺癌的分型，并指导了治疗方案的选择，如是否加入内分泌治疗，是否联合HER-2单抗赫赛汀等。但在预测患者在病程中发生远处转移方面尚缺乏证据，更加无法谈及早期干预。因此，有必要针对乳腺癌远处器官转移设计出一种药物，以抑制乳腺癌细胞的转移。
发明内容
本发明的目的在于克服上述现有技术的不足之处而提供了血清MPO-DNA 在预测乳腺癌预后或抑制乳腺癌远处器官转移中的应用。由于MPO能与DNA 牢固结合形成复合物，有效保护DNA防止机体相关核酸酶的降解，从而能够稳定存在于患者的血清中并被检测到。本发明人通过检测MPO-DNA复合物的水平从而预测乳腺癌远处器官转移情况，辅助乳腺癌患者预后的早期干预。同时，制备了检测血清MPO-DNA的试剂盒，该试剂盒包括用于检测血清中MPO-DNA 表达量的试剂，试剂盒可以辅助现有的预后诊断，以提高患者的生存率；另外， MPO-DNA的抑制剂可以抑制乳腺癌细胞的转移。
为实现上述目的，本发明采取的技术方案为：一种用于乳腺癌预后诊断的试剂盒，包括用于检测血清MPO-DNA表达量的试剂。
作为上述技术方案的改进，所述试剂包括抗MPO单抗和酶标记的抗DNA 单抗。
作为上述技术方案的改进，所述酶标记为HRP标记。
作为上述技术方案的改进，所述试剂还包括入ABTS底物和1％胎牛血清。
另外，本发明还提供一种用于抑制乳腺癌的药物，包括MPO-DNA抑制剂。
作为上述技术方案的改进，还包括药学上可接受的辅料。
另外，本发明还提供所示试剂盒的一种检测方法，其包括以下步骤：
(1)收集乳腺癌患者的外周血，通过离心获得血清；
(2)用抗MPO单抗包被96孔板，然后将HRP标记的抗DNA单抗与血清室温孵育；
(3)加入底物孵育，在OD405处检测MPO-DNA的表达量并进行记录；通过SPSS软件分析相关数据。
本发明的有益效果：本发明提供血清MPO-DNA在预测乳腺癌预后、抑制乳腺癌远处器官转移中的应用以及试剂盒。本发明试剂盒主要用于检测乳腺癌患者乳腺癌细胞远处器官转移中MPO-DNA的表达量，MPO-DNA表达量与乳腺癌患者预后有着强烈相关性，MPO-DNA的表达水平可以作为乳腺癌患者预后的指标；本发明试剂盒具有以下优点：1)适用于所有乳腺癌患者；2)检测方便，检测指标与预后有着明显相关，检测指标单一，操作简单；3)所有试剂大多较为常见，成本控制在可行范围内。另外，本发明还提供了一种新的乳腺癌治疗方案：MPO-DNA抑制剂能抑制乳腺癌细胞的转移。
附图说明
图1显示MPO-DNA表达水平与乳腺癌预后的关系，其中纵坐标表示肝转移生存率；上方曲线样本均来源与MPO-DNA表达水平较低的乳腺癌患者；下方曲线样本均来源与MPO-DNA表达水平较高的乳腺癌患者；曲线括号后面数字表示样本的数量。
图2显示MPO-DNA预测乳腺癌肝转移发生的敏感性。
图3显示降低MPO-DNA表达可显著抑制乳腺癌细向肝组织转移；其中，图片模糊不清楚之处表示乳腺癌细胞的转移。
具体实施方式
为了更加简洁明了的展示本发明的技术方案、目的和优点，下面结合具体实施例及其附图对本发明做进一步的详细描述。
实施例1：血清MPO-DNA的表达与乳腺癌患者预后的相关性
收集医院乳腺癌患者的外周血3-5ml，将外周血置于室温1小时(时间过长则血小板等物质影响MPO-DNA的释放)，室温2500转离心10分钟即可以收集到澄清的上清液(血清)。用抗MPO单抗(serotec公司，货号0400-0002,ABD) 包被96孔板，4℃过夜后次日用1％胎牛血清封闭96孔板，将HRP标记的抗 DNA单抗(Roche公司，货号11774425001)及40μl血清同时加入96孔板，室温孵育2小时；加入ABTS底物孵育，检测OD405，即MPO-DNA的表达量；收集相对应患者的临床预后信息，例如疾病确诊时间，出现肝转移的时间等信息，分析MPO-DNA与乳腺癌患者远处器官转移的预后的相关性，以及在乳腺癌肝转移的预测作用。如图1所示，通过分析MPO-DNA的表达水平与乳腺癌肝转移生存情况，发现MPO-DNA复合物与预后明显相关：MPO-DNA表达越低，乳腺癌患者无肝转移的的生存时间越长；MPO-DNA表达越高，乳腺癌患者无肝转移的生存时间越短。如图2所示，使用MPO-DNA预测乳腺癌预后相关性的敏感性较高。
实施例2：靶向MPO-DNA可明显抑制肿瘤的肝转移
本实施例中的试验动物：C57BL/6小鼠购买于中山大学实验动物中心；PAD4 基因敲除鼠购买于Jackson公司。
在正常的C57BL/6小鼠以及PAD4基因敲除鼠(该鼠可以特异性减少组织以及血清血清MPO-DNA的产生)中，分别在脾脏注射1×106鼠源性乳腺癌细胞株E0771，1个月后于剖出小鼠肝组织，在小动物活体成像系统中检测小鼠肝组织转移灶大小。结果如图3所示，正常的C57BL/6小鼠(Wild type)明显的转移灶，而PAD4基因敲除鼠(PAD4-/-)中的转移灶明显较小，说明降低 MPO-DNA可以抑制肝转移发生。提示MPO-DNA可以作为乳腺癌转移的靶点。
以上所述实施例仅表达了本发明的几种实施方式，其描述较为具体和详细，但并不能因此而理解为对发明专利范围的限制。应当指出的是，对于本领域的普通技术人员来说，在不脱离本发明构思的前提下，还可以做出若干变形和改进，这些都属于本发明的保护范围。因此，本发明专利的保护范围应以所附权利要求为准。

Claims (4)

1.血清MPO-DNA复合物在制备用于预测乳腺癌患者无肝转移的生存时间的试剂盒中的应用，其特征在于，所述试剂盒包括用于收集乳腺癌患者血清的容器以及用于检测血清MPO-DNA表达量的试剂。
2.如权利要求1所述的应用，其特征在于，所述试剂包括抗MPO单抗和酶标记的抗DNA单抗。
3.如权利要求2所述的应用，其特征在于，所述酶标记为HRP标记。
4.如权利要求2或3所述的应用，其特征在于，所述试剂还包括ABTS底物和1%胎牛血清。
CN202010014581.XA 2020-01-07 2020-01-07 血清mpo-dna在预测乳腺癌预后抑制乳腺癌远处器官转移中的应用 Active CN111190008B (zh)

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