乳腺癌中血浆游离DNA含量和完整性的临床应用

doi:10.3969/j.issn.1673-8640.2017.08.003

摘要/Abstract

摘要：

目的探讨血浆循环游离脱氧核糖核酸（cfDNA）含量和完整性在乳腺癌诊断和预后监测中的临床意义。方法收集106例初诊乳腺癌患者血浆标本、72例乳腺良性腺病患者及301名体检健康者血浆标本,提取cfDNA,实时荧光定量聚合酶链反应（PCR）检测cfDNA中100 bp和241 bp长度的人端粒酶逆转录酶（hTERT）的含量,计算cfDNA完整性,分析3个组间差异及与肿瘤负荷的关系。应用受试者工作特征（ROC）曲线,分析100 bp和241 bp hTERT含量及完整性对乳腺癌诊断的灵敏度和特异度。结果 100 bp 和241 bp hTERT含量在乳腺癌患者中明显高于乳腺良性腺病患者和体检健康者,差异有统计学意义（P<0.001）,完整性与淋巴结转移相关（P<0.01）。ROC曲线表明100 bp、241 bp hTERT含量及完整性的曲线下面积（AUC）分别是0.693、0.792、0.865;三者敏感度分别为73.3%、68.0%、66.7%,特异度分别为68%、80%、92%。结论 cfDNA 含量和完整性的检测在乳腺癌的诊疗中有一定临床价值,可作为乳腺癌诊断和预后监测的指标。

关键词: 乳腺癌, 人端粒酶逆转录酶, 循环游离脱氧核糖核酸, 完整性

Abstract:

Objective To investigate the roles of plasma cell-free deoxyribonucleic acid（cfDNA）levels and its integrity in the diagnosis and prognosis of breast cancer. Methods Plasma samples were collected from 106 patients newly diagnosed as breast cancer,72 breast benign disease patients and 301 healthy subjects. Plasma cfDNA was extracted,and then 100 bp and 241 bp human telomerase reverse transcriptases（hTERT）were quantified by real-time fluorescence quantitation polymerase chain reaction（PCR）. The cfDNA integrity was calculated,and the difference among the 3 groups and the relationship with tumor burden were analyzed. The sensitivities and specificities of 100 bp and 241 bp hTERT levels and cfDNA integrity for the diagnosis of breast cancer were evaluated by receiver operating characteristic（ROC）curve. Results The levels of 100 bp and 241 bp hTERT in breast cancer patients were higher than those in breast benign disease patients and healthy subjects（P<0.001）. The integrity of cfDNA was related to lymph node metastasis（P<0.01）. The areas under ROC curves （AUC）of 100 bp and 241 bp hTERT levels and cfDNA integrity were 0.693,0.792 and 0.865. The sensitivities were 73.3%,68.0% and 66.7%,and the specificities were 68%,80% and 92%,respectively. Conclusions The determinations of cfDNA levels and its integrity have certain clinical significance in the diagnosis and treatment of breast cancer,which could be as tumor biomarkers for the diagnosis and prognosis of breast cancer.

Key words: Breast cancer, Human telomerase reverse transcriptase, Cell-free deoxyribonucleic acid, Integrity

中图分类号:

R446.1

顾益凤, 朱自力, 张金业, 张敏, 张小霞, 朱继文. 乳腺癌中血浆游离DNA含量和完整性的临床应用[J]. 检验医学, 2017, 32(8): 671-676.

GU Yifeng, ZHU Zili, ZHANG Jinye, ZHANG Min, ZHANG Xiaoxia, ZHU Jiwen. Clinical application of plasma cell-free deoxyribonucleic acid levels and its integrity in breast cancer[J]. Laboratory Medicine, 2017, 32(8): 671-676.

图/表 6

表1 相关基因的引物序列

引物名称	序列（5'~3'）	循环数	退火温度（℃）	产物长度
β-actin（GenBank#NM-001101）	Sense:ACCGAGCGCGGCTACAG	30	60	265 bp
	antisense:CTTAATGTCACGCACGATTTCC
hTERT（GenBank#NM-000005）	Sense:AACGACGCCGCTTCCTCA	35	52.6	100 bp
	Antisense:TCCCGCACGCTCATCTTCC
hTERT（GenBank#NM-000005）	Sense:GCAGAAGCAACAGGAAACCCAT	35	60	241 bp
	Antisense:TGAACCCAGGAGACGGAGG

图1 实时荧光定量PCR 扩增曲线

图2 241 bp和100 bp hTERT目的基因在各组血浆中的含量注：（a）为241 bp hTERT目的基因在各组血浆中的含量;（b）为100 bp hTERT目的基因在各组血浆中的含量

图3 各组血浆cfDNA完整性

图4 241 bp、100 bp hTERT及游离DNA完整性的ROC曲线分析

图5 有、无淋巴结转移乳腺癌患者血浆cfDNA完整性

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