Genome-wide discovery and validation of diagnostic DNA methylation-based biomarkers for hepatocellular cancer detection in circulating cell free DNA
Genome-wide discovery and validation of diagnostic DNA methylation-based biomarkers for hepatocellular cancer detection in circulating cell free DNA
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DOI：
10.7150/thno.35573
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发表时间：
2019-01-01
期刊：
影响因子：
12.4
通讯作者：
Robertson, Keith D.
中科院分区：
文献类型：
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作者：
Hlady, Ryan A.;Zhao, Xia;Robertson, Keith D.
肝细胞癌是最常见的一种肝癌，其发病率正在上升，但治疗选择仍然有限，尤其是对晚期疾病。由于肝硬变是肝细胞癌发展的主要危险状态，迫切需要标记物来检测这一患者群体中的早期肝细胞癌。表观遗传标记的扰动，如DNA甲基化(5mC)，是包括肝癌在内的人类癌症的一个特征。因此，在从肝硬变到肝细胞癌的发展过程中，识别循环中循环游离DNA(CfDNA)中持续变化的5mC水平的区域可以作为发展早期肝细胞癌微创筛查和监测的标志。方法：为了在肝硬变的背景下发现肝细胞癌的DNA甲基化衍生生物标志物，我们使用Infinium HumanMylation450k珠芯片阵列描绘了患者cfDNA的全基因组5mC景观。我们进一步将这些发现与TCGA和其他公共来源提供的原始组织数据联系起来。利用生物学和统计学框架，我们选择了在原始组织和cfDNA中强有力地区分肝硬变和肝癌的CPG，然后在另一个独立队列中进行验证。结果：通过对cfDNA 5mC图谱的全基因组分析，我们确定了将肝硬变患者与肝硬变背景下的肝癌患者区分开来的CPG。套索回归分析精确地确定了我们发现队列中的一组探针，这些探针在两个独立的数据集中得到了验证。由五个CPG(CG04645914、CG06215569、CG23663760、CG13781744和CG07610777)组成的面板在CFDNA发现和组织验证队列1和2中分别产生了0.9525、0.9714和0.9528的接收者工作特征(AUROC)曲线下的面积。将高甲基化和低甲基化的CPGS组合成一个独立的CFDNA，通过亚硫酸氢盐焦磷酸测序得到AUROC为0.956，而发现的AUROC为0.996。结论：与直接从CFDNA鉴定的标记相比，我们的发现来自原发组织的5mC标记在CFDNA中的表现不佳，揭示了从CFDNA开始寻找液体活检发展的高性能标记的潜在优势。
Hepatocellular carcinoma (HCC), the most prevalent form of liver cancer, is growing in incidence but treatment options remain limited, particularly for late stage disease. As liver cirrhosis is the principal risk state for HCC development, markers to detect early HCC within this patient population are urgently needed. Perturbation of epigenetic marks, such as DNA methylation (5mC), is a hallmark of human cancers, including HCC. Identification of regions with consistently altered 5mC levels in circulating cell free DNA (cfDNA) during progression from cirrhosis to HCC could therefore serve as markers for development of minimally-invasive screens of early HCC diagnosis and surveillance.Methods: To discover DNA methylation derived biomarkers of HCC in the background of liver cirrhosis, we profiled genome-wide 5mC landscapes in patient cfDNA using the Infinium HumanMethylation450k BeadChip Array. We further linked these findings to primary tissue data available from TCGA and other public sources. Using biological and statistical frameworks, we selected CpGs that robustly differentiated cirrhosis from HCC in primary tissue and cfDNA followed by validation in an additional independent cohort.Results: We identified CpGs that segregate patients with cirrhosis, from patients with HCC within a cirrhotic liver background, through genome-wide analysis of cfDNA 5mC landscapes. Lasso regression analysis pinpointed a panel of probes in our discovery cohort that were validated in two independent datasets. A panel of five CpGs (cg04645914, cg06215569, cg23663760, cg13781744, and cg07610777) yielded area under the receiver operating characteristic (AUROC) curves of 0.9525, 0.9714, and 0.9528 in cfDNA discovery and tissue validation cohorts 1 and 2, respectively. Validation of a 5-marker panel created from combining hypermethylated and hypomethylated CpGs in an independent cfDNA set by bisulfite pyrosequencing yielded an AUROC of 0.956, compared to the discovery AUROC of 0.996.Conclusion: Our finding that 5mC markers derived from primary tissue did not perform well in cfDNA, compared to those identified directly from cfDNA, reveals potential advantages of starting with cfDNA to discover high performing markers for liquid biopsy development.