Estimating the abundance of cell-free DNA (cfDNA) fragments shed from a tumor (i.e., circulating tumor DNA (ctDNA)) can approximate tumor burden, which has numerous clinical applications. We derived a novel, broadly applicable statistical method to quantify cancer-indicative methylation patterns within cfDNA to estimate ctDNA abundance, even at low levels. Our algorithm identified differentially methylated regions (DMRs) between a reference database of cancer tissue biopsy samples and cfDNA from individuals without cancer. Then, without utilizing matched tissue biopsy, counts of fragments matching the cancer-indicative hyper/hypo-methylated patterns within DMRs were used to determine a tumor methylated fraction (TMeF; a methylation-based quantification of the circulating tumor allele fraction and estimate of ctDNA abundance) for plasma samples. TMeF and small variant allele fraction (SVAF) estimates of the same cancer plasma samples were correlated (Spearman’s correlation coefficient: 0.73), and synthetic dilutions to expected TMeF of 10−3and 10−4had estimated TMeF within two-fold for 95% and 77% of samples, respectively. TMeF increased with cancer stage and tumor size and inversely correlated with survival probability. Therefore, tumor-derived fragments in the cfDNA of patients with cancer can be leveraged to estimate ctDNA abundance without the need for a tumor biopsy, which may provide non-invasive clinical approximations of tumor burden.
通过检测肿瘤释放至血液中的游离DNA片段（即循环肿瘤DNA）丰度，可近似评估肿瘤负荷，这一方法具有广泛的临床应用前景。本研究开发了一种新颖且普适性强的统计方法，通过量化游离DNA中具有癌症指示性的甲基化模式来估算循环肿瘤DNA丰度，即使在低浓度条件下仍能实现精准检测。该算法首先通过比对癌症组织活检样本参考数据库与健康个体游离DNA数据，识别出差异甲基化区域。随后，在不依赖配对组织活检的情况下，通过统计差异甲基化区域内符合癌症特异性高/低甲基化模式的片段数量，计算出血浆样本的肿瘤甲基化分数（TMeF）——这是一种基于甲基化水平的循环肿瘤等位基因分数定量指标，可用于评估循环肿瘤DNA丰度。对相同癌症血浆样本的分析显示，TMeF与小型变异等位基因分数估算值具有显著相关性（斯皮尔曼相关系数：0.73）。在预期TMeF为10^-3和10^-4的合成稀释实验中，分别有95%和77%的样本其TMeF估算值保持在两倍误差范围内。进一步研究发现，TMeF随癌症分期和肿瘤体积增加而升高，且与患者生存概率呈负相关。因此，通过分析癌症患者游离DNA中的肿瘤源性片段，可在无需肿瘤活检的情况下估算循环肿瘤DNA丰度，这为肿瘤负荷评估提供了非侵入性的临床检测新途径。
肿瘤(癌症)患者之家