Department of Oncology, Johns Hopkins University School of Medicine, Baltimore, MD.
Cepheid, Sunnyvale, CA.
Cancer Res Commun. 2022 Jun;2(6):391-401. doi: 10.1158/2767-9764.crc-22-0133. Epub 2022 Jun 1.
Current molecular liquid biopsy assays to detect recurrence or monitor response to treatment require sophisticated technology, highly trained personnel, and a turnaround time of weeks. We describe the development and technical validation of an automated Liquid Biopsy for Breast Cancer Methylation (LBx-BCM) prototype, a DNA methylation detection cartridge assay that is simple to perform and quantitatively detects nine methylated markers within 4.5 h. LBx-BCM demonstrated high interassay reproducibility when analyzing exogenous methylated DNA (75-300 DNA copies) spiked into plasma (Coefficient of Variation, CV = 7.1 - 10.9%) and serum (CV = 19.1 - 36.1%). It also demonstrated high interuser reproducibility (Spearman = 0.887, < 0.0001) when samples of metastatic breast cancer (MBC, = 11) and normal control ( = 4) were evaluated independently by two users. Analyses of interplatform reproducibility indicated very high concordance between LBx-BCM and the reference assay, cMethDNA, among 66 paired plasma samples (MBC = 40, controls = 26; Spearman = 0.891; 95% CI = 0.825 - 0.933, < 0.0001). LBx-BCM achieved a ROC AUC = 0.909 (95% CI = 0.836 - 0.982), 83% sensitivity and 92% specificity; cMethDNA achieved a ROC AUC = 0.896 (95% CI = 0.817 - 0.974), 83% sensitivity and 92% specificity in test set samples. The automated LBx-BCM cartridge prototype is fast, with performance levels equivalent to the highly sensitive, manual cMethDNA method. Future prospective clinical studies will evaluate LBx-BCM detection sensitivity and its ability to monitor therapeutic response during treatment for advanced breast cancer.
目前，用于检测复发或监测治疗反应的分子液体活检检测需要先进的技术、经过专业培训的人员以及数周的周转时间。我们描述了一种自动化乳腺癌甲基化液体活检（LBx-BCM）原型的开发和技术验证，该原型是一种简单易用的 DNA 甲基化检测试剂盒检测方法，可在 4.5 小时内定量检测 9 个甲基化标志物。LBx-BCM 在分析外源性甲基化 DNA（75-300 个 DNA 拷贝）掺入血浆（变异系数 CV = 7.1-10.9%）和血清（CV = 19.1-36.1%）时表现出良好的组间重现性。当由两名用户独立评估转移性乳腺癌（MBC，n = 11）和正常对照（n = 4）的样本时，它还表现出良好的组间重现性（Spearman = 0.887，<0.0001）。在对 66 对血浆样本（MBC = 40，对照 = 26；Spearman = 0.891；95%置信区间= 0.825-0.933，<0.0001）进行平台间重现性分析时，LBx-BCM 与参考检测方法 cMethDNA 之间的一致性非常高。LBx-BCM 的 ROC AUC = 0.909（95%置信区间= 0.836-0.982），具有 83%的灵敏度和 92%的特异性；cMethDNA 在测试集样本中达到 ROC AUC = 0.896（95%置信区间= 0.817-0.974），83%的灵敏度和 92%的特异性。自动化的 LBx-BCM 试剂盒原型速度很快，性能水平与高度敏感的手动 cMethDNA 方法相当。未来的前瞻性临床研究将评估 LBx-BCM 的检测灵敏度及其在晚期乳腺癌治疗期间监测治疗反应的能力。
