Multi-Purpose Utility of Circulating Plasma DNA Testing in Patients with Advanced Cancers

Tumor genomic instability and selective treatment pressures result in clonal disease evolution; molecular stratification for molecularly targeted drug administration requires repeated access to tumor DNA. We hypothesized that circulating plasma DNA (cpDNA) in advanced cancer patients is largely derived from tumor, has prognostic utility, and can be utilized for multiplex tumor mutation sequencing when repeat biopsy is not feasible. We utilized the Sequenom MassArray System and OncoCarta panel for somatic mutation profiling. Matched samples, acquired from the same patient but at different time points were evaluated; these comprised formalin-fixed paraffin-embedded (FFPE) archival tumor tissue (primary and/or metastatic) and cpDNA. The feasibility, sensitivity, and specificity of this high-throughput, multiplex mutation detection approach was tested utilizing specimens acquired from 105 patients with solid tumors referred for participation in Phase I trials of molecularly targeted drugs. The median cpDNA concentration was 17 ng/ml (range: 0.5–1600); this was 3-fold higher than in healthy volunteers. Moreover, higher cpDNA concentrations associated with worse overall survival; there was an overall survival (OS) hazard ratio of 2.4 (95% CI 1.4, 4.2) for each 10-fold increase in cpDNA concentration and in multivariate analyses, cpDNA concentration, albumin, and performance status remained independent predictors of OS. These data suggest that plasma DNA in these cancer patients is largely derived from tumor. We also observed high detection concordance for critical ‘hot-spot’ mutations (KRAS, BRAF, PIK3CA) in matched cpDNA and archival tumor tissue, and important differences between archival tumor and cpDNA. This multiplex sequencing assay can be utilized to detect somatic mutations from plasma in advanced cancer patients, when safe repeat tumor biopsy is not feasible and genomic analysis of archival tumor is deemed insufficient. Overall, circulating nucleic acid biomarker studies have clinically important multi-purpose utility in advanced cancer patients and further studies to pursue their incorporation into the standard of care are warranted.肿瘤基因组不稳定性与选择性治疗压力可引发克隆性疾病演进；为实施分子靶向药物给药而开展的分子分层（molecular stratification），需要反复获取肿瘤组织DNA。我们提出假说：晚期癌症患者的循环血浆DNA（circulating plasma DNA, cpDNA）主要源自肿瘤组织，具备预后价值，且在无法开展重复活检时，可用于多重肿瘤突变测序。我们采用Sequenom MassArray系统与OncoCarta检测面板进行体细胞突变谱分析。我们评估了同一患者不同时间点获取的配对样本：这些样本包括福尔马林固定石蜡包埋（formalin-fixed paraffin-embedded, FFPE）存档肿瘤组织（原发灶和/或转移灶）以及cpDNA。我们利用105例参与分子靶向药物I期临床试验的实体瘤患者的标本，验证了这种高通量多重突变检测方法的可行性、灵敏度与特异性。cpDNA浓度的中位数为17 ng/ml（范围：0.5~1600 ng/ml），较健康志愿者高出3倍。此外，cpDNA浓度越高，患者总生存期（overall survival, OS）越差：cpDNA浓度每升高10倍，总生存期的风险比为2.4（95%置信区间：1.4~4.2）；多变量分析显示，cpDNA浓度、白蛋白水平与体能状态仍是OS的独立预测因素。上述数据表明，此类癌症患者的血浆DNA主要源自肿瘤组织。我们还观察到，配对的cpDNA与存档肿瘤组织在关键‘热点’突变（KRAS、BRAF、PIK3CA）上的检测一致性较高，且存档肿瘤与cpDNA之间存在显著差异。当无法安全开展重复肿瘤活检，且存档肿瘤的基因组分析被认为不充分时，这种多重测序检测方法可用于晚期癌症患者的血浆体细胞突变检测。总体而言，循环核酸生物标志物研究在晚期癌症患者中具备临床重要的多用途应用价值，未来有必要开展进一步研究以推动其纳入标准诊疗流程。