The confounding effects of next-generation sequencing (NGS) noise on detection of low frequency circulating tumor DNA (ctDNA) without a priori knowledge of solid tumor mutations has limited the applications of circulating cell-free DNA (ccfDNA) in clinical oncology. Here, we use a 118 gene panel and leverage ccfDNA technical replicates to eliminate NGS-associated errors while also enhancing detection of ctDNA from pancreatic ductal adenocarcinomas (PDACs). Pre-operative ccfDNA and tumor DNA were acquired from 14 patients with PDAC (78.6% stage II-III). Post-operative ccfDNA was also collected from 11 of the patients within 100 days of surgery. ctDNA detection was restricted to variants corresponding to pathogenic mutations in PDAC present in both replicates. PDAC-associated pathogenic mutations were detected in pre-operative ccfDNA in four genes (KRAS, TP53, SMAD4, ALK) from five patients. Of the nine ctDNA variants detected (variant allele frequency: 0.08%-1.59%), five had a corresponding mutation in tumor DNA. Pre-operative detection of ctDNA was associated with shorter survival (312 vs. 826 days; ${\chi }^2$=5.4, P = 0.021). Guiding ctDNA detection in pre-operative ccfDNA based on mutations present in tumor DNA yielded a similar survival analysis. Detection of ctDNA in the post-operative ccfDNA with or without tumor-informed guidance was not associated with outcomes. Therefore, the detection of PDAC-derived ctDNA during a broad and untargeted survey of ccfDNA with NGS may be a valuable, non-invasive, prognostic biomarker to integrate into the clinical assessment and management of patients prior to surgery.

下一代测序 (NGS) 噪声对检测低频循环肿瘤 DNA (ctDNA) 的混杂影响，无需先验对实体瘤突变的了解限制了循环游离 DNA (ccfDNA) 在临床肿瘤学中的应用。在这里，我们使用 118 个基因组并利用 ccfDNA 技术复制来消除与 NGS 相关的错误，同时还增强了胰腺导管腺癌 (PDAC) 中 ctDNA 的检测。从 14 名 PDAC 患者（78.6% II-III 期）获得术前 ccfDNA 和肿瘤 DNA。还在手术后 100 天内从 11 名患者中收集了术后 ccfDNA。ctDNA 检测仅限于与两个重复中存在的 PDAC 中的致病突变相对应的变体。在四个基因（ KRAS、TP53、SMAD4、ALK）的术前 ccfDNA 中检测到 PDAC 相关的致病突变) 来自五名患者。在检测到的 9 个 ctDNA 变异体中（变异等位基因频率：0.08%-1.59%），5 个在肿瘤 DNA 中有相应的突变。术前检测 ctDNA 与较短的生存期相关（312 天 vs. 826 天；${\chi }^2$=5.4，P  = 0.021）。根据肿瘤 DNA 中存在的突变指导术前 ccfDNA 中的 ctDNA 检测产生了类似的生存分析。在有或没有肿瘤知情指导的情况下，术后 ccfDNA 中 ctDNA 的检测与结果无关。因此，在使用 NGS 对 ccfDNA 进行广泛和非靶向调查期间检测 PDAC 衍生的 ctDNA 可能是一种有价值的、非侵入性的预后生物标志物，可整合到患者手术前的临床评估和管理中。