Abstract
BackgroundCirculating tumor DNA (ctDNA) offers a convenient way to monitor tumor progression and treatment response. Because tumor mutational profiles are highly variable from person to person, a fixed content panel may be insufficient to track treatment response in all patients.MethodsWe design ctDNA fingerprint panels specific to individual patients which are based on whole exome sequencing and target to high frequency clonal population clusters in patients. We test the fingerprint panels in 313 patients who together have eight tumor types (colorectal, hepatocellular, gastric, breast, pancreatic, and esophageal carcinomas and lung cancer and cholangiocarcinoma) and exposed to multiple treatment methods (surgery, chemotherapy, radiotherapy, targeted-drug therapy, immunotherapy, and combinations of them). We also monitor drug-related mutations in the patients using a pre-designed panel with eight hotspot genes.Results291 (93.0%) designed fingerprint panels harbor less than ten previously known tumor genes. We detected 7475 ctDNA mutations in 238 (76%) patients and 6196 (96.0%) of the mutations are detected in only one test. Both the level of ctDNA content fraction (CCF) and fold change of CCF (between the definitive and proceeding tests) are highly correlated with clinical outcomes (p-values 1.36e-6 for level and 5.64e-10 for fold change, Kruskal–Wallis test). The CCFs of PD patients are an order of magnitude higher than the CCFs of SD and OR patients (median/mean 2.22%/8.96% for SD, 0.18/0.21% for PD, and 0.31/0.54% for OR; pairwise p-values 7.8e-6 for SD ~ PD, 2.7e-4 for OR ~ PD, and 7.0e-3 for SD ~ OR, Wilcoxon rank sum test). The fold change of CCF distinguishes the patient groups even better, which increases for PD, remains stable for SD, and decreases for OR patients (p-values 0.002, ~ 1, and 0.0001 respectively, Wilcoxon signed-rank test). Eleven drug-related mutations are identified from nine out of the 313 patients.ConclusionsThe ctDNA fingerprint method improves both specificity and sensitivity of monitoring treatment response across several tumor types. It can identify tumor relapse/recurrence potentially earlier than imaging-based diagnosis. When augmented with tumor hotspot genes, it can track acquired drug-related mutations in patients.
Highlights
Circulating tumor DNA offers a convenient way to monitor tumor progression and treatment response
Validation of the individualized Circulating tumor DNA (ctDNA) panels in reference samples To determine the detection threshold and sensitivity of the ctDNA fingerprint panels, we performed more than 150 tests for each variant in 10 ng reference samples with total variant allelic frequencies at 15%, 10%, 6%, 3%, 1%, 0.5%, 0.25%, and 0% (See Materials and methods)
Our customized panels achieved a sensitivity of 99.99% when 10 singlenucleotide variants (SNVs) were used in the assessment and the detection threshold was set at 0.25% of the total circulating DNA for individual SNVs
Summary
Circulating tumor DNA (ctDNA) offers a convenient way to monitor tumor progression and treatment response. Circulating tumor DNA (ctDNA) mainly stem from apoptotic or necrotic tumor cells released into the circulation system. They provide accessible snapshots of tumor burden and mutation profile and allow noninvasive longitudinal sampling [1,2,3]. These properties make ctDNA a promising tool to monitor tumor progression and treatment response [4,5,6]. In a triple-negative breast cancer cohort, all of the metastatic patients were found to be ctDNA-positive [11]. Encouraged by these results, clinic trials have been opened to test the clinic utility of ctDNA in the settings of pre-cancer, minimal residual disease, recurrent and/or metastatic disease, which are reviewed in Cescon et al and Araujo et al [6, 12]
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