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Treatment monitoring of colorectal cancer by integrated analysis of plasma concentration and sequencing of circulating tumor DNA

ABSTRACT: Circulating cell-free DNA (cfDNA) analysis is an important tool for cancer monitoring. The patient-specific mutations identified in colorectal cancer (CRC) tissues are usually used to design the cfDNA analysis. Despite high specificity in predicting relapse, the sensitivity in most studies is around 40–50%. To improve this weakness, we designed a cfDNA panel according to the CRC genomic landscape and recurrent-specific mutations. The pathological variants in cfDNA samples from 60 CRC patients were studied by a next-generation sequencing (NGS) method incorporating the dual molecular barcode. Interestingly, patients in the disease positive group had a significantly higher cfDNA concentration than those in the disease negative group. Based on receiver operating characteristic analysis, the cfDNA concentration of 7?ng/mL was selected into the analytical workflow. The sensitivity in determining the disease status was 72.4%, which represented a considerable improvement on prior studies, and the specificity remained high at 80.6%. Compared to standard imaging and laboratory studies, earlier detection of residual disease and clinical benefits were shown on two cases by this cfDNA assay. We conclude this integrative framework of cfDNA analytical pipeline with a satisfactory sensitivity and specificity could be used in postoperative CRC surveillance. Supplementary Information Supplementary information accompanies this paper at 10.1186/s12943-020-01273-8.

SUBMITTER: Yeh Y

PROVIDER: S-EPMC7586655 | biostudies-literature | 2020 Jan

REPOSITORIES: biostudies-literature

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Project description:Postoperative monitoring for patients with colorectal cancer (CRC) requires sensitive biomarkers that are associated with medical response and adjuvant therapy following surgery. Conventional tumor biomarkers [including carcinoembryonic antigen (CEA), CA19-9 and CA125] are widely used, but none of the markers provide high sensitivity or specificity. Previous studies indicated that circulating tumor DNA (ctDNA) is useful for postoperative monitoring of patients with cancer. However, the majority of previous studies involved patients with lung cancer, and therefore further studies are required which investigate patients with CRC. The present study enrolled 11 patients with CRC. All patients underwent surgery, and a number of patients were treated with postoperative chemotherapy. Tumor tissues and serial blood samples were collected from each patient, and somatic mutations of each sample were obtained using next-generation sequencing. The mutation landscape and dynamic changes in mutations for each patient were analyzed, and these results were compared with the changes of CEA levels. A number of driver genes were selected, including tumor protein P53 (TP53), APC and KRAS, to monitor the postoperative outcome of the 11 patients with CRC. Driver mutations were detected in preoperative plasma in 7 patients, with markedly decreased mutation rates detected in postoperative plasma compared with preoperative plasma. Driver mutations were not detected in 4 patients in the preoperative or postoperative plasma. In 1 patient with metastatic rectal cancer, the rate of TP53 mutation increased from 8.95 (preoperative) to 71.4% (postoperative), and a new phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit α mutation emerged. This patient succumbed to mortality six months following surgery, however there were no marked changes in CEA levels during periodic detection of CEA levels. In summary, ctDNA has a high sensitivity and specificity in prediction of the prognosis of patients with CRC.

Project description:BackgroundCirculating tumor DNA (ctDNA) carries information on tumor burden. However, the mutation spectrum is different among tumors. This study was designed to examine the utility of ctDNA for monitoring tumor burden based on an individual mutation profile.MethodologyDNA was extracted from a total of 176 samples, including pre- and post-operational plasma, primary tumors, and peripheral blood mononuclear cells (PBMC), from 44 individuals with colorectal tumor who underwent curative resection of colorectal tumors, as well as nine healthy individuals. Using a panel of 50 cancer-associated genes, tumor-unique mutations were identified by comparing the single nucleotide variants (SNVs) from tumors and PBMCs with an Ion PGM sequencer. A group of the tumor-unique mutations from individual tumors were designated as individual marker mutations (MMs) to trace tumor burden by ctDNA using droplet digital PCR (ddPCR). From these experiments, three major objectives were assessed: (a) Tumor-unique mutations; (b) mutation spectrum of a tumor; and (c) changes in allele frequency of the MMs in ctDNA after curative resection of the tumor.ResultsA total of 128 gene point mutations were identified in 27 colorectal tumors. Twenty-six genes were mutated in at least 1 sample, while 14 genes were found to be mutated in only 1 sample, respectively. An average of 2.7 genes were mutated per tumor. Subsequently, 24 MMs were selected from SNVs for tumor burden monitoring. Among the MMs found by ddPCR with > 0.1% variant allele frequency in plasma DNA, 100% (8 out of 8) exhibited a decrease in post-operation ctDNA, whereas none of the 16 MMs found by ddPCR with < 0.1% variant allele frequency in plasma DNA showed a decrease.ConclusionsThis panel of 50 cancer-associated genes appeared to be sufficient to identify individual, tumor-unique, mutated ctDNA markers in cancer patients. The MMs showed the clinical utility in monitoring curatively-treated colorectal tumor burden if the allele frequency of MMs in plasma DNA is above 0.1%.

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Treatment monitoring of colorectal cancer by integrated analysis of plasma concentration and sequencing of circulating tumor DNA

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