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Heterogeneous mutation pattern in tumor tissue and circulating tumor DNA warrants parallel NGS panel testing.

ABSTRACT: Liquid biopsy by genotyping circulating tumor DNA (ctDNA) has provided a non-invasive approach in assessing tumor genomic alterations in clinical oncology. However, emerging evidence in clinical settings has shown significant discordance in the genomic alterations between matched tumor tissue and blood ctDNA samples, and even between the same set of blood samples analyzed on different testing platforms. Thus, it is necessary to study underlying causes of discrepancies in these studies by genotyping tumor tissue and ctDNA in parallel using next generation sequencing (NGS) panels based on the same technology. Here we enrolled 56 non-small-cell lung cancer (NSCLC) patients and evaluated tumor tissue genotyping and ctDNA based liquid biopsy by parallel NGS panel testing and compared different sample preparation conditions. Somatic mutations in plasma cell-free DNA (cfDNA) were detected in 63.6% patients with early-stage NSCLC and 60% patients with advanced-stage NSCLC. The overall concordance between matched formalin-fixed paraffin-embedded sample and cfDNA was 54.6% in early-stage NSCLC patients and 80% in advanced-stage NSCLC patients. The positive concordance rate was 44.4% and 71.4% in early-stage and advanced-stage patients, respectively. Using fresh frozen tumor samples did not improve the overall concordance rate between matched tumor tissue and cfDNA. Processing blood samples beyond 4 h after blood draw significantly decreased the detection rate of somatic mutations in cfDNA. Thus, the concordance rate between tumor tissue-based and ctDNA-based genotyping in clinical samples can be affected by multiple pre-analytical, analytical and biologic factors. Parallel NGS panel testing on both sample types for each patient may be warranted for effective guidance of cancer targeted therapies and possible early detection of cancer.

SUBMITTER: Guo Q

PROVIDER: S-EPMC6114875 | biostudies-other | 2018 Aug

REPOSITORIES: biostudies-other

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Heterogeneous mutation pattern in tumor tissue and circulating tumor DNA warrants parallel NGS panel testing.

Guo Qiaomei Q Wang Junlei J Xiao Jianfeng J Wang Lin L Hu Xiaomeng X Yu Wenjun W Song Gang G Lou Jiatao J Chen JianFeng J

Molecular cancer 20180828 1

Liquid biopsy by genotyping circulating tumor DNA (ctDNA) has provided a non-invasive approach in assessing tumor genomic alterations in clinical oncology. However, emerging evidence in clinical settings has shown significant discordance in the genomic alterations between matched tumor tissue and blood ctDNA samples, and even between the same set of blood samples analyzed on different testing platforms. Thus, it is necessary to study underlying causes of discrepancies in these studies by genotyp ...[more]

PMID: 30153823

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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%.

Project description:Addressing drug resistance is a core challenge in cancer research, but the degree of heterogeneity in resistance mechanisms in cancer is unclear. In this study, we conducted next-generation sequencing (NGS) of circulating tumor cells (CTC) from patients with advanced cancer to assess mechanisms of resistance to targeted therapy and reveal opportunities for precision medicine. Comparison of the genomic landscapes of CTCs and tissue metastases is complicated by challenges in comprehensive CTC genomic profiling and paired tissue acquisition, particularly in patients who progress after targeted therapy. Thus, we assessed by NGS somatic mutations and copy number alterations (CNA) in archived CTCs isolated from patients with metastatic breast cancer who were enrolled in concurrent clinical trials that collected and analyzed CTCs and metastatic tissues. In 76 individual and pooled informative CTCs from 12 patients, we observed 85% concordance in at least one or more prioritized somatic mutations and CNA between paired CTCs and tissue metastases. Potentially actionable genomic alterations were identified in tissue but not CTCs, and vice versa. CTC profiling identified diverse intra- and interpatient molecular mechanisms of endocrine therapy resistance, including loss of heterozygosity in individual CTCs. For example, in one patient, we observed CTCs that were either wild type for ESR1 (n = 5/32), harbored the known activating ESR1 p.Y537S mutation (n = 26/32), or harbored a novel ESR1 p.A569S (n = 1/32). ESR1 p.A569S was modestly activating in vitro, consistent with its presence as a minority circulating subclone. Our results demonstrate the feasibility and potential clinical utility of comprehensive profiling of archived fixed CTCs. Tissue and CTC genomic assessment are complementary, and precise combination therapies will likely be required for effective targeting in advanced breast cancer patients.Significance: These findings demonstrate the complementary nature of genomic profiling from paired tissue metastasis and circulating tumor cells from patients with metastatic breast cancer. Cancer Res; 78(4); 1110-22. ©2017 AACR.

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Heterogeneous mutation pattern in tumor tissue and circulating tumor DNA warrants parallel NGS panel testing.

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Heterogeneous mutation pattern in tumor tissue and circulating tumor DNA warrants parallel NGS panel testing.

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