Technical considerations for circulating tumor DNA detection in oncology

Franczak, Claire; Filhine-Trésarrieu, Pierre; Gilson, Pauline; Merlin, Jean‐Louis; Au, Lewis; Harlé, Alexandre

doi:10.1080/14737159.2019.1568873

Cited by 31 publications

(24 citation statements)

References 128 publications

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“…Peripheral blood specimens were collected using PAXgene Blood ccfDNA tubes (Qiagen, Germany). Plasma was isolated within six hours in two steps (whole blood was centrifuged at 1.600 g for 20 min at 4 • C, subsequently the plasma fraction was centrifuged at 16.000 g for 10 minutes at 4 • C) following previously published protocols [24,25]. The plasma samples were stored at −70 • C until further processing.…”

Section: Discussionmentioning

confidence: 99%

Quantitative Analysis and Monitoring of EZH2 Mutations Using Liquid Biopsy in Follicular Lymphoma

Nagy

Bátai

Balogh

et al. 2020

Genes

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Recent advances in molecular technologies enable sensitive and quantitative assessment of circulating tumor DNA, offering a noninvasive disease monitoring tool for patients with malignant disorders. Here, we demonstrated on four follicular lymphoma cases that circulating tumor DNA based EZH2 mutation analysis performed by a highly sensitive droplet digital PCR method may be a valuable treatment monitoring approach in EZH2 mutant follicular lymphoma. EZH2 variant allele frequencies changed in parallel with the volume of metabolically active tumor sites observed on 18F-fluorodeoxyglucose positron emission tomography combined with computer tomography (PET-CT) scans. Variant allele frequencies of EZH2 mutations decreased or were eliminated rapidly upon successful treatment, with treatment failure being associated with elevated EZH2 variant allele frequencies. We also demonstrated spatial heterogeneity in a patient with two different EZH2 mutations in distinct anatomical sites, with both mutations simultaneously detected in the liquid biopsy specimen. In summary, circulating tumor DNA based EZH2 mutation analysis offers a rapid, real-time, radiation-free monitoring tool for sensitive detection of EZH2 mutations deriving from different anatomical sites in follicular lymphoma patients receiving immunochemotherapy.

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Section: Discussionmentioning

confidence: 99%

Quantitative Analysis and Monitoring of EZH2 Mutations Using Liquid Biopsy in Follicular Lymphoma

Nagy

Bátai

Balogh

et al. 2020

Genes

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show abstract

“…For blood collection, the ratio of plasma germline cfDNA is lower than that of serum; thus, it is more suitable for ctDNA isolation 24. Plasma separation should be performed within a short time since the half-time of ctDNA is only about 2 hrs in standard EDTA tubes 25.…”

Section: Methodologies In Exploring Cfdna/ctdnamentioning

confidence: 99%

“…There is a variety of PCR-based methods, each with its characteristics, including real-time PCR, co-amplification at lower denaturation temperature-PCR, methylation-specific PCR, digital PCR or droplet digital PCR (ddPCR), “Beads, Emulsion, Amplification, Magnetics digital PCR” (BEAMing) and so on 24. Designed to detect hotspot mutations, these PCR-based methods are relatively time-saving and economical.…”

Section: Methodologies In Exploring Cfdna/ctdnamentioning

confidence: 99%

“…Next-Generation Sequencing (NGS)-based methods such as tagged-amplicon deep sequencing (TamSeq), the Safe Sequencing System (SafeSeqS), CAncer Personalized Profiling by deep Sequencing (CAPP-Seq) and targeted error correction sequencing (TEC-Seq) are available to assess genetic alterations with different detection capacities 24. Whole Genome Sequencing (WGS) and Whole Exome Sequencing (WES) technologies allow for the identification of novel alterations with no pre-existing knowledge 32,33.…”

Section: Methodologies In Exploring Cfdna/ctdnamentioning

confidence: 99%

“…Whole Genome Sequencing (WGS) and Whole Exome Sequencing (WES) technologies allow for the identification of novel alterations with no pre-existing knowledge 32,33. The ability of NGS-based methods to detect copy number variations (CNVs) is an advantage over PCR 24. Though efficient, these methods are relatively time-consuming and require professional bioinformatics analysis.…”

Section: Methodologies In Exploring Cfdna/ctdnamentioning

confidence: 99%

See 2 more Smart Citations

<p>Circulating Cell-Free DNA or Circulating Tumor DNA in the Management of Ovarian and Endometrial Cancer</p>

Chen¹,

Zhang²,

Wang³

et al. 2019

OTT

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Ovarian cancer (OC) is the most lethal cancer of all gynecological malignancies, while endometrial cancer (EC) is the most common one. Current strategies for OC/EC diagnosis consist of the extraction of a solid tissue from the affected area. This sample enables the study of specific biomarkers and the genetic nature of the tumor. However, the tissue extraction is risky and painful for the patient and in some cases is unavailable in inaccessible tumors. Moreover, a tissue biopsy is expensive and requires a highly skilled gynecological surgery to pinpoint accurately which cannot be applied repeatedly. New alternatives that overcome these drawbacks are rising up nowadays, such as liquid biopsy. A liquid biopsy is the analysis of biomarkers in a non-solid biological tissue, mainly blood, which has remarkable advantages over the traditional method. The most studied cancer non-invasive biomarkers are circulating tumor cells (CTCs), circulating tumor DNA (ctDNA), and circulating free DNA (cfDNA). These circulating biomarkers play a key role in the understanding of metastasis and tumorigenesis, which could provide a better insight into the evolution of the tumor dynamics during treatment and disease progression. Liquid biopsy is an emerging non-invasive, safe and effective method with considerable potential for clinical diagnosis and treatment management in patients with OC and EC. Analysis of cfDNA and ctDNA will provide a better characterization of biomarkers and give rise to a wide range of clinical applications, such as early detection of OC/EC, the prediction of treatment responses due to the discovery of personalized tumor-related biomarkers, and therapeutic response monitoring.

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Circulating tumor DNA predicts efficacy of a dual AKT/p70S6K inhibitor (LY2780301) plus paclitaxel in metastatic breast cancer: plasma analysis of the TAKTIC phase IB/II study

et al. 2022

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The phosphatidylinositol-3-kinase (PI3K)/AKT/mammalian target of rapamycin (mTOR) pathway is frequently activated in HER2-negative breast cancer and may play a role in taxane resistance. The phase IB/II TAKTIC trial (NCT01980277) has shown that combining a dual AKT and p70 ribosomal protein S6 kinase (p70S6K) inhibitor (LY2780301) taken orally with weekly paclitaxel in HER2-negative advanced breast cancer is feasible, with preliminary evidence of efficacy. We wanted to explore whether circulating tumor DNA (ctDNA) may be a surrogate marker of treatment efficacy in this setting. Serial plasma samples were collected and cell-free DNA was sequenced using low-coverage whole-genome sequencing, and analysis was completed with droplet digital polymerase chain reaction (PCR) for some patients with driver mutations. Baseline tumor fraction (TF) and TF after 7 weeks on treatment were compared to progression-free survival (PFS) and the overall response rate. We also explored circulating copy number alterations associated with treatment failure. Of the 51 patients enrolled in the TAKTIC trial, at least one plasma sample was available for 44 cases (96 timepoints). All patients with tumor TP53, PI3KCA, or AKT1 mutations harbored at least one of these alterations in plasma. TF at inclusion was correlated with PFS (6m-PFS was 92% for ctDNAneg patients vs 68% for ctDNApos cases; hazard ratio [HR] = 3.45, 95% confidence interval ], P = 0.007). ctDNA status at week 7 was not correlated with prognosis. Even though most circulating copy number alterations were conserved at disease progression, some genomic regions of interest were altered in post-progression samples. In conclusion, ctDNA detection

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Technical considerations for circulating tumor DNA detection in oncology

Cited by 31 publications

References 128 publications

Quantitative Analysis and Monitoring of EZH2 Mutations Using Liquid Biopsy in Follicular Lymphoma

Quantitative Analysis and Monitoring of EZH2 Mutations Using Liquid Biopsy in Follicular Lymphoma

<p>Circulating Cell-Free DNA or Circulating Tumor DNA in the Management of Ovarian and Endometrial Cancer</p>

Circulating tumor DNA predicts efficacy of a dual AKT/p70S6K inhibitor (LY2780301) plus paclitaxel in metastatic breast cancer: plasma analysis of the TAKTIC phase IB/II study

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