Noninvasive Identification and Monitoring of Cancer Mutations by Targeted Deep Sequencing of Plasma DNA

  title={Noninvasive Identification and Monitoring of Cancer Mutations by Targeted Deep Sequencing of Plasma DNA},
  author={Tim Forshew and Muhammed Murtaza and Christine A. Parkinson and Davina Gale and Dana Wai Yi Tsui and Fiona Kaper and Sarah-Jane Dawson and Anna M Piskorz and Mercedes Jimenez-Linan and D. R. Bentley and James Hadfield and Andrew P. May and Carlos Caldas and James D. Brenton and Nitzan Rosenfeld},
  journal={Science Translational Medicine},
  pages={136ra68 - 136ra68}
Sizable genomic regions were screened and low-frequency mutations were identified in circulating DNA of cancer patients using tagged-amplicon deep sequencing (TAm-Seq). Deep Sequencing Tumor DNA in Plasma Five liters of circulating blood contain millions of copies of the genome, broken into short fragments; in cancer patients, a small fraction is circulating tumor DNA (ctDNA). An even smaller number harbor mutations that affect cancer outcome. Looking for diagnostic answers in circulating DNA… 
Utility of targeted deep sequencing for detecting circulating tumor DNA in pancreatic cancer patients
It is demonstrated that ctDNA levels measured by targeted deep sequencing sensitively indicate the presence of cancer and correlate well with clinical responses to therapy and disease progression in PDAC patients.
Characterization of DNA lesions associated with cell-free DNA by targeted deep sequencing
The data suggested that cytosine deamination increased in cfDNA compared to that in cellular gDNA, which can contribute to a better understanding of cfDNA-associated DNA damage, which will enable the accurate analysis of somatic variants present in cf DNA at an extremely low frequency.
Ultradeep targeted sequencing of circulating tumor DNA in plasma of early and advanced breast cancer
This pilot study endorses the use of targeted NGS for non‐invasive molecular profiling of breast cancer and recommendsaired sequencing of plasma ctDNA and WBC should be implemented to improve accurate interpretation of liquid biopsy.
Selecting short DNA fragments in plasma improves detection of circulating tumour DNA
Size selection allows detection of tumour alterations masked by non-tumour DNA in plasma and could help overcome sensitivity limitations of liquid biopsy for applications in early diagnosis, detection of minimal residual disease, and genomic profiling.
ctDNA monitoring using patient-specific sequencing and integration of variant reads
It is shown that sensitivity for ctDNA detection in plasma can be improved by analyzing hundreds to thousands of mutations that are first identified by tumor genotyping, and the INtegration of VAriant Reads (INVAR) pipeline, which combines custom error-suppression methods and signal-enrichment approaches based on biological features of ct DNA is described.
Whole genome deep sequencing analysis of cell-free DNA in samples with low tumour content
Deep sequencing analysis of plasma samples revealed higher fraction of unique somatic mutations in plasma samples, which were not detected in matched tumour samples, and Sequencing of di-nucleosome bound cfDNA fragments may increase recovery of tumour mutations from plasma.
Characterization of Factors Affecting the Detection Limit of EGFR p.T790M in Circulating Tumor DNA
Both the sequencing platform and the specific nucleotide change affect the limit of detection and should therefore be determined in the validation process of new assays, supporting the approach to determine the detection limit.
High-sensitivity monitoring of ctDNA by patient-specific sequencing panels and integration of variant reads
The INtegration of VAriant Reads (INVAR) analysis pipeline, which combines patient-specific mutation lists with both custom error-suppression methods and signal enrichment based on biological features of ctDNA, is described and demonstrated that INVAR can be generalized and allows improved detection of cTDNA from whole-exome and low-depth whole-genome sequencing data.
Efficacy comparison of targeted next-generation sequencing in the identification of somatic mutations in circulating tumor DNA from different stages of lung cancer
It appears that ctDNA NGS-based deep sequencing is a feasible approach to identify mutations in patients with Stage IV NSCLC, however, additional methods with higher sensitivity and specificity are needed to improve the successful application of this platform in the earlier stages ofNSCLC.


Detection of tumor mutations in the presence of excess amounts of normal DNA
A screening method, termed PPEM (PNA-directed PCR, primer extension, MALDI-TOF), that can detect as few as 3 copies of mutant alleles in the presence of a 10,000-fold excess of normal allele in a robust and specific manner and is amenable to high-throughput automation.
Detection of low prevalence somatic mutations in solid tumors with ultra-deep targeted sequencing
With an improved performance when run on the Illumina Miseq, the UDT-Seq assay is well suited for clinical applications to guide therapy and study clonal selection in heterogeneous samples.
High-throughput detection of actionable genomic alterations in clinical tumor samples by targeted, massively parallel sequencing.
A sequencingbased approach to identifying genomic alterations in FFPE tumor samples is described, affirm the feasibility and clinical utility of targeted sequencing in the oncology arena and provide a foundation for genomics-based stratification of cancer patients.
High Throughput Interrogation of Somatic Mutations in High Grade Serous Cancer of the Ovary
Sequenom analysis using OncoMap on DNA extracted from FFPE ovarian cancer samples is feasible and leads to the detection of potentially druggable mutations.
Circulating mutant DNA to assess tumor dynamics
It is found that ctDNA measurements could be used to reliably monitor tumor dynamics in subjects with cancer who were undergoing surgery or chemotherapy, and it is suggested that this personalized genetic approach could be generally applied to individuals with other types of cancer.
Detection of mutations in EGFR in circulating lung-cancer cells.
Molecular analysis of circulating tumor cells from the blood of patients with lung cancer offers the possibility of monitoring changes in epithelial tumor genotypes during the course of treatment, and shows that a reduction in the number of captured cells was associated with a radiographic tumor response; an increase in theNumber of cells wasassociated with tumor progression, with the emergence of additional EGFR mutations in some cases.
Detection and quantification of mutations in the plasma of patients with colorectal tumors.
  • F. Diehl, M. Li, B. Vogelstein
  • Biology, Medicine
    Proceedings of the National Academy of Sciences of the United States of America
  • 2005
Patients with advanced colorectal cancers consistently contained mutant adenomatous polyposis coli (APC) DNA molecules in their plasma, which has implications for the mechanisms through which tumor DNA is released into the circulation and for diagnostic tests based on this phenomenon.
Single-Molecule Detection of Epidermal Growth Factor Receptor Mutations in Plasma by Microfluidics Digital PCR in Non–Small Cell Lung Cancer Patients
The sensitive detection and accurate quantification of low abundance EGFR mutations in tumor tissues and plasma by microfluidics digital PCR would be useful for predicting treatment response, monitoring disease progression and early detection of treatment failure associated with acquired drug resistance.
Replacing PCR with COLD-PCR enriches variant DNA sequences and redefines the sensitivity of genetic testing
Co-amplification at lower denaturation temperature PCR (COLD-PCR), a novel form of PCR that amplifies minority alleles selectively from mixtures of wild-type and mutation-containing sequences irrespective of the mutation type or position on the sequence, is described.