Nicolas Pécuchet1,2, Yves Rozenholc3, Eleonora Zonta1, Daniel Pietrasz1, Audrey Didelot1, Pierre Combe1,2, Laure Gibault4, Jean-Baptiste Bachet1,5, Valérie Taly1, Elizabeth Fabre1,2, Hélène Blons1,6, Pierre Laurent-Puig7,6. 1. INSERM UMR-S1147, CNRS SNC 5014, Paris Sorbonne Cité Université, Paris, France-Equipe labélisée Ligue Contre le cancer. 2. Department of Medical Oncology, Hôpital Européen Georges Pompidou (HEGP), Assistance Publique-Hôpitaux de Paris, Paris, France. 3. MERIT-UMR IRD 216, Paris Sorbonne Cité Université, Paris, France. 4. Department of Pathology, Hôpital Européen Georges Pompidou (HEGP), Assistance Publique-Hôpitaux de Paris, Paris, France. 5. Department of Gastro-enterology, Hôpital Pitié-Salpêtrière, Assistance Publique-Hôpitaux de Paris, Paris, France. 6. Department of Biochemistry, Pharmacogenetic and Molecular Oncology Unit, Hôpital Européen Georges Pompidou (HEGP), Assistance Publique-Hôpitaux de Paris, Paris, France. 7. INSERM UMR-S1147, CNRS SNC 5014, Paris Sorbonne Cité Université, Paris, France-Equipe labélisée Ligue Contre le cancer; pierre.laurent-puig@parisdescartes.fr.
Abstract
BACKGROUND: Detecting single-nucleotide variations and insertions/deletions in circulating tumor DNA is challenging because of their low allele frequency. The clinical use of circulating tumor DNA to characterize tumor genetic alterations requires new methods based on next-generation sequencing. METHODS: We developed a method based on quantification of error rate of each base position [position error rate (PER)]. To identify mutations, a binomial test was used to compare the minor-allele frequency to the measured PER at each base position. This process was validated in control samples and in 373 plasma samples from patients with lung or pancreatic cancer. RESULTS: Minimal mutated allele frequencies were 0.003 for single-nucleotide variations and 0.001 for insertions/deletions. Independent testing performed by droplet digital PCR (n = 231 plasma samples) showed strong agreement with the base-PER method (κ = 0.90). CONCLUSIONS: Targeted next-generation sequencing analyzed with the base-PER method represents a robust and low cost method to detect circulating tumor DNA in patients with cancer.
BACKGROUND: Detecting single-nucleotide variations and insertions/deletions in circulating tumor DNA is challenging because of their low allele frequency. The clinical use of circulating tumor DNA to characterize tumor genetic alterations requires new methods based on next-generation sequencing. METHODS: We developed a method based on quantification of error rate of each base position [position error rate (PER)]. To identify mutations, a binomial test was used to compare the minor-allele frequency to the measured PER at each base position. This process was validated in control samples and in 373 plasma samples from patients with lung or pancreatic cancer. RESULTS: Minimal mutated allele frequencies were 0.003 for single-nucleotide variations and 0.001 for insertions/deletions. Independent testing performed by droplet digital PCR (n = 231 plasma samples) showed strong agreement with the base-PER method (κ = 0.90). CONCLUSIONS: Targeted next-generation sequencing analyzed with the base-PER method represents a robust and low cost method to detect circulating tumor DNA in patients with cancer.
Authors: Yoshitaka Zaimoku; Bhavisha A Patel; Sharon D Adams; Ruba Shalhoub; Emma M Groarke; Audrey Ai Chin Lee; Sachiko Kajigaya; Xingmin Feng; Olga Julia Rios; Holly Eager; Lemlem Alemu; Diego Quinones Raffo; Colin O Wu; Willy A Flegel; Neal S Young Journal: Blood Date: 2021-12-30 Impact factor: 25.476
Authors: S Stasik; C Schuster; C Ortlepp; U Platzbecker; M Bornhäuser; J Schetelig; G Ehninger; G Folprecht; C Thiede Journal: Biomol Detect Quantif Date: 2018-01-09