Open AccessApplication of circulating tumor DNA in prospective clinical oncology trials – standardization of preanalytical conditions
Author(s) -
Dessel Lisanne F.,
Beije Nick,
Helmijr Jean C.A.,
Vitale Silvia R.,
Kraan Jaco,
Look Maxime P.,
Wit Ronald,
Sleijfer Stefan,
Jansen Maurice P.H.M.,
Martens John W.M.,
Lolkema Martijn P.
Publication year - 2017
Publication title -
molecular oncology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.332
H-Index - 88
eISSN - 1878-0261
pISSN - 1574-7891
DOI - 10.1002/1878-0261.12037
Subject(s) - dna , digital polymerase chain reaction , polymerase chain reaction , dna extraction , somatic cell , circulating tumor dna , microbiology and biotechnology , whole blood , liquid biopsy , biology , medicine , cancer , immunology , genetics , gene
Circulating tumor DNA (ct DNA ) has emerged as a potential new biomarker with diagnostic, predictive, and prognostic applications for various solid tumor types. Before beginning large prospective clinical trials to prove the added value of utilizing ct DNA in clinical practice, it is essential to investigate the effects of various preanalytical conditions on the quality of cell‐free DNA (cf DNA ) in general and of ct DNA in particular in order to optimize and standardize these conditions. Whole blood samples were collected from patients with metastatic cancer bearing a known somatic variant. The following preanalytical conditions were investigated: (a) different time intervals to plasma isolation (1, 24, and 96 h) and (b) different preservatives in blood collection tubes ( EDTA , CellSave, and BCT ). The quality of cf DNA /ct DNA was assessed by DNA quantification, digital polymerase chain reaction ( dPCR ) for somatic variant detection and a β‐actin fragmentation assay for DNA contamination from lysed leukocytes. In 11 (69%) of our 16 patients, we were able to detect the known somatic variant in ct DNA . We observed a time‐dependent increase in cf DNA concentrations in EDTA tubes, which was positively correlated with an increase in wild‐type copy numbers and large DNA fragments (> 420 bp). Using different preservatives did not affect somatic variant detection ability, but did stabilize cf DNA concentrations over time. Variant allele frequency was affected by fluctuations in cf DNA concentration only in EDTA tubes at 96 h. Both CellSave and BCT tubes ensured optimal ct DNA quality in plasma processed within 96 h after blood collection for downstream somatic variant detection by dPCR .