title: Integrated digital error suppression for improved detection of circulating tumor DNA.
abstract: High-throughput sequencing of circulating tumor DNA (ctDNA) promises to facilitate personalized cancer therapy. However, low quantities of cell-free DNA (cfDNA) in the blood and sequencing artifacts currently limit analytical sensitivity. To overcome these limitations, we introduce an approach for integrated digital error suppression (iDES). Our method combines in silico elimination of highly stereotypical background artifacts with a molecular barcoding strategy for the efficient recovery of cfDNA molecules. Individually, these two methods each improve the sensitivity of cancer personalized profiling by deep sequencing (CAPP-Seq) by about threefold, and synergize when combined to yield ∼15-fold improvements. As a result, iDES-enhanced CAPP-Seq facilitates noninvasive variant detection across hundreds of kilobases. Applied to non-small cell lung cancer (NSCLC) patients, our method enabled biopsy-free profiling of EGFR kinase domain mutations with 92% sensitivity and >99.99% specificity at the variant level, and with 90% sensitivity and 96% specificity at the patient level. In addition, our approach allowed monitoring of NSCLC ctDNA down to 4 in 10(5) cfDNA molecules. We anticipate that iDES will aid the noninvasive genotyping and detection of ctDNA in research and clinical settings.
authors.affiliation: Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, California, USA.|Division of Oncology, Department of Medicine, Stanford Cancer Institute, Stanford University, Stanford, California, USA.
authors.forename: Aaron M
authors.initials: AM
authors.lastname: Newman
authors.affiliation: Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, California, USA.|Department of Radiation Oncology, Stanford University, Stanford, California, USA.|Stanford Cancer Institute, Stanford University, Stanford, California, USA.
authors.forename: Alexander F
authors.initials: AF
authors.lastname: Lovejoy
authors.affiliation: Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, California, USA.|Division of Oncology, Department of Medicine, Stanford Cancer Institute, Stanford University, Stanford, California, USA.|Stanford Cancer Institute, Stanford University, Stanford, California, USA.
authors.forename: Daniel M
authors.initials: DM
authors.lastname: Klass
authors.affiliation: Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, California, USA.|Division of Oncology, Department of Medicine, Stanford Cancer Institute, Stanford University, Stanford, California, USA.|Department of Bioengineering, Stanford University, Stanford, California, USA.
authors.forename: David M
authors.initials: DM
authors.lastname: Kurtz
authors.affiliation: Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, California, USA.
authors.forename: Jacob J
authors.initials: JJ
authors.lastname: Chabon
authors.affiliation: Division of Oncology, Department of Medicine, Stanford Cancer Institute, Stanford University, Stanford, California, USA.
authors.forename: Florian
authors.initials: F
authors.lastname: Scherer
authors.affiliation: Stanford Cancer Institute, Stanford University, Stanford, California, USA.
authors.forename: Henning
authors.initials: H
authors.lastname: Stehr
authors.affiliation: Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, California, USA.|Division of Oncology, Department of Medicine, Stanford Cancer Institute, Stanford University, Stanford, California, USA.
authors.forename: Chih Long
authors.initials: CL
authors.lastname: Liu
authors.affiliation: Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, California, USA.|Department of Radiation Oncology, Stanford University, Stanford, California, USA.
authors.forename: Scott V
authors.initials: SV
authors.lastname: Bratman
authors.affiliation: Department of Radiation Oncology, Stanford University, Stanford, California, USA.
authors.forename: Carmen
authors.initials: C
authors.lastname: Say
authors.affiliation: Stanford Cancer Institute, Stanford University, Stanford, California, USA.
authors.forename: Li
authors.initials: L
authors.lastname: Zhou
authors.affiliation: Department of Radiation Oncology, Stanford University, Stanford, California, USA.
authors.forename: Justin N
authors.initials: JN
authors.lastname: Carter
authors.affiliation: Department of Pathology, Stanford University, Stanford, California, USA.
authors.forename: Robert B
authors.initials: RB
authors.lastname: West
authors.affiliation: Division of Oncology, Department of Medicine, Stanford Cancer Institute, Stanford University, Stanford, California, USA.|Stanford Cancer Institute, Stanford University, Stanford, California, USA.
authors.forename: George W
authors.initials: GW
authors.lastname: Sledge
authors.affiliation: Division of Thoracic Surgery, Department of Cardiothoracic Surgery, Stanford School of Medicine, Stanford University, Stanford, California, USA.
authors.forename: Joseph B
authors.initials: JB
authors.lastname: Shrager
authors.affiliation: Department of Radiation Oncology, Stanford University, Stanford, California, USA.
authors.forename: Billy W
authors.initials: BW
authors.lastname: Loo
authors.affiliation: Division of Oncology, Department of Medicine, Stanford Cancer Institute, Stanford University, Stanford, California, USA.
authors.forename: Joel W
authors.initials: JW
authors.lastname: Neal
authors.affiliation: Division of Oncology, Department of Medicine, Stanford Cancer Institute, Stanford University, Stanford, California, USA.
authors.forename: Heather A
authors.initials: HA
authors.lastname: Wakelee
authors.affiliation: Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, California, USA.|Division of Oncology, Department of Medicine, Stanford Cancer Institute, Stanford University, Stanford, California, USA.|Department of Radiation Oncology, Stanford University, Stanford, California, USA.
authors.forename: Maximilian
authors.initials: M
authors.lastname: Diehn
authors.affiliation: Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, California, USA.|Division of Oncology, Department of Medicine, Stanford Cancer Institute, Stanford University, Stanford, California, USA.|Stanford Cancer Institute, Stanford University, Stanford, California, USA.|Division of Hematology, Department of Medicine, Stanford Cancer Institute, Stanford University, Stanford, California, USA.
authors.forename: Ash A
authors.initials: AA
authors.lastname: Alizadeh
mesh_terms: D000465:Algorithms; D016477:Artifacts; D014408:Biomarkers, Tumor / Q000097:blood* / Q000235:genetics*; D004273:DNA, Neoplasm / Q000097:blood* / Q000235:genetics* / Q000302:isolation & purification; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006801:Humans; D009360:Neoplastic Cells, Circulating / Q000378:metabolism; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D012815:Signal Processing, Computer-Assisted; D018511:Systems Integration
chemical_list: D014408:Biomarkers, Tumor; D004273:DNA, Neoplasm