Back to Search
Start Over
Reality of Single Circulating Tumor Cell Sequencing for Molecular Diagnostics in Pancreatic Cancer.
- Source :
-
The Journal of molecular diagnostics : JMD [J Mol Diagn] 2016 Sep; Vol. 18 (5), pp. 688-696. Date of Electronic Publication: 2016 Jul 01. - Publication Year :
- 2016
-
Abstract
- To understand the potential and limitations of circulating tumor cell (CTC) sequencing for molecular diagnostics, we investigated the feasibility of identifying the ubiquitous KRAS mutation in single CTCs from pancreatic cancer (PC) patients. We used the NanoVelcro/laser capture microdissection CTC platform, combined with whole genome amplification and KRAS Sanger sequencing. We assessed both KRAS codon-12 coverage and the degree that allele dropout during whole genome amplification affected the detection of KRAS mutations from single CTCs. We isolated 385 single cells, 163 from PC cell lines and 222 from the blood of 12 PC patients, and obtained KRAS sequence coverage in 218 of 385 single cells (56.6%). For PC cell lines with known KRAS mutations, single mutations were detected in 67% of homozygous cells but only 37.4% of heterozygous single cells, demonstrating that both coverage and allele dropout are important causes of mutation detection failure from single cells. We could detect KRAS mutations in CTCs from 11 of 12 patients (92%) and 33 of 119 single CTCs sequenced, resulting in a KRAS mutation detection rate of 27.7%. Importantly, KRAS mutations were never found in the 103 white blood cells sequenced. Sequencing of groups of cells containing between 1 and 100 cells determined that at least 10 CTCs are likely required to reliably assess KRAS mutation status from CTCs.<br /> (Copyright © 2016 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.)
- Subjects :
- Biomarkers, Tumor
Cell Line, Tumor
DNA Mutational Analysis
Genotype
Humans
Molecular Diagnostic Techniques
Neoplastic Cells, Circulating pathology
Polymerase Chain Reaction
Proto-Oncogene Proteins p21(ras) genetics
Reproducibility of Results
Sensitivity and Specificity
Sequence Analysis, DNA
Microchip Analytical Procedures methods
Neoplastic Cells, Circulating metabolism
Pancreatic Neoplasms diagnosis
Pancreatic Neoplasms genetics
Single-Cell Analysis methods
Subjects
Details
- Language :
- English
- ISSN :
- 1943-7811
- Volume :
- 18
- Issue :
- 5
- Database :
- MEDLINE
- Journal :
- The Journal of molecular diagnostics : JMD
- Publication Type :
- Academic Journal
- Accession number :
- 27375074
- Full Text :
- https://doi.org/10.1016/j.jmoldx.2016.03.006