Ion Torrent next-generation sequencing for routine identification of clinically relevant mutations in colorectal cancer patients.
OBJECTIVE
To evaluate the accuracy, consumable cost and time around testing (TAT) of a next-generation sequencing (NGS) assay, the Ion Torrent AmpliSeq Colon and Lung Cancer Panel, as an alternative to Sanger sequencing to genotype KRAS, NRAS and BRAF in colorectal cancer patients.


METHODS
The Ion Torrent panel was first verified on cell lines and on control samples and then prospectively applied to routine specimens (n=114), with Sanger sequencing as reference.


RESULTS
The Ion Torrent panel detected mutant alleles at the 5% level on cell lines and correctly classified all control tissues. The Ion Torrent assay was successfully carried out on most (95.6%) routine diagnostic samples. Of these, 12 (11%) harboured mutations in the BRAF gene and 47 (43%) in either of the two RAS genes, in two cases with a low abundance of RAS mutant allele which was missed by Sanger sequencing. The mean TAT, from sample receipt to reporting, was 10.4 (Sanger) and 13.0 (Ion Torrent) working days. The consumable cost for genotyping KRAS, NRAS and BRAF was €196 (Sanger) and €187 (Ion Torrent).


CONCLUSIONS
Ion Torrent AmpliSeq Colon and Lung Cancer Panel sequencing is as robust as Sanger sequencing in routine diagnostics to select patients for anti-epidermal growth factor receptor (EGFR) therapy for metastatic colorectal cancer.
Department of Public Health, Pathological section, University of Naples Federico II, Naples, Italy.
Umberto
U
Malapelle
Department of Public Health, Pathological section, University of Naples Federico II, Naples, Italy.
Elena
E
Vigliar
Department of Public Health, Pathological section, University of Naples Federico II, Naples, Italy.
Roberta
R
Sgariglia
Department of Public Health, Pathological section, University of Naples Federico II, Naples, Italy.
Claudio
http://orcid.org/0000-0002-7479-6457
C
Bellevicine
Mediterranean Institute of Oncology (IOM), Catania, Italy PhD Programme in Biotechnology and Clinical Medicine, Sapienza University, Rome, Italy.
Lorenzo
L
Colarossi
Mediterranean Institute of Oncology (IOM), Catania, Italy.
Domenico
D
Vitale
CNR/IEOS, Institute of Experimental Endocrinology and Oncology, National Research Council, Naples, Italy Department of Molecular Medicine and Medical Biotechnology (DMMBM), University of Naples Federico II, Naples, Italy.
Pierlorenzo
P
Pallante
Department of Public Health, Pathological section, University of Naples Federico II, Naples, Italy.
Giancarlo
G
Troncone
D014408:Biomarkers, Tumor / Q000235:genetics; D015179:Colorectal Neoplasms / Q000235:genetics*; D004252:DNA Mutational Analysis / Q000191:economics / Q000379:methods*; D005838:Genotype; D059014:High-Throughput Nucleotide Sequencing / Q000191:economics / Q000379:methods*; D006801:Humans
D014408:Biomarkers, Tumor
BRAF; Ion Torrent; NGS; cetuximab; colon cancer; KRAS; NRAS; panitumumab