Clinical application of targeted next-generation sequencing for colorectal cancer patients: a multicentric Belgian experience

Abstract

International guidelines made RAS (KRAS and NRAS) status a prerequisite for the use of anti-EGFR agents for metastatic colorectal cancer (CRC) patients. Daily, new data emerges on the theranostic and prognostic role of molecular biomarkers; this is a strong incentive for a validated, sensitive, and broadly available molecular screening test. Next-generation sequencing (NGS) has begun to supplant other technologies for genomic profiling. We report here our 2 years of clinical practice using NGS results to guide therapeutic decisions. The Ion Torrent AmpliSeq colon/lung cancer panel, which allows mutation detection in 22 cancer-related genes, was prospectively used in clinical practice (BELAC ISO 15189 accredited method). The DNA of 741 formalin-fixed paraffin-embedded CRC tissues, including primary tumors and metastasis, was obtained from 14 different Belgian institutions and subjected to targeted NGS. Of the tumors tested, 98% (727) were successfully sequenced and 89% (650) harbored at least one mutation. KRAS, BRAF and NRAS mutations were found in 335 (46%), 78 (11%) and 32 (4%) samples, respectively. These mutation frequencies were consistent with those reported in public databases. Moreover, mutations and amplifications in potentially actionable genes were identified in 464 samples (64%), including mutations in PIK3CA (14%), ERBB2 (0.4%), AKT1 (0.6%), and MAP2K1 (0.1%), as well as amplifications of ERBB2 (0.3%) and EGFR (0.3%). The median turnaround time between reception of the sample in the laboratory and report release was 8 calendar days. Overall, the AmpliSeq colon/lung cancer panel was successfully applied in daily practice and provided reliable clinically relevant information for CRC patients.

Keywords: colorectal cancer; next-generation sequencing.

Figure 1. Pie chart of the distribution of institution contributions

Figure 2. Histogram of the turnaround time measured in calendar days

Figure 3. Molecular profile of CRC samples

Molecular alterations in different genes (rows) are indicated for each CRC sample (columns). A full square indicates that a mutation was found (in the gene), a hatched square indicates that an amplification was found (in the gene), whereas an empty square indicated that no mutation was detected (in the gene).

Figure 4

Histograms of the distribution of base read depth (A, C, E) and of the distribution of amplicon read depth (B, D, F) for a “typical” case (A, B) without gene amplification, for an EGFR-amplified case (C, D) and for an ERBB2-amplified case (E, F).