Rapid detection of mutations in decalcified lung cancer bone metastasis.

Detection of molecular alterations in lung cancer bone metastasis (LCBM) is particularly difficult when decalcification procedure is needed. The Idylla™ real-time (RT)-PCR is compared to the routine method used in our laboratory, which combines next generation and Sanger sequencing, for the detection of mutations in LCBM. LCBM subjected to EDTA or formic acid decalcification were analysed for mutational status using two methods: first, the Ion Torrent Ampliseq next generation sequencing (NGS) assay +/- Sanger sequencing was used prospectively; then, the fully-automated, RT-PCR based molecular testing system Idylla™ Mutation Test was applied retrospectively. Out of the 34 LCBM assayed, 14 (41.2%) were unsuitable for NGS analysis and five remained unsuitable after additional Sanger sequencing (5/34, 14.7%). Using Idylla™, valid results were observed for 33/34 samples (97.1%). The concordance between the NGS +/- Sanger sequencing method and the RT-PCR method was 89.7% (26/29), one false positive S768I mutation and two false negative results were observed using Idylla™; one of these false negative cases was diagnosed by Sanger sequencing with a rare exon 19 mutation not covered by the Idylla™ Mutation Test design. Detection of mutations in decalcified LCBM is challenging using NGS, more than half of samples showing invalid results. Alternative methods should thus be preferred to spare clinical samples and decrease delay. The Idylla™ Mutation Test shows a good performance on decalcified bone samples and could be used as a first step. In case of negative results, a sequencing approach is mandatory to check the presence of rare mutations sensitive to EGFR tyrosine kinase inhibitors.