Abstract
Background: Parallel sequencing technologies have become integrated into clinical practice. This study evaluated the TruSight Tumor 170 assay for the simultaneous detection of somatic gene mutations (SNPs and indels), gene fusions and CNVs, and its implementation into routine diagnostics. Methods: Forty-four formalin-fixed, paraffin-embedded tissue samples analyzed previously with validated methods were evaluated with the TruSight Tumor 170 assay (Illumina). For data analysis the TruSight Tumor 170 app, the BaseSpace Variant Interpreter (Illumina), and the Molecular Health Guide Software (Molecular Health) were used. Results: All somatic gene mutations were identified when covered by the assay. Two high-level MET amplifications were detected by CNV analysis. Focal MET amplifications with a copy number below 10 were not reliably detected at the DNA-level. Twenty-one of 31 fusions and splice variants were confirmed with the assay on the RNA-level. The remaining eight aberrations were incorrect by previous methods. In two cases, no splicing was observed. Conclusions: The TruSight Tumor 170 gives reliable results even if low DNA and RNA concentrations are applied in comparison to other methods and can be used in a routine workflow to detect somatic gene mutations, gene fusions, and splice variants. However, we were not able to detect most focal gene amplifications/deletions.
Highlights
In the last decade, high-quality molecular analysis of formalin-fixed, paraffin-embedded (FFPE) tissue has become crucial for personalized treatment strategies in routine clinical practice [1]
We evaluated the TruSight Tumor 170 assay on FFPE tumor samples with a variety of known genetic aberrations for the simultaneous detection of somatic gene mutations (SNPs and indels), gene fusions, and copy number variations (CNVs) and its implementation into routine diagnostics
The 42 formalin-fixed and paraffin embedded (FFPE) tumor samples with known genetic aberrations including somatic gene mutations (SNPs and indels), CNVs, gene fusions, and splice variants and two FFPE control samples (Quantitative Multiplex Reference Standard (FFPE) from Horizon Discovery, Cambridge, United Kingdom were included in the cohort
Summary
High-quality molecular analysis of formalin-fixed, paraffin-embedded (FFPE) tissue has become crucial for personalized treatment strategies in routine clinical practice [1]. Targeted parallel sequencing provides a high through-put, fast, and cost effective technology, and offers a more comprehensive and accurate approach for genome wide analysis and the detection of somatic mutations [2,3]. Many institutions use amplicon-based parallel sequencing approaches for the detection of somatic gene mutations. With this method, target regions are enriched by multiplex PCR. Amplicon-based approaches are time saving and cost effective for the detection of single nucleotide variants (SNVs), insertions, deletions (indels), or duplications on the DNA-level [6]. Amplicon-based panels cannot detect all relevant genomic alterations like SNVs, indels, CNV, or gene fusions in one assay. This study evaluated the TruSight Tumor 170 assay for the simultaneous detection of somatic gene mutations (SNPs and indels), gene fusions and CNVs, and its implementation into routine diagnostics
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