Feasibility and clinical utility of a pan-solid tumor targeted RNA fusion panel: A single center experience

Performance of an RNA-Based Next-Generation Sequencing Assay for Combined Detection of Clinically Actionable Fusions and Hotspot Mutations in NSCLC

“…Furthermore, however, as RNA, DNA, and ambiguous reads captured for every GSP2 are used to call a variant, we noted that the RNA-based estimated of VAF seemed to be closer to the DNA-based one when the proportion of DNA reads, or DNA/RNA reads ratio, was higher. The performance of AMP-PCR chemistry in solid tumors has been abundantly described for fusion detection 5,12,13 and combined variant/fusion detection using parallel DNA and RNA workflows. 14,15 In contrast, the detection of clinically relevant hotspot mutations using RNA-based NGS is very limited and no study was identified for NSCLC with this strategy.…”

“…The performance of AMP-PCR chemistry in solid tumors has been abundantly described for fusion detection 5 , 12 , 13 and combined variant/fusion detection using parallel DNA and RNA workflows. 14 , 15 In contrast, the detection of clinically relevant hotspot mutations using RNA-based NGS is very limited and no study was identified for NSCLC with this strategy.…”

Next Generation Sequencing for Gene Fusion Analysis in Lung Cancer: A Literature Review

“…Most available kits are coupled with proper analysis systems and a number of total reads and fusion-specific reads to define a reliable test result is dependent on each panel and software. Notably, some NGS gene fusion tests have proved to be able to reveal low allele fractions with an excellent limit of detection and in some studies also samples with a low percentage of tumor cells had adequate results [ 31 , 40 , 41 ]. Another important technical aspect of gene fusion analysis by NGS methods is the possibility to characterize fusion variants.…”

“…The most common anchored mPCR panels are produced by ArcherDX (ArcherDX, Boulder, CO, USA) and have provided excellent results. These panels require a higher RNA input than mPCR assay—about 200 ng [ 39 , 40 ]—and are compatible with both Illumina and ThermoFisher sequencing platforms. Cohen and collaborators evaluated a total of 297 FFPE specimens using the Archer comprehensive Thyroid and Lung Panel with the related analysis software and workflow on an Ion PGM platform.…”

“…Overall fusion variants had an incidence of 19% with a dropout rate of 18%, equal to 30% on cytological specimens, they included also decalcified samples and many of them failed not only RNA but also DNA testing [ 39 ]. Hindi and collaborators, using a custom Archer panel on a NextSeq500 platform (Illumina), analyzed 72 retrospective and 84 prospective FFPE cases (decalcified samples were excluded) and they reported a sensitivity, specificity and reproducibility of 100%; dilution studies confirmed that the limit of detection was equal to 12.5% tumor cells [ 40 ]. Other anchor mPCR NGS panels for gene fusion analysis are produced also by Asuragen (Asuragen, Austin, Texas, USA) and Qiagen (Qiagen, Düsseldorf, Germany) [ 22 ].…”