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"Down the Ocular" - Webcast - Listen to Dr. Bharathi Vennapusa discuss various topics related to NTRK gene fusion testing methodologies and recommendations for stratifying patients with IHC testing prior to NGS confirmation.
1. What kinds of testing methods can be used to detect NTRK gene fusions? (click for answer)
NTRK gene fusions can be detected using various testing methods, including NGS, immunohistochemistry (IHC), which detects protein expression of both fusion and wild-type; fluorescence in situ hybridization (FISH), and reverse transcriptase polymerase chain reaction (RT-PCR).
2. How does each testing method work and what are the advantages and limitations of each? (click for answer)
NGS relies on DNA or ribonucleic acid (RNA) sequencing technologies that are capable of processing multiple DNA or RNA sequences in parallel, and provides the most comprehensive view of several biomarkers. NGS can detect fusions in all 3 NTRK genes (must have NTRK 1, 2, 3 gene-fusion coverage), as well as the NTRK gene fusion partner and position.
However, turnaround time for results with NGS may be longer than with IHC or FISH.
IHC is a protein biomarker test that utilizes specific antibodies to detect expression of the TRK component of the wild type and fusion protein. An antibody directed against a conserved region may detect fusion or wild-type proteins derived from TRKA, TRKB, or TRKC proteins.
Like IHC, FISH break-apart probes can distinguish fusion variants. FISH assays can often be labor intensive and more expensive than IHC when performing multiple assays.
RT-PCR requires multiple primer sets for each gene, since the location of the gene rearrangement is not known. It is very reliable for known fusion variants but requires multiple reactions with specific primers for known variants and can miss detection of unknown/untested variants.
3. Is there a companion diagnostic assay for selecting NTRK fusion-positive eligible patients? (click for answer)
On October 23, 2020, the Food and Drug Administration approved the next-generation sequencing (NGS)-based FoundationOne CDx test (Foundation Medicine, Inc.) as a companion diagnostic to identify fusions in neurotrophic receptor tyrosine kinase (NTRK) genes, NTRK1, NTRK2, and NTRK3, in DNA isolated from tumor tissue specimens from patients with solid tumors eligible for treatment with larotrectinib (VITRAKVI, Bayer Healthcare Pharmaceuticals, Inc.).
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