RNA-based NGS – the best way to identify NTRK gene fusion


How to identify NTRK gene fusions?

IHC testing vs FISH testing vs RT-PCR testing vs NGS testing

NTRK gene fusions can serve as biomarkers for pan-cancer TRK inhibitors. Using biomarker-driven companion diagnostics, clinicians are able to predict the effects of targeted agents against those rare genetic mutations that are potentially tumorigenic across pan-cancer. Immunohistochemistry (IHC), Fluorescence in situ hybridization (FISH), Reverse Transcription Polymerase Chain Reaction (RT-PCR), and Next-generation Sequencing (NGS) testing are commonly used to identify NTRK gene fusions.

The IHC method visualizes the expression of cancer-related protein in cancer tissue. The FISH method uses multi-coloured fluorescent probes to detect break points in a gene. The RT-PCR method amplifies known RNA fusion transcripts for subsequent detection. The NGS technology is a massively parallel method that can sequence both known and novel gene fusions.

Here is a comparison summary table for different methods:
Detection Pros Cons
IHC method TRK protein expression, which is potentially driven by NTRK gene fusions
  • Rapid results
  • Low costs
  • TRK proteins are also expressed in rapid-growing tissues without the presence of NTRK gene fusions
  • Cannot determine fusion partners and breakpoints
FISH method Translocations, amplifications, or deletions on chromosomes
  • Rapid results
  • Low costs
  • Cannot determine fusion partners and breakpoints
  • Requires extra works from experienced and qualified pathologists to carefully interpret the test results
RT-PCR method Fusion transcripts with specific NTRK gene fusion combinations
  • A relatively sensitive test
  • Available in most molecular genetic laboratories
  • Cannot detect novel fusion partners
  • Difficult to amplify across large introns
NGS testing Fusions partners and positions among any NTRK genes
  • Comprehensive and high throughput
  • Potential for multiplexed testing
  • Relatively expensive

Why we should consider RNA-based NGS to identify NTRK gene fusions?

Both RNA-based and DNA-based NGS are capable of multiplexed assessment for different fusion targets. However, RNA-based NGS is preferred for identifying NTRK gene fusions, due to its capability to detect expressed fusion transcripts directly, with more accurate breakpoint readout. RNA-based NGS avoids the difficulties of sequencing larger intronic regions associated with NTRK gene fusions. It also has a faster sequencing process compare to DNA-based NGS. Its detection accuracy can vary depending on the choice of library preparation methods, for example, hybridization-based capture or amplicon-based approach.

Why choose Codex for NGS testing?

Codex provides comprehensive cancer screening using next-generation sequencing (NGS) technology, which helps identify genetic mutations associated with 523 cancer-related genes and 9 types of cancer. The CoGenesis® Bioinformatics platforms allow comprehensive and accurate profiling for data analysis. It provides AI-assisted matching of genomic variants. The platform is fully scalable with shortened computation time. It improves the query performance of complex biological queries.

Codex’s laboratory is using the latest sequencer – Illumina NextSeq 2000, for next-generation sequencing (NGS). It simplifies sequencing workflows and minimizes the need for additional resources. Therefore, sequencing turnaround time is reduced.

Codex Genetics applies a DNA and RNA hybridization-based approach for detecting a broad range of NTRK gene fusions. This approach allows us to detect a larger number of genes and genomic regions, while offering a high level of sensitivity for detecting fusions with novel fusion partners.