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Updated: Jan 16, 2026
Oncogenic Gene Fusion Detection Using Anchored Multiplex Polymerase Chain Reaction Followed by Next Generation Sequencing
Published on: July 5, 2019
Fusion Gene Detection in Driver Mutation-Negative Melanomas Using RNA-Based Anchored Multiplex Polymerase Chain
Tokimasa Hida1, Masashi Idogawa2, Sayuri Sato1
1Department of Dermatology, Sapporo Medical University School of Medicine, Sapporo, Japan.
Abstract:
Advanced melanoma is typically treated with immune checkpoint inhibitors (ICIs) and targeted therapies. However, their efficacy is limited in acral and mucosal melanomas, which are more prevalent in non-White populations and often exhibit low tumor mutational burden and lack BRAF mutations. Fusion genes, widely used as therapeutic targets in other cancers, may represent alternative targets in these melanoma subtypes. This study aimed to detect fusion genes in Japanese melanomas lacking major driver mutations (BRAF, RAS, NF1, or KIT) using a custom RNA-based anchored multiplex polymerase chain reaction (AMP) panel. RNA extracted from 14 tumors, primarily formalin-fixed paraffin-embedded specimens, was analyzed using a custom Archer FUSIONPlex panel. Libraries were successfully generated in 80% of cases, and two in-frame fusions-MAD1L1::BRAF and CIC::MEGF8-were identified (17%). MAD1L1::BRAF retained the BRAF kinase domain and may be targetable by MEK inhibitors. CIC::MEGF8, a novel fusion in melanoma, may result in transcriptional dysregulation through CIC loss of function. This Method paper outlines the AMP workflow, including troubleshooting strategies and quality control criteria, and demonstrates its applicability to clinical samples. Our findings support the utility of RNA-based fusion detection in driver-negative melanomas and the potential of fusion genes as actionable targets.
Insights
Fusion genes were identified in Japanese melanomas lacking common mutations, offering new therapeutic targets. This RNA-based approach detects actionable gene fusions in driver-negative melanoma.
Area of Science:
- Oncology
- Genetics
- Molecular Biology
Background:
- Advanced melanoma treatment relies on immune checkpoint inhibitors (ICIs) and targeted therapies.
- Efficacy of current treatments is limited in acral and mucosal melanomas, common in non-White populations.
- These melanoma subtypes often lack major driver mutations like BRAF, necessitating alternative therapeutic strategies.
Purpose of the Study:
- To detect fusion genes in Japanese melanomas that lack common driver mutations (BRAF, RAS, NF1, KIT).
- To evaluate the utility of RNA-based anchored multiplex polymerase chain reaction (AMP) for identifying fusion genes in clinical samples.
Main Methods:
- Analysis of RNA from 14 Japanese melanoma tumors, mostly formalin-fixed paraffin-embedded.
- Utilized a custom Archer FUSIONPlex panel for RNA-based fusion detection.
- Developed and validated an anchored multiplex PCR (AMP) workflow, including quality control.
Main Results:
- Successfully generated libraries from 80% of analyzed samples.
- Identified two in-frame fusion genes: MAD1L1::BRAF and CIC::MEGF8 (17% of cases).
- MAD1L1::BRAF retains the BRAF kinase domain, suggesting potential MEK inhibitor targeting. CIC::MEGF8 is a novel fusion potentially causing transcriptional dysregulation.
Conclusions:
- RNA-based fusion detection is a valuable method for driver-negative melanomas.
- Fusion genes represent promising actionable therapeutic targets for specific melanoma subtypes.
- The described AMP workflow is applicable to clinical samples for fusion gene identification.
