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Updated: May 23, 2025
Targeted RNA Sequencing Assay to Characterize Gene Expression and Genomic Alterations
Published on: August 4, 2016
Identification of diagnostic and prognostic genetic alterations in uveal melanoma using RNA sequencing
Rogier J Nell1, Mieke Versluis2, Davy Cats3
1Department of Ophthalmology, Leiden University Medical Center, PO Box 9600, 2300 RC, Leiden, The Netherlands. r.j.nell@lumc.nl.
Abstract:
Uveal melanoma is a lethal intraocular tumour, in which the presence of various genetic alterations correlates with the risk of metastatic dissemination and survival. Here, we tested the detectability of all key mutations and chromosomal changes from RNA sequencing data in 80 primary uveal melanomas studied by The Cancer Genome Atlas (TCGA) initiative, and in five prospective cases. Whereas unsupervised gene expression profiling strongly indicated the presence of chromosome 3 alterations, it was not reliable in identifying other alterations. Though, the presence of both chromosome 3 and 8q copy number alterations could be successfully inferred from expressed allelic imbalances of heterozygous common single nucleotide polymorphisms. Most mutations were adequately recognised in the RNA by their nucleotide changes (all genes), alternative splicing around the mutation (BAP1) and transcriptome-wide aberrant splicing (SF3B1). Notably, in the TCGA cohort we detected previously unreported mutations in BAP1 (n = 3) and EIF1AX (n = 5), that were missed by the original DNA sequencing. In our prospective cohort, all genetic alterations were successfully identified by combining the described approaches. In conclusion, a transcriptional analysis presents insights into the expressed tumour genotype and its phenotypic consequences and may augment or even substitute DNA-based approaches, with potential applicability in research and clinical practice.
Insights
RNA sequencing can detect genetic mutations and chromosomal changes in uveal melanoma, offering a powerful tool for understanding tumor genotype and guiding clinical practice.
Area of Science:
- Oncology
- Genetics
- Ophthalmology
Background:
- Uveal melanoma is a deadly eye cancer with genetic alterations linked to metastasis and survival.
- Accurate detection of these genetic changes is crucial for prognosis and treatment.
Purpose of the Study:
- To assess the detectability of key mutations and chromosomal alterations in uveal melanoma using RNA sequencing data.
- To explore RNA sequencing as a potential alternative or supplement to DNA-based methods for uveal melanoma genetic profiling.
Main Methods:
- Analysis of RNA sequencing data from 80 The Cancer Genome Atlas (TCGA) uveal melanoma cases and 5 prospective cases.
- Utilized gene expression profiling, analysis of expressed allelic imbalances, and detection of nucleotide changes and aberrant splicing patterns.
Main Results:
- RNA sequencing reliably inferred chromosome 3 alterations and copy number alterations (chromosomes 3 and 8q) via expressed allelic imbalances.
- Most mutations were detected through nucleotide changes, BAP1 splicing alterations, and SF3B1 transcriptome-wide aberrant splicing.
- Identified novel BAP1 and EIF1AX mutations in the TCGA cohort missed by prior DNA sequencing.
Conclusions:
- Transcriptional analysis provides insights into the expressed tumor genotype and its phenotypic impact in uveal melanoma.
- RNA sequencing can effectively identify genetic alterations, potentially augmenting or replacing DNA-based approaches in research and clinical settings.
