Related Experiment Video
Updated: May 23, 2026
06:41
In Vivo Functional Study of Disease-associated Rare Human Variants Using Drosophila
Published on: August 20, 2019
From Uncertainty to Pathogenicity: Resolving a CSF1R Variant of Uncertain Significance Using Long-Read
Charles Wade1, Kylie Montgomery2,3, Gabriela E Jones4
1Department of Neuroinflammation, Queen Square Institute of Neurology, University College London, London, UK.
Summary
Targeted RNA sequencing resolved a CSF1R-related disorder variant of uncertain significance (VUS) by identifying a novel exon-skipping isoform. This functional evidence enabled reclassification, aiding definitive diagnosis for leukoencephalopathy patients.
Area of Science:
- Genomics and transcriptomics
- Neurogenetics
- Molecular diagnostics
Background:
- CSF1R-related disorder (CSF1R-RD) is a severe, progressive leukoencephalopathy.
- Many CSF1R variants remain classified as variants of uncertain significance (VUS), hindering genetic testing.
- Accurate variant classification is crucial for pre-symptomatic and prenatal diagnosis.
Purpose of the Study:
- To investigate a CSF1R splice-region variant classified as VUS in a patient with CSF1R-RD.
- To determine the transcript-level consequences of the identified CSF1R variant.
- To establish the pathogenicity of the variant and enable definitive molecular diagnosis.
Main Methods:
- Targeted long-read RNA sequencing of peripheral blood was performed.
- Transcriptomic analysis was used to identify novel isoforms and assess transcript abundance.
- Variant pathogenicity was evaluated using ACMG and ACGS guidelines.
Main Results:
- A novel exon 20-skipping isoform of CSF1R was identified, absent in controls.
- This isoform accounted for approximately 63% of transcripts, leading to reduced canonical transcript levels.
- The exon skipping causes a frameshift and premature termination codon, disrupting the tyrosine kinase domain, leading to reclassification from VUS to likely pathogenic.
Conclusions:
- Targeted long-read transcriptomic sequencing provides decisive functional evidence for non-canonical splice variants in CSF1R-RD.
- Transcript-level assessment is a valuable adjunct to genomic testing for variants of uncertain significance.
- This approach facilitates definitive molecular diagnosis in clinically compelling cases.
