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Updated: Nov 29, 2025
Author Spotlight: Advancing VRL Diagnosis Using Cell-Free DNA Extraction from Vitreous Humor
Published on: January 12, 2024
Single B-Cell Genomic Analyses Differentiate Vitreoretinal Lymphoma from Chronic Inflammation
Wei Jian Tan1, Mona Meng Wang2, Paola Ricciardi Castagnoli1
1A. Menarini Biomarkers Singapore Pte. Ltd., Singapore.
Insights
Analyzing single B cells from vitreous fluid using immunoglobulin heavy chain (IGH) and MYD88 mutations can distinguish vitreoretinal lymphoma (VRL) from chronic inflammation. This genomic approach offers a promising diagnostic tool for VRL.
Area of Science:
- Ophthalmology
- Hematology
- Genomics
Background:
- Distinguishing vitreoretinal lymphoma (VRL) from chronic inflammation is critical for appropriate patient management.
- Current diagnostic methods may not always provide definitive differentiation.
Purpose of the Study:
- To evaluate the utility of single B-cell genomic analysis from vitreous fluid for differentiating VRL from chronic inflammation.
- To identify key genomic features, including immunoglobulin heavy chain (IGH) and MYD88L265P mutations, and copy number profiles, for diagnostic purposes.
Main Methods:
- Retrospective analysis of vitreous biopsies from patients with VRL and chronic inflammation.
- Single B-cell isolation using the DEPArray NxT system.
- Genomic analysis including IGH gene rearrangement, MYD88L265P mutation detection, and genome-wide copy number profiling.
Main Results:
- Significantly higher frequencies of dominant IGH and MYD88L265P mutations were observed in VRL patients compared to those with chronic inflammation.
- Single-cell analysis revealed clonal B-cell populations and characteristic copy number aberrations in VRL cases, including a BCL2/JH translocation.
- Copy number profiles showed high similarity within patients with VRL but no common genome-wide signatures across different VRL patients.
Conclusions:
- Single B-cell genomic characterization of IGH, MYD88L265P mutation, and copy number profiles is a viable method for VRL diagnosis.
- These proof-of-concept findings in a small cohort warrant further investigation in larger patient populations.
Purpose:
To test whether analyzing DEPArray (Menarini Silicon Biosystems) isolated single B cells from the vitreous fluid can reveal crucial genomic and clinicopathological features to distinguish patients with vitreoretinal lymphoma (VRL) from those with chronic inflammation using immunoglobulin heavy chain (IGH), disease biomarker myeloid differentiation primary response 88 (MYD88)L265P mutation, and copy number profiling.
Design:
A single-center, retrospective study.
Participants:
Remnant vitreous biopsies from 7 patients with VRL and 4 patients with chronic inflammation were acquired for molecular analysis.
Methods:
Vitreous fluid samples were prefixed in PreservCyt (Hologic) and underwent cytologic analysis and immunohistochemistry examination. Single cells were isolated using the DEPArray NxT system, followed by downstream genomic analysis.
Main Outcome Measures:
The frequencies of the dominant IGH and MYD88L265P mutation and the genome-wide copy number aberration (CNA) profiles of individual vitreous-isolated B cells were characterized.
Results:
An average of 10 to 13 vitreous B cells were used in the single-cell IGH and MYD88 analyses. Higher frequencies of dominant IGH (88.8% ± 13.2%) and MYD88L265P mutations (35.0% ± 31.3%) were detected in patients with VRL than in patients with chronic inflammation (65.9% ± 13.4% and 1.5% ± 2.6% for IGH and MYD88L265P, respectively). In a cytology-proven VRL case, all 15 vitreous isolated B cells were derived from the same clone with 100% paired IGH: immunoglobulin light chain (IGK) sequences. Genome-wide copy number profiling revealed a high degree of similarity between B cells from the same patient with VRL, with extensive gains and losses at the same areas across the whole genome. In addition, 14 of 15 B cells showed a BCL2/JH t(14;18) translocation, confirming cellular malignancy with a clonal origin. Clustering analysis of the copy number profiles revealed that malignant B cells derived from different patients with VRL had no common genome-wide signatures.
Conclusions:
Single B-cell genomic characterization of the IGH, MYD88L265P mutation, and copy number profile enables VRL diagnosis. Because our study involved only a small cohort, these meaningful proof-of-concept data now warrant further investigation in a larger patient cohort.
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