Single B-Cell Genomic Analyses Differentiate Vitreoretinal Lymphoma from Chronic Inflammation.

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) 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 MYD88 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 MYD88 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 MYD88, 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, MYD88 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.