Cytologic and Molecular Diagnostics for Vitreoretinal Lymphoma: Current Approaches and Emerging Single-Cell Analyses

Abstract

Vitreoretinal lymphoma (VRL) is a rare ocular malignancy that manifests as diffuse large B-cell lymphoma. Early and accurate diagnosis is essential to prevent mistreatment and to reduce the high morbidity and mortality associated with VRL. The disease can be diagnosed using various methods, including cytology, immunohistochemistry, cytokine analysis, flow cytometry, and molecular analysis of bulk vitreous aspirates. Despite these options, VRL diagnosis remains challenging, as samples are often confounded by low cellularity, the presence of debris and non-target immunoreactive cells, and poor cytological preservation. As such, VRL diagnostic accuracy is limited by both false-positive and false-negative outcomes. Missed or inappropriate diagnosis may cause delays in treatment, which can have life-threatening consequences for patients with VRL. In this review, we summarize current knowledge and the diagnostic modalities used for VRL diagnosis. We also highlight several emerging molecular techniques, including high-resolution single cell-based analyses, which may enable more comprehensive and precise VRL diagnoses.

Keywords: cytology; diffuse large B-cell lymphoma; molecular diagnostics; precision medicine; single-cell analysis; vitreoretinal lymphoma.

Figure 1

Comparison of bulk cell-vs. single cell-based MYD88 analysis. (A) A heterogeneous cellular sample with MYD88^WT (red) or MYD88^L265P mutants (blue) was lysed for bulk-cell myeloid differentiation primary response 88 (MYD88) analysis. (B) An electropherogram showing sequencing peaks for MYD88^WT and MYD88^L265P. The black arrows throughout indicate the location of a T→C point mutation corresponding to MYD88^WT (CTG) or MYD88^L265P (CCG), respectively. (C) Individual cells with MYD88^WT (red/red), heterozygous (red/blue), or homozygous MYD88^L265P mutant (blue/blue) were isolated and lysed for single cell-based MYD88 analysis. (D) An electropherogram showing clean and well-resolved sequencing peaks for MYD88^WT, heterozygous (red/blue), or homozygous MYD88^L265P alleles from the single cells. The black arrows throughout indicate the location of a T→C point mutation corresponding to MYD88^WT (CTG) or MYD88^L265P (CCG), respectively. WT, wild-type MYD88^WT; mutant, MYD88^L265P.