Abstract
BACKGROUND: Epstein-Barr virus (EBV)(+) and EBV(-) primary CNS lymphomas (PCNSL) carry distinct mutational landscapes, but their transcriptional and epigenetic profiles have not been integrated and compared. This precludes further insights into pathobiology and molecular differences, relevant for classification and targeted therapy. MATERIAL AND METHODS: 23 EBV- and 15 EBV+ PCNSL, histologically classified as diffuse large B-cell lymphomas, were subjected to RNA-Sequencing and EPIC methylation arrays. Unsupervised clustering analyses were performed. Differentially expressed and differentially methylated genes were identified and integrated. Patient-derived cell lines were used to validate findings and identify novel targeted therapies. RESULTS: Two distinct transcriptional clusters were found, which separated EBV- and EBV+ PCNSL (p < 0.0001). The EBV+transcriptional signature contained genes (GPR15, FCER2/CD23, SLAMF1/CD150) closely regulated by EBV oncogenes in B-cells. Pathway enrichment analysis uncovered enhanced B-cell receptor (BCR) and WNT/beta-catenin signaling in EBV- lymphomas, whereas Interleukin-10, NOTCH, and viral life cycle pathways were upregulated in EBV+ PCNSL. Correspondingly, BCR-associated SYK kinase activity was enriched in EBV- tumors while JAK2 was overrepresented in EBV+ PCNSL. Epigenetic profiling revealed reduced global promoter methylation in EBV+ PCNSL. Two methylation clusters were recognized, which separated EBV- and EBV+ PCNSL (p < 0.0001). Epigenetic profiles were distinct from 2,788 other brain tumor and non-malignant reference samples. Promoter region hypermethylation of CD79B, a BCR subunit critical for sustained proliferation in EBV- disease, highly correlated (R = -0.7) with its transcriptional downregulation in EBV+ PCNSL. EBV- patient-derived PCNSL cell lines were particularly sensitive to BTK inhibition whereas EBV+ cell lines could be specifically targeted using JAK/STAT inhibitors. CONCLUSIONS: EBV+ and EBV- PCNSL harbor distinct transcriptional and epigenetic profiles, corroborating them as distinctive biological subtypes. Uncovered differences provide novel insights into their pathobiology, may guide molecular diagnostics and targeted therapies.