Epstein–Barr virus (EBV)-associated diffuse large B-cell lymphoma not otherwise specified (DLBCL NOS) constitute a distinct clinicopathological entity in the current World Health Organization (WHO) classification. However, its genomic features remain sparsely characterized. Here, we combine whole-genome sequencing (WGS), targeted amplicon sequencing (tNGS), and fluorescence in situ hybridization (FISH) from 47 EBV + DLBCL (NOS) cases to delineate the genomic landscape of this rare disease. Integrated WGS and tNGS analysis clearly distinguished this tumor type from EBV-negative DLBCL due to frequent mutations in ARID1A (45%), KMT2A/KMT2D (32/30%), ANKRD11 (32%), or NOTCH2 (32%). WGS uncovered structural aberrations including 6q deletions (5/8 patients), which were subsequently validated by FISH (14/32 cases). Expanding on previous reports, we identified recurrent alterations in CCR6 (15%), DAPK1 (15%), TNFRSF21 (13%), CCR7 (11%), and YY1 (6%). Lastly, functional annotation of the mutational landscape by sequential gene set enrichment and network propagation predicted an effect on the nuclear factor κB (NFκB) pathway (CSNK2A2, CARD10), IL6/JAK/STAT (SOCS1/3, STAT3), and WNT signaling (FRAT1, SFRP5) alongside aberrations in immunological processes, such as interferon response. This first comprehensive description of EBV + DLBCL (NOS) tumors substantiates the evidence of its pathobiological independence and helps stratify the molecular taxonomy of aggressive lymphomas in the effort for future therapeutic strategies.

爱泼斯坦-巴尔病毒（EBV）相关弥漫性大 B 细胞淋巴瘤（DLBCL NOS）在当前世界卫生组织（WHO）分类中构成了一个独特的临床病理学实体。然而，它的基因组特征仍然很少被描述。在这里，我们结合来自 47 例 EBV + DLBCL (NOS) 病例的全基因组测序 (WGS)、靶向扩增子测序 (tNGS) 和荧光原位杂交 (FISH) 来描绘这种罕见疾病的基因组图谱。由于ARID1A (45%)、KMT2A/KMT2D ( 32/30 %)、ANKRD11 (32%) 或NOTCH2 的频繁突变，综合 WGS 和 tNGS 分析清楚地将这种肿瘤类型与 EBV 阴性 DLBCL 区分开来（32%）。WGS 发现了结构异常，包括 6q 缺失（5/8 例患者），随后通过 FISH 验证（14/32 例）。扩展之前的报告，我们发现了CCR6 (15%)、DAPK1 (15%)、TNFRSF21 (13%)、CCR7 (11%) 和YY1 (6%) 的复发性改变。最后，通过序列基因集富集和网络传播对突变景观的功能注释预测了对核因子 κB (NFκB) 通路 ( CSNK2A2 , CARD10 )、IL6/JAK/STAT ( SOCS1/3 , STAT3 ) 和 WNT 信号传导的影响（FRAT1，SFRP5) 以及免疫过程中的异常，例如干扰素反应。这是对 EBV + DLBCL (NOS) 肿瘤的首次全面描述，证实了其病理生物学独立性的证据，并有助于对侵袭性淋巴瘤的分子分类进行分层，以制定未来的治疗策略。