Characterization of double-negative B cells within the NPC microenvironment

Abstract

<p>Introduction: IgD-CD27- double-negative (DN) B cells are involved in various autoimmune and infectious diseases. A DN B cell population was identified within the tumor microenvironment (TME) of nasopharyngeal carcinoma (NPC), a malignancy highly associated with Epstein-Barr Virus (EBV) infection. The DN B cell population was associated with poor clinical outcomes. However, the mechanisms associating DN B cells with poor NPC prognosis are unknown. This study aims to characterize NPC-infiltrating DN B cells and elucidate their interactions with the NPC TME.</p><p>Methods: We analyzed the single-cell RNA-seq (scRNA-seq) data from four NPC cohorts. Fifty-nine samples from 43 NPC patients and 6 healthy subjects, consisting of 10 NPC-patient derived peripheral blood samples, 43 NPC tissue samples, and 6 normal tissue samples; and a total of 463,913 cells were included in this study.</p><p>Results: NPC-infiltrating B cells were divided into 14 unique subclusters, with NPC tumor samples being enriched in DN B cells compared to normal tissue samples (P = 0.07) and NPC-patient-derived blood samples (P < 0.01). The majority of NPC-infiltrating DN B cells do not express marker genes associated with previously described DN B subtypes, such as CR2 and CXCR5 for DN1, ITGAX and TBX21 for DN2, TBX21 for DN3 and IGHE for DN4 B cells. However, we observed a small proportion of NPC-infiltrating DN B cells expressing DN1-associated marker genes. Some of the genes differentially expressed by the DN B cells, such as HNRNPH1 and DDX17, are involved in mRNA processing. Network functional enrichment analysis identified biological pathways involved in mRNA processing, viral genome replication regulation, and viral responses to be enriched in DN B cells. This suggests the DN B cells may have altered mRNA processing, while the presence of EBV in the NPC TME might affect those DN B cells. We next conducted trajectory analysis to investigate the cellular development in relation to mRNA processing from the NPC-infiltrating DN B cells. They were identified as an intermediary B cell development state, suggesting the DN B cell population may represent a B cell developmental block within the NPC TME. Finally, we compared the gene expression profiles of NPC cancer cells from NPC-high and NPC-low tumors and revealed an association between DN B cell content and NPC tumors with high expression of immune-related genes, such as Human Leukocyte Antigen (HLA) genes and interferon-induced genes.</p><p>Conclusion: We characterized the NPC-infiltrating DN B cells and revealed their potential alterations in mRNA-splicing and differentiation. Further research is needed to study the interactions between DN B cells with NPC tumors with high immune-related gene expression, as well as the potential effects of EBV on the DN B cells.</p><p>Acknowledgments: This study is supported by the Theme-based Research Scheme (TBRS) (T12-703/22-R).</p>