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939 Unique profile of memory-derived regulatory B cells in viral-driven hepatocellular carcinoma
  1. Shiyong Neo and
  2. Kong-Peng Lam
  1. Agency for Science, Technology and Research, Singapore, Singapore
  • Journal for ImmunoTherapy of Cancer (JITC) preprint. The copyright holder for this preprint are the authors/funders, who have granted JITC permission to display the preprint. All rights reserved. No reuse allowed without permission.


Background The functional plasticity of tumor-infiltrating B (TIL-B) cells spans from an array of anti-tumor responses to non-canonical immune suppression but yet, its influence on the tumor microenvironment (TME) is often underappreciated. Previous studies demonstrated how humoral immune responses have considerable impact on the tumor microenvironment and to the progressive development of hepatocellular carcinoma (HCC). Despite high dimensional immune profiling of HCC being performed extensively in recent years, the focus of the immune oncology field is largely emphasized on the biology of T cells and myeloid cells.

Methods In the present study, we integrated single cell RNA sequencing and BCR sequencing to reveal the development trajectory of regulatory B cells (Bregs) within tumor and non-tumor tissues with a cohort of HCC patients. To further validate our transcriptomics findings, flow cytometric analysis on a separate validatory patient cohort was performed. Additional in vitro functional assays were also perform to further substantiate our findings.

Results The combinatory analysis of gene expression and BCR clonotypes revealed a developmental trajectory of regulatory B cells (Bregs) across HCC etiologies. Shared regulons between Bregs and plasma cells were also revealed in a single cell gene regulatory network. Furthermore, by flow cytometry (FACS) analysis, the Breg phenotype was found to resemble age-associated B cells (ABCs) to co-express Tbet and CD11c and being the non-switch memory (non-SM) B cells, which expressed higher levels of PD-L1, CD25 and granzyme B. Using in vitro assays, we demonstrated that the presence of HCC tumor cells could induce the upregulation of these suppressive factors on peripheral blood B cells, at the same time, resulting in differential capacity to co-stimulate T cell proliferation.

Conclusions Taken together, our study had for the first time, uncover a potential association between inflammatory ABCs and cancer. We highlighted the significance of Bregs in HCC and their potential involvement in immune tolerance during conventional immune checkpoint therapies. With deeper understanding of viral-acquired memory and the development of immune resistance, harnessing the immunogenicity of viral-driven cancers may pave the way forward for the development of novel therapeutic interventions for HCC.

Acknowledgements We would like to thank the flow cytometry core services of the Singapore Immunology Network and clinical coordinators of National Cancer Centre, Singapore for their technical assistance. This work is funded by the National Research Foundation of Singapore (NRF-CRP26–2021RS-0001) and National Medical Research Council (NMRC/OFLCG/003/2018).

Ethics Approval Peripheral blood were collected from and in accordance with the Health Sciences Authority (Ref. #201306–04). Peripheral blood, non-tumor and tumor tissues were collected from HCC patients in National Cancer Centre, Singapore in accordance with institutional review board (CIRB Ref:2019/2303).

This is an open access article distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited, appropriate credit is given, any changes made indicated, and the use is non-commercial. See

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