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Immune landscape of tertiary lymphoid structures in hepatocellular carcinoma (HCC) treated with neoadjuvant immune checkpoint blockade

Shu D.H. et al. (BioRxiv) DOI: 10.1101/2023.10.16.562104

Immune landscape of tertiary lymphoid structures in hepatocellular carcinoma (HCC) treated with neoadjuvant immune checkpoint blockade


  • Tertiary lymphoid structures

  • Neoadjuvant immune checkpoint blockade

  • Hepatocellular carcinoma

Main Findings

This study focuses on tertiary lymphoid structures (TLS) formation after neoadjuvant immune checkpoint blockade (ICB) treatment. Shu et al. found that the density of TLS in tumors was highest in patients that responded to ICB and was associated with significantly longer relapse-free survival. Notably, by investigating immune cell populations by a combination of bulk RNA-sequencing, and imaging mass cytometry, they identify two TLS types: mature TLS with a classical germinal center and involuted TLS characterized by a ring of B cells.

The obtained results show that involuted TLS are associated with complete tumor responses and B cells undergoing antigen-driven positive selection. The authors identified T peripheral helper (Tph) cells (CD4+ T cell subset) within tumor-associated lymphoid structures to significantly influence immune responses.

The authors then used laser capture of individual TLS to sequence both T- and B-cell receptors. High density TLS following ICB was associated with expanded T and B cell repertoire. Importantly, they found that while most B cell clones are TLS-specific, the most abundant T cell clones were shared between the different TLS of the same tumor. By comparing TCR clonotypes between peripheral blood (obtained by single-cell RNA-sequencing) and tumor associated TLS, the authors undertake to infer T cell phenotype present in the tumor. They confirmed the presence in both mature and involuted TLS of abundant CD8 T cell effector memory (TEM) populations exhibiting high clonality and clonal expansion within the tumors, with some showing cytotoxic features. These results deepen our understanding of TLS formation and their role in immune cell dynamics after neoadjuvant ICB therapy.


  • The inference of T cell phenotype based on shared TCR clones between peripheral blood and TLS would benefit further validation.

  • Low amount of involuted TLS samples (n=3) may hinder the assessment of the TCR diversity.

  • The study of low density TLS by imaging mass cytometry in partially responding patients may be of interest to understand the formation of intratumoral TLS and the impact of ICB.


This study illuminates the role of tertiary lymphoid structures (TLS) forming in the tumors of hepatocellular carcinoma (HCC) patients that received pre-operative neoadjuvant immunotherapy. Neoadjuvant immunotherapy uses the primary tumor to enhance antitumor immunity and reduce the risk of cancer recurrence after surgery. While this approach has shown promise, that the mechanisms involved in the contribution of TLS were not well understood. The study's findings reveal that in HCC treated with neoadjuvant immunotherapy, intratumoral TLS are associated with superior pathologic responses and improved relapse-free survival, aligning with similar data from other solid tumors treated with immune checkpoint blockade (ICB).

Additionally, the research sheds light on the dissolution of TLS, which may play a functional role in supporting the contraction and memory phase of the intratumoral adaptive immune response. The identification of T cell populations within TLS, such as the CXCR5-CXCR3+ CD4+ T cells, suggests potential for tailored treatment approaches based on TLS involvement. While the study focused on HCC, the findings may have broader implications in the field of oncology, potentially influencing treatment approaches for various cancer types. The pre-operative neoadjuvant ICB could potentially be used to induce TLS in various tumors, launching an early anticancer response and potentially increasing the long-term survival of patients.


Reviewed by Ramojus Balevicius as part of a cross-institutional journal club between the Icahn School of Medicine at Mount Sinai, the University of Oxford, the Karolinska Institute and the University of Toronto.

The authors declare no conflict of interests in relation to their involvement in the review.

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