Effect of Tertiary Lymphoid Structures on Prognosis of Patients with Hepatocellular Carcinoma and Preliminary Exploration of Its Formation Mechanism

Abstract

Background: Tertiary lymphoid structures (TLSs) are formed by the aggregation of tumour-infiltrating lymphocytes (TILs), which is driven by chemokines or cytokines in the tumour microenvironment. Studies have shown that TLSs are associated with good prognosis in patients with various solid tumours and can improve patient responses to immunotherapy. However, the role of TLSs in hepatocellular carcinoma (HCC) remains controversial, and the underlying molecular mechanism is unclear. Methods: According to haematoxylin-eosin (HE) staining results, HCC patients in Xijing Hospital data and TCGA data were divided into TLS+ and TLS- groups, and Kaplan–Meier (KM) analysis was performed to assess overall survival (OS) and recurrence-free survival (RFS). Immunofluorescence (IF) and immunohistochemistry (IHC) were used to identify TILs in the TLS+ group. Lymphocyte-specific protein tyrosine kinase (LCK), a molecule involved in TLS formation, was explored in LinkedOmics. TILs were divided into two groups by drawing receiver operating characteristic (ROC) curves to calculate cut-off values. Spearman correlation analysis was used to calculate the correlation between LCK and TILs, and the molecular pathways by which LCK regulates immunotherapy were clarified through enrichment analysis. The half-maximal inhibitory concentration (IC50) distribution of sorafenib was observed in groups that varied in LCK expression. Results: According to the HE results, 61 cases in the Xijing Hospital cohort and 195 cases in the TCGA cohort had TLSs, while 89 cases and 136 cases did not. The KM results showed that TLSs had no effect on the OS of HCC patients but significantly affected RFS. The IF/IHC results showed that higher TIL numbers in TLSs were correlated with better prognosis in HCC patients. Spearman correlation analysis showed that LCK expression was positively correlated with TIL numbers. Enrichment analysis showed that upregulation of LCK expression mainly regulated the cytokine signalling pathway, the chemokine signalling pathway and T-cell activation. The IC50 scores of sorafenib in HCC patients with high LCK expression were lower, and the sensitivity was higher. Conclusion: TLSs mainly affected the early RFS of HCC patients but had no effect on OS. The high expression of the TLS formation-related gene LCK can increase the sensitivity of HCC patients to ICIs.

Tertiary lymphoid structures in cancer: immune mechanisms and clinical implications

“…Published in December 2023, a meta‐analysis focusing on digestive system tumors further confirms the association between intratumoral localization of TLS and improved survival outcomes 91 . It is noteworthy that TLS derived from inflammation or the tumor itself may exhibit distinct functionalities, with the former creating a favorable environment for the growth of malignant cells, 92 while tumor‐induced TLS usually contributes to antitumor immunity 93 . The spatial distribution of immune cells within TLS also warrants investigation.…”

“… 91 It is noteworthy that TLS derived from inflammation or the tumor itself may exhibit distinct functionalities, with the former creating a favorable environment for the growth of malignant cells, 92 while tumor‐induced TLS usually contributes to antitumor immunity. 93 The spatial distribution of immune cells within TLS also warrants investigation. A study focusing on the spatial distribution of tumor‐associated neutrophils in bladder cancer patients revealed a decrease in the density of these immunosuppressive cells with increasing distance from TLS.…”

Prognostic value of tertiary lymphoid structures (TLS) in digestive system cancers: a systematic review and meta-analysis

“…Absence No OS, DFS Ling YH [ 31 ] 2022 China ESCC 394 I-III 2008–2017 Median:68.57 month HE、IHC TLS Presence vs. Absence No OS, DFS Jiang Q [ 32 ] 2022 China GC 292 I-IV 1983–2016 Until 2020.1 HE、IHC Mature TLS High vs. Low Yes OS Kemi N [ 33 ] 2022 Finland GC 721 I-IV 2002–2008 Median: 28.0 month HE TLS diameter High vs. Low NA OS Zhan Z [ 34 ] 2022 China CRC 203 I-IV 2014.01–2017.07 Median: 50.0 month IHC TLS High vs. None or Low Yes OS, DFS Li JH [ 35 ] 2022 China HCC 150 I-...…”

The combination of TLSs and the neutrophil-to-macrophage ratio predicts early recurrence in patients with hepatocellular carcinoma

“…TLSs, as lymphocyte aggregates found in nonlymphoid areas, are known to be involved in chronic in ammation, transplant rejection, autoimmune in ammation, and tumors [1,7,10,11,25,26]. They share functional and developmental similarities with lymph nodes and play a role in local immune function as part of the tumor microenvironment (TME) [10].…”

“…TLSs, also known as ectopic lymph node structures, are speci c aggregations of lymphocytes in nonlymphoid tissues and are frequently found in conditions such as chronic in ammation, transplant rejection reactions, autoimmune diseases, and cancers [7,10,11]. Like secondary lymph nodes, TLSs likely function similarly to secondary lymph nodes, leading most researchers to believe that TLSs play a role in HCC similar to that of lymph nodes [12,13].…”