TCF1+CD4+ T cells and microglia co-orchestrate tertiary lymphoid structures to enhance prognosis and immunotherapy in non-small cell lung cancer with brain metastases

BACKGROUND: Non-small cell lung cancer brain metastases (NSCLC-BM) confer a poor prognosis, with only a minority of patients responding to immune checkpoint inhibitors (ICIs). Tertiary lymphoid structures (TLSs) have been linked to prognosis and response to immunotherapy across various tumours. Given the unique immune environment of brain metastases, TLSs within NSCLC-BM might exhibit distinct characteristics. Nevertheless, the function and formation mechanisms of these intracranial TLSs remain inadequately understood. METHODS: We retrospectively analysed 120 resected NSCLC-BM samples. Heterogeneity of TLS characteristics and functional states in NSCLC-BM was assessed using H&E and multiplex immunohistochemistry (mIHC). Associations with OS and iPFS were evaluated, and a nomogram incorporating TLS score was developed. Potential mechanisms underlying TLS formation were explored using public scRNA-seq datasets and mIHC, and were functionally validated in vivo in the LC-BrM mouse model. FINDINGS: TLSs were primarily intratumoral and immature. Elevated intratumoral TLS scores independently predicted prolonged OS and iPFS, and predicted benefit from postoperative ICIs. TLS-rich tumours exhibited increased CD8+ T cells, FOXP3-CD4+ T cells, and B cells. Mechanistically, TCF1+CD4+ T cells and microglia co-localised within TLSs and might facilitate their formation via LTβ/LTβR signalling. Meanwhile, stromal CXCL12 was further identified as a crucial chemoattractant for these cells and correlated with favourable outcomes of ICI therapy. Functionally, the combination of rmCXCL12 and anti-PD-1 promotes TLS formation and intracranial tumour control in an LTβR-dependent manner. INTERPRETATION: These findings highlight the significance of TLSs as prognostic biomarkers and potential therapeutic targets in patients with NSCLC-BM. FUNDING: This work was supported by the Natural Science Foundation of China (82573031); the CQMU Program for Youth Innovation in Future Medicine (W0172); the Postdoctoral Fellowship Program of the China Postdoctoral Science Foundation (GZC20233356).