Intratumoral CXCL13⁺ CD160⁺ CD8⁺ T cells promote the formation of tertiary lymphoid structures to enhance the efficacy of immunotherapy in advanced gastric cancer

Background: Stage IV gastric cancer is a highly heterogeneous and lethal tumor with few therapeutic strategies. The combination of programmed cell death protein 1 inhibitors and chemotherapy is currently the standard frontline treatment regimen for advanced gastric cancer. Nevertheless, it remains a great challenge to screen the beneficiaries of immunochemotherapy and expand indications for this treatment regimen.

Methods: We conducted a pathological assessment to ascertain the importance of tertiary lymphoid structures based on the tissue samples collected from patients with stage IV gastric cancer (n=15) both prior to and following immunochemotherapy treatment. Additionally, we used spatial (n=10) and single-cell transcriptional analysis (n=97) to investigate the key regulators of tertiary lymphoid structures (TLSs). Multiplex immunofluorescence and image analysis (n=34) were performed to explore the association between tumor-infiltrating CXCL13⁺ CD160⁺ CD8⁺ T cells and TLSs. The relationship between CXCL13⁺ CD160⁺ CD8⁺ T cells and the responsiveness to immunotherapy was also evaluated by multiplex immunofluorescence and image analysis approaches (n=15). Furthermore, we explored the intrinsic characteristics of CXCL13⁺ CD160⁺ CD8⁺ T cells through various experimental techniques, including quantitative reverse transcription-PCR, western blot, and flow cytometry.

Results: We found that responders exhibited higher levels of TLSs and CXCL13⁺ CD160⁺ CD8⁺ T cells in biopsy tissues prior to immunochemotherapy compared with non-responders. Following conversion therapy, responders also had a higher percentage of mature TLSs and a higher number of CXCL13⁺ CD160⁺ CD8⁺ T cells in surgical resections. Moreover, we discovered that vitamin B₆ in CD160⁺ CD8⁺ T cells could reduce the ubiquitination modification of HIF-1α by MDM2, thereby attenuating the degradation of HIF-1α. Consequently, this led to the transcriptional upregulation of CXCL13 expression, facilitating the recruitment of CXCR5⁺ B cells and the formation of TLSs.

Conclusion: The number and maturity of TLSs, along with the extent of CXCL13⁺ CD160⁺ CD8⁺ T-cell infiltration, might function as potential indicators for assessing the effectiveness of immunotherapy in treating gastric malignancies. Furthermore, our research suggests that vitamin B₆ could enhance the secretion of CXCL13 by CD160⁺ CD8⁺ T cells by reducing the degradation of HIF-1α. Additionally, we demonstrate that vitamin B₆ supplementation or targeting pyridoxal kinase could substantially improve the efficacy of immunotherapies for gastric cancer.