Multiplexed imaging mass cytometry elicits p-S6 as a therapeutic and prognostic factor in diffuse large B-cell lymphoma.

Diffuse large B-cell lymphoma (DLBCL) is a heterogeneous disease characterized by distinct morphological, genetic, and clinical features. DLBCL with Myc/Bcl-2 double-expression (DE) has been positively correlated with poor prognosis and a low response rate to chemotherapy. However, molecular mechanisms underlying the malignant progression of DE DLBCL, and the tumor microenvironment of DE DLBCL has not been fully elucidated. In this study, we assessed protein expression in 18 DLBCL patient samples using imaging mass cytometry with a panel of 26 metal-tagged antibodies, identifying eight cell types within the tumor microenvironment. Although the ratio of immune cells did not significantly differ between DE and non-DE tissues, a strong interaction between fibroblasts and exhausted CD8+ T cells was specifically observed in DE tissues. By comparing protein expression ratios between DE and non-DE tissues, we found that p-S6 levels were significantly higher in B cells and fibroblasts of DE samples compared to non-DE samples. In our retrospective study, p-S6 independently predicted an inferior prognosis in DLBCL patients (n=71, HR = 5.758; 95% CI: 1.297-25.558; p=0.021). Additionally, co-culture with fibroblasts accelerated the in vitro and in vivo growth of DLBCL cells, and enrichment of the mTOR/S6 pathways was identified using bulk RNA sequencing. Conversely, specific inhibition of p-S6 suppressed DLBCL cell proliferation and exhibited synergistic effects with chemotherapy in vivo. In summary, our findings elucidated the specific tumor microenvironment in DE DLBCL tissues and identified p-S6 as a promising therapeutic target to enhance the efficacy of chemotherapy.