Probes for INS

TIM4 has previously been associated with antitumor immunity, yet the pattern of expression and the function of this receptor across human cancer tissues remains poorly explored. Here we combined extensive immunolabeling of human tissues with in-silico analysis of pan-cancer transcriptomic datasets to explore the clinical significance of TIM4 expression. Our results unveil that TIM4 is expressed on a fraction of cavity macrophages (CATIM4+MΦ) of carcinoma patients. Moreover, we uncover a high expression of TIM4 on macrophages of the T-cell zone of the carcinoma-associated tertiary lymphoid structures (TLSTIM4+MΦ). In-silico analysis of a pan-cancer dataset revealed a positive correlation between TIM4 expression and markers of B cells, effector CD8+ T cells and a 12-chemokine signature defining tertiary lymphoid structure. In addition, TLSTIM4+MΦ were enriched in cancers displaying microsatellite instability and high CD8+ T-cell infiltration, confirming their association with immune-reactive tumors. Both CATIM4+MΦ and TLSTIM4+MΦ express FOLR2, a marker of tissue-resident MΦ. However, CATIM4+MΦ had higher expression of the immunosuppressive molecules TREM2, IL10 and TGFβ as compared to TLSTIM4+MΦ. By analyzing a scRNA-seq dataset of tumor-associated myeloid cells we identified two TIM4+FOLR2+ clusters coherent with CATIM4+MΦ and TLSTIM4+MΦ. We defined specific gene-signatures for each subset and found that the CATIM4+ MΦ signature was associated with worse patient survival. In contrast, TLSTIM4+MΦ gene-signature positively correlate with better prognosis. Together these data illustrate that TIM4 marks two distinct macrophage populations with distinct phenotype and tissue localization and that may have opposing roles in tumor immunity.

Laryngeal squamous cell carcinomas (LSCCs) typically have an excellent prognosis for stage I tumors but a significant risk of locoregional and distant recurrence for intermediate to advanced disease. This study will investigate the clinical relevance of the tumor microenvironment in a large cohort of treatment-naïve patients affected by stage II-IV LSCC.Whole slide-based digital pathology analysis was applied to measure six immune cell populations identified by immunohistochemistry (IHC) staining for CD3, CD8, CD20, CD66b, CD163 and CD38. Survival analysis was performed by Cox proportional hazards models and unsupervised hierarchical clustering using the k-means method. Double IHC staining and in-situ hybridisation by RNAscope allowed further analysis of a protumoral B cell population.A cohort of 98 patients was enrolled and analysed. The cluster of immune-infiltrated LSCCs demonstrated a significantly worse disease-specific survival rate. We also discovered a new association between high CD20+ B cells and a greater risk of distant recurrence. The phenotypic analysis of infiltrating CD20+ B cells showed a naïve (BCL6-CD27-Mum1-) regulatory phenotype, producing TGFβ but not IL10, according to an active TGFβ pathway, as proved by positive pSMAD2 staining.The identification of regulatory B cells in the context of LSCC, along with the activation of the TGFβ pathway, could provide the basis for new trials investigating the efficacy of already available molecules targeting the TGFβ pathway in the treatment of LSCC.