Dataset Information

MET overexpression contributes to STAT4-PD-L1 signaling activation associated with tumor-associated, macrophages-mediated immunosuppression in primary glioblastomas.

ABSTRACT:

Background

Dysregulated receptor tyrosine kinases, such as the mesenchymal-epidermal transition factor (MET), have pivotal role in gliomas. MET and its interaction with the tumor microenvironment have been previously implicated in secondary gliomas. However, the contribution of MET gene to tumor cells' ability to escape immunosurveillance checkpoints in primary gliomas, especially in glioblastoma (GBM), which is a WHO grade 4 glioma with the worst overall survival, is still poorly understood.

Methods

We investigated the relationship between MET expression and glioma microenvironment by using multiomics data and aimed to understand the potential implications of MET in clinical practice through survival analysis. RNA expression data from a total of 1243 primary glioma samples (WHO grades 2-4) were assembled, incorporating The Cancer Genome Atlas, Chinese Glioma Genome Atlas, and GSE16011 data sets.

Results

Pearson's correlation test from the three data sets indicated that MET showed a robust correlation with programmed death-ligand 1 (PD-L1) and STAT pathways. Western blot analysis revealed that in GBM cell lines (N33 and LN229), PD-L1 and phosphorylated STAT4 were upregulated by MET activation treatment with hepatocyte growth factor and were downregulated on MET suppression by PLB-1001. Tumor tissue microarray analysis indicated a positive correlation between MET and PD-L1 and macrophage-associated markers. Chromatin immunoprecipitation-PCR assay showed enrichment of STAT4 in the PD-L1 DNA. Transwell co-culture and chemotaxis assays revealed that knockdown of MET in GBM cells inhibited macrophage chemotaxis. Moreover, we performed CIBERSORTx and single-cell RNA sequencing data analysis which revealed an elevated number of macrophages in glioma samples with MET overexpression. Kaplan-Meier survival analysis indicated that activation of the MET/STAT4/PD-L1 pathway and upregulation of macrophages were associated with shorter survival time in patients with primary GBM.

Conclusions

These data indicated that the MET-STAT4-PD-L1 axis and tumor-associated macrophages might enforce glioma immune evasion and were associated with poor prognosis in GBM samples, suggesting potential clinical strategies for targeted therapy combined with immunotherapy in patients with primary GBM.

SUBMITTER: Wang QW

PROVIDER: S-EPMC8527154 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BACKGROUND:Tumor-associated macrophages (TAMs) facilitate tumor progression via establishment of an immunosuppressive tumor microenvironment (TME). However, it is poorly understood how tumor cells could functionally modulate TAMs. Our previous work indicated that tumor cell-released autophagosomes (TRAPs), a type of LC3-II+ double-membrane extracellular vesicles (EVs) was sufficient to suppress anti-tumor immune responses by inducing IL-10-producing B cells and immune suppressive neutrophils. Here, we hypothesized that TRAPs may participate in regulating macrophage polarization. METHODS:TRAPs isolated from multiple murine tumor cell lines and pleural effusions or ascites of cancer patients were incubated with bone marrow-derived macrophages (BMDMs) and monocytes, respectively. Cellular phenotypes were examined by flow cytometry, ELISA and quantitative PCR. TRAPs treated BMDMs were tested for the ability to suppress T-cell proliferation in vitro, and for promotion of tumor growth in vivo. Transwell chamber and neutralization antibodies were added to ascertain the inhibitory molecules expressed on BMDMs exposed to TRAPs. Knockout mice were used to identify the receptors responsible for TRAPs-induced BMDMs polarization and the signaling mechanism was examined by western blot. Autophagy-deficient tumors were profiled for phenotypic changes of TAMs and IFN-? secretion of T cells by flow cytometry. The phenotype of monocytes from pleural effusions or ascites of cancer patients was assessed by flow cytometry. RESULTS:TRAPs converted macrophages into an immunosuppressive M2-like phenotype characterized by the expression of PD-L1 and IL-10. These macrophages inhibited the proliferation of both CD4+ and CD8+ T cells in vitro, and promoted tumor growth mainly through PD-L1 in vivo. TRAPs-induced macrophage polarization was dependent on TLR4-mediated MyD88-p38-STAT3 signaling. In vivo studies indicated that disruption of autophagosome formation in B16F10 cells by silencing the autophagy gene Beclin1 resulted in a remarkable delay in tumor growth, which was associated with reduced autophagosome secretion, TAMs reprogramming and enhanced T cell activation. Moreover, the levels of LC3B+ EVs appeared to correlate significantly with up-regulation of PD-L1 and IL-10 in matched monocytes from effusions or ascites of cancer patients, and TRAPs isolated from these samples could also polarize monocytes to an M2-like phenotype with increased expression of PD-L1, CD163 and IL-10, decreased expression of HLA-DR, and T cell-suppressive function. CONCLUSIONS:These findings suggest the TRAPs-PD-L1 axis as a major driver of immunosuppression in the TME by eliciting macrophage polarization towards an M2-like phenotype, and highlight the potential novel therapeutic approach of simultaneously targeting autophagy and PD-L1.