Fig. 4: α-KG-OXGR1-MAPK signaling pathway inhibits macrophage antigen presentation function.

a Gene Ontology (GO) analysis of RNA-seq after 24 h co-culture of tumor CM with or without α-KG (8 μM) and THP-1 cells. b Volcano plot to depict the downregulated HLA-DRA, HLA-DPA1, and HLA-DQA1 expression changes of immune response observed in the RNA-seq data. c Gene Set Enrichment Analysis (GSEA) indicating that positive regulation of MAPK cascade was significantly upregulated after α-KG accumulation. d Western blot analysis showed the expression of the MAPK pathway and HLA-DR, CD74 changes after 24 h co-culture (n = 3 biological replicates). The samples derive from the same experiment but different gels for p-ERK, ERK, and HLA-DR, another for CD74 and p-P38, another for p-JNK, JNK, and another for P38 and α-Tubulin were processed in parallel. e Diagram of interaction between α-KG and OXGR1 protein using molecular docking. f Western blot analysis showed the expression of p-ERK, HLA-DR, and CD74 changes after 24 h co-culture (n = 3 biological replicates). The samples derive from the same experiment but different gels for p-ERK, ERK, and HLA-DR, another for CD74, and another for OXGR1 and α-Tubulin were processed in parallel. g Schematic diagram of THP-1 with or without OXGR1 depletion and tumor CM with α-KG (0, 8 μM). Created in BioRender. Zhang, N. (2025) https://BioRender.com/a10o519. After 24 h, macrophages were used for flow analysis. h Flow cytometry analysis of HLA-DR⁺ MFI values from data (g) (n = 4 biological replicates). i Schematic diagram of subcutaneous tumor treatment. Created in BioRender. Zhang, N. (2025) https://BioRender.com/j80q160. And representative images of vehicle, montelukast, α-KG or combined on day 26 (n = 5 biologically independent samples). j Tumor growth of LTP-C9 with overexpression of PDHA1 WT were injected into C57BL/6 mice (n = 5 biologically independent samples), with vehicle, montelukast, α-KG or combined treatment, beginning on day 14. k Tumor volume from data (i) (n = 5 biologically independent samples). l Tumor weight from data (i) (n = 5 biologically independent samples). Data are presented as mean values ± SD, and P values were calculated using two-tailed unpaired Student’s t-tests (d, f, h, k, l) or 2way ANOVA (j). Each Western blot was independently repeated three times. Mac macrophages, CM conditioned medium, MFI mean fluorescence intensity. Source data are provided as a Source Data File.