The XOR-IDH3α axis controls macrophage polarization in hepatocellular carcinoma

•Loss of XOR in monocyte-derived TAMs rather than resident Kupffer cells promotes the progression of liver cancer. •Loss of XOR increases α-KG generation in monocyte-derived TAMs by increasing the activity of IDH3α. •Loss of XOR enhances the production of adenosine and KYNA in monocyte-derived TAMs via increased activity of IDH3α. •XOR-null monocyte-derived TAMs promote the exhaustion of CD8+ T cells via immunosuppressive metabolites including adenosine and KYNA. Background & Aims Tumor-associated macrophages (TAMs) are indispensable in the hepatocellular carcinoma (HCC) tumor microenvironment. Xanthine oxidoreductase (XOR), also known as xanthine dehydrogenase (XDH), participates in purine metabolism, uric acid production, and macrophage polarization to a pro-inflammatory phenotype. However, the role of XOR in HCC-associated TAMs is unclear. Methods We evaluated the XOR level in macrophages isolated from HCC tissues and paired adjacent tissues. We established diethylnitrosamine/carbon tetrachloride (CCl4)-induced and orthotopically implanted HCC mouse models using mice with Xdh-specific depletion in the myeloid cell lineage (Xdhf/fLyz2cre) or Kupffer cells (Xdhf/fClec4fcre). We determined metabolic differences using specific methodologies, including metabolomics and metabolic flux. Results We found that XOR expression was downregulated in HCC TAMs and positively correlated with patient survival, which was strongly related to the characteristics of the tumor microenvironment, especially hypoxia. Using HCC-inflicted mice (Xdhf/fLyz2cre and Xdhf/fClec4fcre), we revealed that XOR loss in monocyte-derived TAMs rather than Kupffer cells promoted their M2 polarization and CD8+ T-cell exhaustion, which exacerbated HCC progression. In addition, the tricarboxylic acid cycle was disturbed, and the generation of α-ketoglutarate was enhanced within XOR-depleted macrophages. XOR inhibited α-ketoglutarate production by interacting with IDH3α catalytic sites (K142 and Q139). The increased IDH3α activity caused increased adenosine and kynurenic acid production in TAMs, which enhanced the immunosuppressive effects of TAMs and CD8+ T cells. Conclusions The XOR-IDH3α axis mediates TAM polarization and HCC progression and may be a small-molecule therapeutic or immunotherapeutic target against suppressive HCC TAMs. Impact and implications Immunotherapies have been widely applied to the treatment of hepatocellular carcinoma (HCC), but to date they have been associated with unsatisfactory efficacy. The tumor microenvironment of HCC is full of different infiltrating immune cells. Tumor-associated macrophages (TAMs) are vital components in the tumor microenvironment and are involved in HCC progression. Herein, we confirm the downregulation of XOR expression in TAMs isolated from human HCC. The loss of XOR in monocyte-derived macrophages increases IDH3 activity and results in an increase in α-ketoglutarate production, which can promote M2-like polarization. Additionally, XOR-null TAMs derived from monocytes promote CD8+ T-cell exhaustion via the upregulation of immunosuppressive metabolites, including adenosine and kynurenic acid. Given the prevalence and high rate of incidence of HCC and the need for improved therapeutic options for patients, our findings identify potential therapeutic targets that may be further studied to develop improved therapies. Tumor-associated macrophages (TAMs) are indispensable in the hepatocellular carcinoma (HCC) tumor microenvironment. Xanthine oxidoreductase (XOR), also known as xanthine dehydrogenase (XDH), participates in purine metabolism, uric acid production, and macrophage polarization to a pro-inflammatory phenotype. However, the role of XOR in HCC-associated TAMs is unclear. We evaluated the XOR level in macrophages isolated from HCC tissues and paired adjacent tissues. We established diethylnitrosamine/carbon tetrachloride (CCl4)-induced and orthotopically implanted HCC mouse models using mice with Xdh-specific depletion in the myeloid cell lineage (Xdhf/fLyz2cre) or Kupffer cells (Xdhf/fClec4fcre). We determined metabolic differences using specific methodologies, including metabolomics and metabolic flux. We found that XOR expression was downregulated in HCC TAMs and positively correlated with patient survival, which was strongly related to the characteristics of the tumor microenvironment, especially hypoxia. Using HCC-inflicted mice (Xdhf/fLyz2cre and Xdhf/fClec4fcre), we revealed that XOR loss in monocyte-derived TAMs rather than Kupffer cells promoted their M2 polarization and CD8+ T-cell exhaustion, which exacerbated HCC progression. In addition, the tricarboxylic acid cycle was disturbed, and the generation of α-ketoglutarate was enhanced within XOR-depleted macrophages. XOR inhibited α-ketoglutarate production by interacting with IDH3α catalytic sites (K142 and Q139). The increased IDH3α activity caused increased adenosine and kynurenic acid production in TAMs, which enhanced the immunosuppressive effects of TAMs and CD8+ T cells. The XOR-IDH3α axis mediates TAM polarization and HCC progression and may be a small-molecule therapeutic or immunotherapeutic target against suppressive HCC TAMs.