Purine nucleoside phosphorylase (PNP) enables the breakdown and recycling of guanine nucleosides. PNP insufficiency in humans is paradoxically associated with both immunodeficiency and autoimmunity, but the mechanistic basis for these outcomes is incompletely understood. Here, we identify two immune lineage-dependent consequences of PNP inactivation dictated by distinct gene interactions. During T cell development, PNP inactivation is synthetically lethal with downregulation of the dNTP triphosphohydrolase SAMHD1. This interaction requires deoxycytidine kinase activity and is antagonized by microenvironmental deoxycytidine. In B lymphocytes and macrophages, PNP regulates Toll-like receptor 7 signaling by controlling the levels of its (deoxy)guanosine nucleoside ligands. Overriding this regulatory mechanism promotes germinal center formation in the absence of exogenous antigen and accelerates disease in a mouse model of autoimmunity. This work reveals that one purine metabolism gene protects against immunodeficiency and autoimmunity via independent mechanisms operating in distinct immune lineages and identifies PNP as a potentially novel metabolic immune checkpoint.
Rationale: Stimulator of Interferon Genes (STING) is a mediator of immune recognition of cytosolic DNA, which plays important roles in cancer, cytotoxic therapies, and infections with certain pathogens. While pharmacological STING activation stimulates potent anti-tumor immune responses in animal models, clinically applicable pharmacodynamic biomarkers that inform of the magnitude, duration, and location of immune activation elicited by systemic STING agonists are yet to be described. We investigated whether systemic STING activation induces metabolic alterations in immune cells that can be visualized by Positron Emission Tomography (PET) imaging.Methods: C57BL/6 mice were treated with systemic STING agonists and imaged with 18 Ffluorodeoxyglucose ( 18 F-FDG) PET after 24 hours. Splenocytes were harvested 6 hours after STING agonist administration and analyzed by single-cell RNA sequencing and flow cytometry. 18 F-FDG uptake in total splenocytes and immunomagnetically enriched splenic B and T lymphocytes from STING agonist-treated mice was measured by gamma counting. In mice bearing prostate or pancreas cancer tumors, the effects of STING agonist treatment on 18 F-FDG uptake, T lymphocyte activation marker levels, and tumor growth were evaluated.Results: Systemic delivery of structurally distinct STING agonists in mice significantly increased 18 F-FDG uptake in the spleen. The average spleen SUVmax in control mice was 1.90 (range 1.56-2.34) compared to 4.55 (range 3.35-6.2) in STING agonist-treated mice (P < 0.0001).Single-cell transcriptional and flow cytometry analyses of immune cells from systemic STING agonist-treated mice revealed enrichment of a glycolytic transcriptional signature in both T and B lymphocytes which correlated with the induction of immune cell activation markers. In tumorbearing mice, STING agonist administration significantly delayed tumor growth and increased 18 F-FDG uptake in secondary lymphoid organs.
Conclusion:These findings reveal hitherto unknown functional links between STING signaling and immunometabolism and suggest that 18 F-FDG PET could provide a widely applicable approach to measure the pharmacodynamic effects of systemic STING agonists at a wholebody level and guide their clinical development.
Purpose: STING (Stimulator of Interferon Genes) agonists are currently in development for treatment of solid tumors, including pancreatic ductal adenocarcinoma (PDAC). Response rates to STING agonists alone have been promising yet modest and combination therapies will likely be required to elicit their full potency. We sought to identify combination therapies and mechanisms that augment the tumor-cell intrinsic effect of therapeutically relevant STING agonists apart from their known effects on tumor immunity. Experimental Design: We screened 430 kinase inhibitors to identify synergistic effectors of tumor cell death with diABZI, an intravenously administered and systemically available STING agonist. We deciphered the mechanisms of synergy with STING agonism that cause tumor cell death in vitro and tumor regression in vivo. Results: We found that MEK inhibitors caused the greatest synergy with diABZI and that this effect was most pronounced in cells with high STING expression. MEK inhibition enhanced the ability of STING agonism to induce Type I interferon-dependent cell death in vitro and tumor regression in vivo. We parsed NF-κB-dependent and independent mechanisms that mediate STING-driven Type I interferon production and show that MEK signaling inhibits this effect by suppressing NF-κB activation. Conclusions: Our results highlight the cytotoxic effects of STING agonism on PDAC cells that are independent of tumor immunity, and that these therapeutic benefits of STING agonism can be synergistically enhanced by MEK inhibition.
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