DNA damage repair kinase DNA-PK and cGAS synergize to induce cancer-related inflammation in glioblastoma.

Cytosolic DNA promotes inflammatory responses upon detection by the cyclic GMP-AMP (cGAMP) synthase (cGAS). It has been suggested that cGAS downregulation is an immune escape strategy harnessed by tumor cells. Here, we used glioblastoma cells that show undetectable cGAS levels to address if alternative DNA detection pathways can promote pro-inflammatory signaling. We show that the DNA-PK DNA repair complex (i) drives cGAS-independent IRF3-mediated type I Interferon responses and (ii) that its catalytic activity is required for cGAS-dependent cGAMP production and optimal downstream signaling. We further show that the cooperation between DNA-PK and cGAS favors the expression of chemokines that promote macrophage recruitment in the tumor microenvironment in a glioblastoma model, a process that impairs early tumorigenesis but correlates with poor outcome in glioblastoma patients. Thus, our study supports that cGAS-dependent signaling is acquired during tumorigenesis and that cGAS and DNA-PK activities should be analyzed concertedly to predict the impact of strategies aiming to boost tumor immunogenicity.

Protocol to induce and assess cGAS-STING pathway activation in vitro.

The cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) pathway plays a pivotal role in several cellular processes including pathogen recognition and inflammatory responses. We describe a protocol to activate the cGAS-STING pathway in murine cells using nucleic acids transfection. We describe how to prepare the nucleic acid probes and validate activation of the pathway by western blot and gene expression analysis. The protocol can be applied to investigate cGAS-STING signaling in both murine and human cell lines. For complete details on the use and execution of this protocol, please refer to Vila et al. (2022).

Activation of STING in the pancreatic tumor microenvironment: A novel therapeutic opportunity.

Pancreatic ductal adenocarcinoma (PDAC) is a cancer of poor prognosis that presents with a dense desmoplastic stroma that contributes to therapeutic failure. PDAC patients are mostly unresponsive to immunotherapy. However, hopes to elicit response to immunotherapy have emerged with novel strategies targeting the Stimulator of Interferon Genes (STING) protein, which is a major regulator of tumor-associated inflammation. Combination of STING agonists with conventional immunotherapy approaches has proven to potentiate therapeutic benefits in several cancers. However, recent data underscore that the output of STING activation varies depending on the cellular and tissue context. This suggests that tumor heterogeneity, and in particular the heterogeneity of the tumor microenvironment (TME), is a key factor determining whether STING activation would bear benefits for patients. In this review, we discuss the potential benefits of STING activation in PDAC. To this aim, we describe the major components of the PDAC TME, and the expected consequences of STING activation.

STING orchestrates the crosstalk between polyunsaturated fatty acid metabolism and inflammatory responses.

Concerted alteration of immune and metabolic homeostasis underlies several inflammation-related pathologies, ranging from metabolic syndrome to infectious diseases. Here, we explored the coordination of nucleic acid-dependent inflammatory responses and metabolic homeostasis. We reveal that the STING (stimulator of interferon genes) protein regulates metabolic homeostasis through inhibition of the fatty acid desaturase 2 (FADS2) rate-limiting enzyme in polyunsaturated fatty acid (PUFA) desaturation. STING ablation and agonist-mediated degradation increased FADS2-associated desaturase activity and led to accumulation of PUFA derivatives that drive thermogenesis. STING agonists directly activated FADS2-dependent desaturation, promoting metabolic alterations. PUFAs in turn inhibited STING, thereby regulating antiviral responses and contributing to resolving STING-associated inflammation. Thus, we have unveiled a negative regulatory feedback loop between STING and FADS2 that fine-tunes inflammatory responses. Our results highlight the role of metabolic alterations in human pathologies associated with aberrant STING activation and STING-targeting therapies.

Nucleic Acid Immunity and DNA Damage Response: New Friends and Old Foes.

The maintenance of genomic stability in multicellular organisms relies on the DNA damage response (DDR). The DDR encompasses several interconnected pathways that cooperate to ensure the repair of genomic lesions. Besides their repair functions, several DDR proteins have emerged as involved in the onset of inflammatory responses. In particular, several actors of the DDR have been reported to elicit innate immune activation upon detection of cytosolic pathological nucleic acids. Conversely, pattern recognition receptors (PRRs), initially described as dedicated to the detection of cytosolic immune-stimulatory nucleic acids, have been found to regulate DDR. Thus, although initially described as operating in specific subcellular localizations, actors of the DDR and nucleic acid immune sensors may be involved in interconnected pathways, likely influencing the efficiency of one another. Within this mini review, we discuss evidences for the crosstalk between PRRs and actors of the DDR. For this purpose, we mainly focus on cyclic GMP-AMP (cGAMP) synthetase (cGAS) and Interferon Gamma Inducible Protein 16 (IFI16), as major PRRs involved in the detection of aberrant nucleic acid species, and components of the DNA-dependent protein kinase (DNA-PK) complex, involved in the repair of double strand breaks that were recently described to qualify as potential PRRs. Finally, we discuss how the crosstalk between DDR and nucleic acid-associated Interferon responses cooperate for the fine-tuning of innate immune activation, and therefore dictate pathological outcomes. Understanding the molecular determinants of such cooperation will be paramount to the design of future therapeutic approaches.