Immunologie et Neurogénétique Expérimentales et Moléculaires

Cyclophosphamide is one of several clinically important cancer drugs whose therapeutic efficacy is due in part to their ability to stimulate antitumor immune responses. Studying mouse models, we demonstrate that cyclophosphamide alters the composition of microbiota in the small intestine and induces the translocation of selected species of Gram-positive bacteria into secondary lymphoid organs. There, these bacteria stimulate the generation of a specific subset of "pathogenic" T helper 17 (pT(H)17) cells and memory T(H)1 immune responses. Tumor-bearing mice that were germ-free or that had been treated with antibiotics to kill Gram-positive bacteria showed a reduction in pT(H)17 responses, and their tumors were resistant to cyclophosphamide. Adoptive transfer of pT(H)17 cells partially restored the antitumor efficacy of cyclophosphamide. These results suggest that the gut microbiota help shape the anticancer immune response.

We have recently shown that low intensity, intermediate frequency, electric fields inhibit by an anti-microtubule mechanism of action, cancerous cell growth in vitro. Using implanted electrodes, these fields were also shown to inhibit the growth of dermal tumors in mice. The present study extends these findings to additional cell lines [human breast carcinoma; MDA-MB-231, and human non-small-cell lung carcinoma (H1299)] and to animal tumor models (intradermal B16F1 melanoma and intracranial F-98 glioma) using external insulated electrodes. These findings led to the initiation of a pilot clinical trial of the effects of TTFields in 10 patients with recurrent glioblastoma (GBM). Median time to disease progression in these patients was 26.1 weeks and median overall survival was 62.2 weeks. These time to disease progression and OS values are more than double the reported medians of historical control patients. No device-related serious adverse events were seen after >70 months of cumulative treatment in all of the patients. The only device-related side effect seen was a mild to moderate contact dermatitis beneath the field delivering electrodes. We conclude that TTFields are a safe and effective new treatment modality which effectively slows down tumor growth in vitro, in vivo and, as demonstrated here, in human cancer patients.

Instability in the composition of gut bacterial communities (dysbiosis) has been linked to common human intestinal disorders, such as Crohn's disease and colorectal cancer. Here, we show that dysbiosis caused by Nod2 deficiency gives rise to a reversible, communicable risk of colitis and colitis-associated carcinogenesis in mice. Loss of either Nod2 or RIP2 resulted in a proinflammatory microenvironment that enhanced epithelial dysplasia following chemically induced injury. The condition could be improved by treatment with antibiotics or an anti-interleukin-6 receptor-neutralizing antibody. Genotype-dependent disease risk was communicable via maternally transmitted microbiota in both Nod2-deficient and WT hosts. Furthermore, reciprocal microbiota transplantation reduced disease risk in Nod2-deficient mice and led to long-term changes in intestinal microbial communities. Conversely, disease risk was enhanced in WT hosts that were recolonized with dysbiotic fecal microbiota from Nod2-deficient mice. Thus, we demonstrated that licensing of dysbiotic microbiota is a critical component of disease risk. Our results demonstrate that NOD2 has an unexpected role in shaping a protective assembly of gut bacterial communities and suggest that manipulation of dysbiosis is a potential therapeutic approach in the treatment of human intestinal disorders.

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Limited experimental evidence bridges nutrition and cancer immunosurveillance. Here, we show that ketogenic diet (KD) - or its principal ketone body, 3-hydroxybutyrate (3HB), most specifically in intermittent scheduling - induced T cell-dependent tumor growth retardation of aggressive tumor models. In conditions in which anti-PD-1 alone or in combination with anti-CTLA-4 failed to reduce tumor growth in mice receiving a standard diet, KD, or oral supplementation of 3HB reestablished therapeutic responses. Supplementation of KD with sucrose (which breaks ketogenesis, abolishing 3HB production) or with a pharmacological antagonist of the 3HB receptor GPR109A abolished the antitumor effects. Mechanistically, 3HB prevented the immune checkpoint blockade-linked upregulation of PD-L1 on myeloid cells, while favoring the expansion of CXCR3+ T cells. KD induced compositional changes of the gut microbiota, with distinct species such as Eisenbergiella massiliensis commonly emerging in mice and humans subjected to carbohydrate-low diet interventions and highly correlating with serum concentrations of 3HB. Altogether, these results demonstrate that KD induces a 3HB-mediated antineoplastic effect that relies on T cell-mediated cancer immunosurveillance.

Gait disorders and postural instability, which are commonly observed in elderly patients with Parkinson disease (PD), respond poorly to dopaminergic agents used to treat other parkinsonian symptoms. The brain structures underlying gait disorders and falls in PD and aging remain to be characterized. Using functional MRI in healthy human subjects, we have shown here that activity of the mesencephalic locomotor region (MLR), which is composed of the pedunculopontine nucleus (PPN) and the adjacent cuneiform nucleus, was modulated by the speed of imagined gait, with faster imagined gait activating a discrete cluster within the MLR. Furthermore, the presence of gait disorders in patients with PD and in aged monkeys rendered parkinsonian by MPTP intoxication correlated with loss of PPN cholinergic neurons. Bilateral lesioning of the cholinergic part of the PPN induced gait and postural deficits in nondopaminergic lesioned monkeys. Our data therefore reveal that the cholinergic neurons of the PPN play a central role in controlling gait and posture and represent a possible target for pharmacological treatment of gait disorders in PD.

RATIONALE: Pulmonary fibrosis is a devastating as yet untreatable disease. We previously investigated the endogenous mediators released on lung injury and showed that uric acid is a danger signal activating Nod-like receptor family, pyrin domain containing 3 (NLRP3) inflammasome in lung inflammation and fibrosis (Gasse et al., Am J Respir Crit Care Med 2009;179:903-913). OBJECTIVES: Here we address the role of extracellular adenosine triphosphate (eATP) in pulmonary inflammation and fibrosis. METHODS: ATP was quantified in bronchoalveolar lavage fluid (BALF) of control subjects and patients with idiopathic pulmonary fibrosis. The contribution of eATP as a danger signal was assessed in a murine model of lung fibrosis induced by airway-administered bleomycin (BLM), an intercalating agent that causes DNA strand breaks. MEASUREMENTS AND MAIN RESULTS: Fibrotic patients have elevated ATP content in BALF in comparison with control individuals. In mice, we report an early increase in eATP levels in BALF on BLM administration. Modulation of eATP levels with the ATP-degrading enzyme apyrase greatly reduced BLM-induced inflammatory cell recruitment, lung IL-1β, and tissue inhibitor of metalloproteinase (TIMP)-1 production, while administration of ATP-γS, a stable ATP derivative, enhanced inflammation. P2X(7) receptor-deficient mice presented dramatically reduced lung inflammation, with reduced fibrosis markers such as lung collagen content and matrix-remodeling proteins TIMP-1 and matrix metalloproteinase-9. The acute inflammation depends on a functional pannexin-1 hemichannel protein. In vitro, ATP is released by pulmonary epithelial cells on BLM-induced stress and this is partly dependent on the presence of functional P2X(7) receptor and pannexin-1 hemichannel. CONCLUSIONS: ATP released from BLM-injured lung cells constitutes a major endogenous danger signal that engages the P2X(7) receptor/pannexin-1 axis, leading to IL-1β maturation and lung fibrosis.

Acute cigarette smoke exposure of the airways (two cigarettes twice daily for three days) induces acute inflammation in mice. In this study, we show that airway inflammation is dependent on Toll-like receptor 4 and IL-1R1 signaling. Cigarette smoke induced a significant recruitment of neutrophils in the bronchoalveolar space and pulmonary parenchyma, which was reduced in TLR4-, MyD88-, and IL-1R1-deficient mice. Diminished neutrophil influx was associated with reduced IL-1, IL-6, and keratinocyte-derived chemokine levels and matrix metalloproteinase-9 activity in the bronchoalveolar space. Further, cigarette smoke condensate (CSC) induced a macrophage proinflammatory response in vitro, which was dependent on MyD88, IL-1R1, and TLR4 signaling, but not attributable to LPS. Heat shock protein 70, a known TLR4 agonist, was induced in the airways upon smoke exposure, which probably activates the innate immune system via TLR4/MyD88, resulting in airway inflammation. CSC-activated macrophages released mature IL-1beta only in presence of ATP, whereas CSC alone promoted the TLR4/MyD88 signaling dependent production of IL-1alpha and pro-IL-1beta implicating cooperation between TLRs and the inflammasome. In conclusion, acute cigarette exposure results in LPS-independent TLR4 activation, leading to IL-1 production and IL-1R1 signaling, which is crucial for cigarette smoke induced inflammation leading to chronic obstructive pulmonary disease with emphysema.

By triggering immunogenic cell death, some anticancer compounds, including anthracyclines and oxaliplatin, elicit tumor-specific, interferon-γ-producing CD8(+) αβ T lymphocytes (Tc1 CTLs) that are pivotal for an optimal therapeutic outcome. Here, we demonstrate that chemotherapy induces a rapid and prominent invasion of interleukin (IL)-17-producing γδ (Vγ4(+) and Vγ6(+)) T lymphocytes (γδ T17 cells) that precedes the accumulation of Tc1 CTLs within the tumor bed. In T cell receptor δ(-/-) or Vγ4/6(-/-) mice, the therapeutic efficacy of chemotherapy was compromised, no IL-17 was produced by tumor-infiltrating T cells, and Tc1 CTLs failed to invade the tumor after treatment. Although γδ T17 cells could produce both IL-17A and IL-22, the absence of a functional IL-17A-IL-17R pathway significantly reduced tumor-specific T cell responses elicited by tumor cell death, and the efficacy of chemotherapy in four independent transplantable tumor models. Adoptive transfer of γδ T cells restored the efficacy of chemotherapy in IL-17A(-/-) hosts. The anticancer effect of infused γδ T cells was lost when they lacked either IL-1R1 or IL-17A. Conventional helper CD4(+) αβ T cells failed to produce IL-17 after chemotherapy. We conclude that γδ T17 cells play a decisive role in chemotherapy-induced anticancer immune responses.

MyD88, the common adapter involved in TLR, IL-1, and IL-18 receptor signaling, is essential for the control of acute Mycobacterium tuberculosis (MTB) infection. Although TLR2, TLR4, and TLR9 have been implicated in the response to mycobacteria, gene disruption for these TLRs impairs only the long-term control of MTB infection. Here, we addressed the respective role of IL-1 and IL-18 receptor pathways in the MyD88-dependent control of acute MTB infection. Mice deficient for IL-1R1, IL-18R, or Toll-IL-1R domain-containing adaptor protein (TIRAP) were compared with MyD88-deficient mice in an acute model of aerogenic MTB infection. Although primary MyD88-deficient macrophages and dendritic cells were defective in cytokine production in response to mycobacterial stimulation, IL-1R1-deficient macrophages exhibited only a reduced IL-12p40 secretion with unaffected TNF, IL-6, and NO production and up-regulation of costimulatory molecules CD40 and CD86. Aerogenic MTB infection of IL-1R1-deficient mice was lethal within 4 wk with 2-log higher bacterial load in the lung and necrotic pneumonia but efficient pulmonary CD4 and CD8 T cell responses, as seen in MyD88-deficient mice. Mice deficient for IL-18R or TIRAP controlled acute MTB infection. These data demonstrate that absence of IL-1R signal leads to a dramatic defect of early control of MTB infection similar to that seen in the absence of MyD88, whereas IL-18R and TIRAP are dispensable, and that IL-1, together with IL-1-induced innate response, might account for most of MyD88-dependent host response to control acute MTB infection.

Liver fibrosis is a severe, life-threatening clinical condition resulting from nonresolving hepatitis of different origins. IL-17A is critical in inflammation, but its relation to liver fibrosis remains elusive. We find increased IL-17A expression in fibrotic livers from HBV-infected patients undergoing partial hepatectomy because of cirrhosis-related early-stage hepatocellular carcinoma in comparison with control nonfibrotic livers from uninfected patients with hepatic hemangioma. In fibrotic livers, IL-17A immunoreactivity localizes to the inflammatory infiltrate. In experimental carbon tetrachloride-induced liver fibrosis of IL-17RA-deficient mice, we observe reduced neutrophil influx, proinflammatory cytokines, hepatocellular necrosis, inflammation, and fibrosis as compared with control C57BL/6 mice. IL-17A is produced by neutrophils and T lymphocytes expressing the Th17 lineage-specific transcription factor Retinoic acid receptor-related orphan receptor γt. Furthermore, hepatic stellate cells (HSCs) isolated from naive C57BL/6 mice respond to IL-17A with increased IL-6, α-smooth muscle actin, collagen, and TGF-β mRNA expression, suggesting an IL-17A-driven fibrotic process. Pharmacologic ERK1/2 or p38 inhibition significantly attenuated IL-17A-induced HSC activation and collagen expression. In conclusion, IL-17A(+) Retinoic acid receptor-related orphan receptor γt(+) neutrophils and T cells are recruited into the injured liver driving a chronic, fibrotic hepatitis. IL-17A-dependent HSC activation may be critical for liver fibrosis. Thus, blockade of IL-17A could potentially benefit patients with chronic hepatitis and liver fibrosis.

The NLRP3 inflammasome is a protein complex involved in IL-1β and IL-18 processing that senses pathogen- and danger-associated molecular patterns (PAMPs and DAMPs). One step- or two step-models have been proposed to explain the tight regulation of IL-1β production during inflammation. Moreover, cellular stimulation triggers adenosine triphosphate (ATP) release and subsequent activation of purinergic receptors at the cell surface. Importantly some studies have reported roles for extracellular ATP, in NLRP3 inflammasome activation in response to PAMPs and DAMPs. In this mini review, we will discuss the link between active ATP release, purinergic signaling and NLRP3 inflammasome activation. We will focus on the role of autocrine or paracrine ATP export in particle-induced NLRP3 inflammasome activation and discuss how particle activators are competent to induce maturation and secretion of IL-1β through a process that involves, as a first event, extracellular release of endogenous ATP through hemichannel opening, and as a second event, signaling through purinergic receptors that trigger NLRP3 inflammasome activation. Finally, we will review the evidence for ATP as a key pro-inflammatory mediator released by dying cells. In particular we will discuss how cancer cells dying via autophagy trigger ATP-dependent NLRP3 inflammasome activation in the macrophages engulfing them, eliciting an immunogenic response against tumors.

Toll-like receptors (TLRs) such as TLR2 and TLR4 have been implicated in host response to mycobacterial infection. Here, mice deficient in the TLR adaptor molecule myeloid differentiation factor 88 (MyD88) were infected with Mycobacterium tuberculosis (MTB). While primary MyD88(-/-) macrophages and DCs are defective in TNF, IL-12, and NO production in response to mycobacterial stimulation, the upregulation of costimulatory molecules CD40 and CD86 is unaffected. Aerogenic infection of MyD88(-/-) mice with MTB is lethal within 4 weeks with 2 log(10) higher CFU in the lung; high pulmonary levels of cytokines and chemokines; and acute, necrotic pneumonia, despite a normal T cell response with IFN-gamma production to mycobacterial antigens upon ex vivo restimulation. Vaccination with Mycobacterium bovis bacillus Calmette-Guerin conferred a substantial protection in MyD88(-/-) mice from acute MTB infection. These data demonstrate that MyD88 signaling is dispensable to raise an acquired immune response to MTB. Nonetheless, this acquired immune response is not sufficient to compensate for the profound innate immune defect and the inability of MyD88(-/-) mice to control MTB infection.

Silica particles induce lung inflammation and fibrosis. Here we show that stimulator of interferon genes (STING) is essential for silica-induced lung inflammation. In mice, silica induces lung cell death and self-dsDNA release in the bronchoalveolar space that activates STING pathway. Degradation of extracellular self-dsDNA by DNase I inhibits silica-induced STING activation and the downstream type I IFN response. Patients with silicosis have increased circulating dsDNA and CXCL10 in sputum, and patients with fibrotic interstitial lung disease display STING activation and CXCL10 in the lung. In vitro, while mitochondrial dsDNA is sensed by cGAS-STING in dendritic cells, in macrophages extracellular dsDNA activates STING independent of cGAS after silica exposure. These results reveal an essential function of STING-mediated self-dsDNA sensing after silica exposure, and identify DNase I as a potential therapy for silica-induced lung inflammation.

Oxidative stress plays an important role in the pathogenesis of chronic obstructive pulmonary disease (COPD) caused by cigarette smoke and characterized by chronic inflammation, alveolar destruction (emphysema) and bronchiolar obstruction. Ozone is a gaseous constituent of urban air pollution resulting from photochemical interaction of air pollutants such as nitrogen oxide and organic compounds. While acute exposure to ozone induces airway hyperreactivity and neutrophilic inflammation, chronic ozone exposure in mice causes activation of oxidative pathways resulting in cell death and a chronic bronchial inflammation with emphysema, mimicking cigarette smoke-induced COPD. Therefore, the chronic exposure to ozone has become a model for studying COPD. We review recent data on mechanisms of ozone induced lung disease focusing on pathways causing chronic respiratory epithelial cell injury, cell death, alveolar destruction, and tissue remodeling associated with the development of chronic inflammation and AHR. The initial oxidant insult may result from direct effects on the integrity of membranes and organelles of exposed epithelial cells in the airways causing a stress response with the release of mitochondrial reactive oxygen species (ROS), DNA, and proteases. Mitochondrial ROS and mitochondrial DNA activate NLRP3 inflammasome and the DNA sensors cGAS and STING accelerating cell death pathways including caspases with inflammation enhancing alveolar septa destruction, remodeling, and fibrosis. Inhibitors of mitochondrial ROS, NLRP3 inflammasome, DNA sensor, cell death pathways, and IL-1 represent novel therapeutic targets for chronic airways diseases underlined by oxidative stress.

The international ImMunoGeneTics information system (IMGT) (http://imgt.cines.fr), created in 1989, by the Laboratoire d'ImmunoGenetique Moleculaire LIGM (Universite Montpellier II and CNRS) at Montpellier, France, is a high-quality integrated knowledge resource specializing in the immunoglobulins (IGs), T cell receptors (TRs), major histocompatibility complex (MHC) of human and other vertebrates, and related proteins of the immune systems (RPI) that belong to the immunoglobulin superfamily (IgSF) and to the MHC superfamily (MhcSF). IMGT includes several sequence databases (IMGT/LIGM-DB, IMGT/PRIMER-DB, IMGT/PROTEIN-DB and IMGT/MHC-DB), one genome database (IMGT/GENE-DB) and one three-dimensional (3D) structure database (IMGT/3Dstructure-DB), Web resources comprising 8000 HTML pages (IMGT Marie-Paule page), and interactive tools. IMGT data are expertly annotated according to the rules of the IMGT Scientific chart, based on the IMGT-ONTOLOGY concepts. IMGT tools are particularly useful for the analysis of the IG and TR repertoires in normal physiological and pathological situations. IMGT is used in medical research (autoimmune diseases, infectious diseases, AIDS, leukemias, lymphomas, myelomas), veterinary research, biotechnology related to antibody engineering (phage displays, combinatorial libraries, chimeric, humanized and human antibodies), diagnostics (clonalities, detection and follow up of residual diseases) and therapeutical approaches (graft, immunotherapy and vaccinology). IMGT is freely available at http://imgt.cines.fr.

Toll-like receptors (TLRs) such as TLR2 and TLR4 have been implicated in host response to mycobacterial infection. Here, mice deficient in the TLR adaptor molecule myeloid differentiation factor 88 (MyD88) were infected with Mycobacterium tuberculosis (MTB). While primary MyD88–/– macrophages and DCs are defective in TNF, IL-12, and NO production in response to mycobacterial stimulation, the upregulation of costimulatory molecules CD40 and CD86 is unaffected. Aerogenic infection of MyD88–/– mice with MTB is lethal within 4 weeks with 2 log10 higher CFU in the lung; high pulmonary levels of cytokines and chemokines; and acute, necrotic pneumonia, despite a normal T cell response with IFN-γ production to mycobacterial antigens upon ex vivo restimulation. Vaccination with Mycobacterium bovis bacillus Calmette-Guérin conferred a substantial protection in MyD88–/– mice from acute MTB infection. These data demonstrate that MyD88 signaling is dispensable to raise an acquired immune response to MTB. Nonetheless, this acquired immune response is not sufficient to compensate for the profound innate immune defect and the inability of MyD88–/– mice to control MTB infection.

Deposition of uric acid crystals in joints causes the acute and chronic inflammatory disease known as gout and prolonged airway exposure to silica crystals leads to the development of silicosis, an irreversible fibrotic pulmonary disease. Aluminum salt (Alum) crystals are frequently used as vaccine adjuvant. The mechanisms by which crystals activate innate immunity through the Nlrp3 inflammasome are not well understood. Here, we show that uric acid, silica and Alum crystals trigger the extracellular delivery of endogenous ATP, which just precedes the secretion of mature interleukin-1β (IL-1β) by macrophages, both events depending on purinergic receptors and connexin/pannexin channels. Interestingly, not only ATP but also ADP and UTP are involved in IL-1β production upon these Nlrp3 inflammasome activators through multiple purinergic receptor signaling. These findings support a pivotal role for nucleotides as danger signals and provide a new molecular mechanism to explain how chemically and structurally diverse stimuli can activate the Nlrp3 inflammasome.

OBJECTIVE: To investigate the longitudinal dose-effect relationship between dopamine replacement therapy and impulse control disorders (ICDs) in Parkinson disease (PD). METHODS: We used data from a multicenter longitudinal cohort of consecutive patients with PD with ≤5 years' disease duration at baseline followed up annually up to 5 years. ICDs were evaluated during face-to-face semistructured interviews with movement disorder specialists. Generalized estimating equations and Poisson models with robust variance were used to study the association between several time-dependent definitions of dopamine agonist (DA) use, taking dose and duration of treatment into account, and ICDs at each visit. Other antiparkinsonian drugs were also examined. RESULTS: Among 411 patients (40.6% women, mean age 62.3 years, average follow-up 3.3 years, SD 1.7 years), 356 (86.6%) took a DA at least once since disease onset. In 306 patients without ICDs at baseline, the 5-year cumulative incidence of ICDs was 46.1% (95% confidence interval [CI] 37.4-55.7, DA ever users 51.5% [95% CI 41.8-62.1], DA never users 12.4% [95% CI 4.8-30.0]). ICD prevalence increased from 19.7% at baseline to 32.8% after 5 years. ICDs were associated with ever DA use (prevalence ratio 4.23, 95% CI 1.78-10.09). Lifetime average daily dose and duration of treatment were independently associated with ICDs with significant dose-effect relationships. Similar analyses for levodopa were not in favor of a strong association. ICDs progressively resolved after DA discontinuation. CONCLUSION: In this longitudinal study of patients with PD characterized by a high prevalence of DA treatment, the 5-year cumulative incidence of ICDs was ≈46%. ICDs were strongly associated with DA use with a dose-effect relationship; both increasing duration and dose were associated with ICDs. ICDs progressively resolved after DA discontinuation. CLINICALTRIALSGOV IDENTIFIER: NCT01564992.

Stimulator of interferon genes (STING) contributes to immune responses against tumors and may control viral infection including SARS-CoV-2 infection. However, activation of the STING pathway by airway silica or smoke exposure leads to cell death, self-dsDNA release, and STING/type I IFN dependent acute lung inflammation/ARDS. The inflammatory response induced by a synthetic non-nucleotide-based diABZI STING agonist, in comparison to the natural cyclic dinucleotide cGAMP, is unknown. A low dose of diABZI (1 µg by endotracheal route for 3 consecutive days) triggered an acute neutrophilic inflammation, disruption of the respiratory barrier, DNA release with NET formation, PANoptosis cell death, and inflammatory cytokines with type I IFN dependent acute lung inflammation. Downstream upregulation of DNA sensors including cGAS, DDX41, IFI204, as well as NLRP3 and AIM2 inflammasomes, suggested a secondary inflammatory response to dsDNA as a danger signal. DNase I treatment, inhibition of NET formation together with an investigation in gene-deficient mice highlighted extracellular DNA and TLR9, but not cGAS, as central to diABZI-induced neutrophilic response. Therefore, activation of acute cell death with DNA release may lead to ARDS which may be modeled by diABZI. These results show that airway targeting by STING activator as a therapeutic strategy for infection may enhance lung inflammation with severe ARDS. STING agonist diABZI induces neutrophilic lung inflammation and PANoptosis A, Airway STING priming induce a neutrophilic lung inflammation with epithelial barrier damage, double-stranded DNA release in the bronchoalvelolar space, cell death, NETosis and type I interferon release. B, 1. The diamidobenzimidazole (diABZI), a STING agonist is internalized into the cytoplasm through unknown receptor and induce the activation and dimerization of STING followed by TBK1/IRF3 phosporylation leading to type I IFN response. STING activation also leads to NF-kB activation and the production of pro-inflammatory cytokines TNFα and IL-6. 2. The activation of TNFR1 and IFNAR1 signaling pathway results in ZBP1 and RIPK3/ASC/CASP8 activation leading to MLKL phosphorylation and necroptosis induction. 3. This can also leads to Caspase-3 cleavage and apoptosis induction. 4. Self-dsDNA or mtDNA sensing by NLRP3 or AIM2 induces inflammsome formation leading to Gasdermin D cleavage enabling Gasdermin D pore formation and the release mature IL-1β and pyroptosis. NLRP3 inflammasome formation can be enhanced by the ZBP1/RIPK3/CASP8 complex. 5. A second signal of STING activation with diABZI induces cell death and the release of self-DNA which is sensed by cGAS and form 2'3'-cGAMP leading to STING hyper activation, the amplification of TBK1/IRF3 and NF-kB pathway and the subsequent production of IFN-I and inflammatory TNFα and IL-6. This also leads to IFI204 and DDX41 upregulation thus, amplifying the inflammatory loop. The upregulation of apoptosis, pyroptosis and necroptosis is indicative of STING-dependent PANoptosis.