Inherited human RelB deficiency impairs innate and adaptive immunity to infection.

We report two unrelated adults with homozygous (P1) or compound heterozygous (P2) private loss-of-function variants of V-Rel Reticuloendotheliosis Viral Oncogene Homolog B (RELB). The resulting deficiency of functional RelB impairs the induction of NFKB2 mRNA and NF-κB2 (p100/p52) protein by lymphotoxin in the fibroblasts of the patients. These defects are rescued by transduction with wild-type RELB complementary DNA (cDNA). By contrast, the response of RelB-deficient fibroblasts to Tumor Necrosis Factor (TNF) or IL-1ß via the canonical NF-κB pathway remains intact. P1 and P2 have low proportions of naïve CD4+ and CD8+ T cells and of memory B cells. Moreover, their naïve B cells cannot differentiate into immunoglobulin G (IgG)- or immunoglobulin A (IgA)-secreting cells in response to CD40L/IL-21, and the development of IL-17A/F-producing T cells is strongly impaired in vitro. Finally, the patients produce neutralizing autoantibodies against type I interferons (IFNs), even after hematopoietic stem cell transplantation, attesting to a persistent dysfunction of thymic epithelial cells in T cell selection and central tolerance to some autoantigens. Thus, inherited human RelB deficiency disrupts the alternative NF-κB pathway, underlying a T- and B cell immunodeficiency, which, together with neutralizing autoantibodies against type I IFNs, confers a predisposition to viral, bacterial, and fungal infections.

Overexpression of ESYT3 improves radioimmune responses through activating cGAS-STING pathway in lung adenocarcinoma.

BACKGROUND: Radiotherapy can modulate systemic antitumor immunity, while immune status in the tumor microenvironment also influences the efficacy of radiotherapy, but relevant molecular mechanisms are poorly understood in lung adenocarcinoma (LUAD). METHODS: In this study, we innovatively proposed a radiotherapy response classification for LUAD, and discovered ESYT3 served as a tumor suppressor and radioimmune response sensitizer. ESYT3 expression was measured both in radioresistant and radiosensitive LUAD tissues and cells. The influence of ESYT3 on radiotherapy sensitivity and resistance was then investigated. Interaction between ESYT3 and STING was evaluated through multiple immunofluorescent staining and coimmunoprecipitation, and downstream molecules were further analyzed. In vivo models were constructed to assess the combination treatment efficacy of ESYT3 overexpression with radiotherapy. RESULTS: We found that radioresistant subtype presented immunosuppressive state and activation of DNA damage repair pathways than radiosensitive subtype. ESYT3 expression was remarkably attenuated both in radioresistant LUAD tissues and cells. Clinically, low ESYT3 expression was linked with radioresistance. Overexpression of ESYT3 enabled to alleviate radioresistance, and sensitize LUAD cells to DNA damage induced by irradiation. Mechanically, ESYT3 directly interacted with STING, and activated cGAS-STING signaling, subsequently increasing the generation of type I IFNs as well as downstream chemokines CCL5 and CXCL10, thus improving radioimmune responses. The combination treatment of ESYT3 overexpression with radiotherapy had a synergistic anticancer effect in vitro and in vivo. CONCLUSIONS: In summary, low ESYT3 expression confers resistance to radiotherapy in LUAD, and its overexpression can improve radioimmune responses through activating cGAS-STING-dependent pathway, thus providing an alternative combination therapeutic strategy for LUAD patients.

Different activation of STAT1 and STAT2 phosphorylation by IFNc, IFNd, and IFNh in tilapia.

Type I IFNs are a subset of cytokines exerting their antiviral effects mainly through the JAK-STAT signalling. Immunogenetic studies have shown that fish possess key components of IFN-JAK-STAT cascade, but the information about the distinct responses of STAT1 and STAT2 to different IFNs is rather limited in fish. Here, we identified and cloned STAT1 and STAT2 genes (named as On-STAT1 and On-STAT2) from tilapia, Oreochromis niloticus. On-STAT1 and On-STAT2 genes were detected in all orangs/tissues examined, and were rapidly induced in spleen, head kidney, and liver following the stimulation of poly(I:C). In addition, the stimulation of poly(I:C), poly(A:T), and different subgroups of recombinant IFNs could induce the expression of On-STAT1 and On-STAT2 in TA-02 cells with distinct induction levels. Importantly, On-STAT2 was rapidly phosphorylated by all three subgroups of IFNs, but the phosphorylation of On-STAT1 was only observed in IFNc- and IFNh-treated TA-02 cells, reflecting the distinct activation of STAT by different subgroups of fish IFNs. The present results thus contribute to better understanding of the JAK-STAT signalling mediated by different subgroups of IFNs in fish.

Regulation of Type I Interferon and Autophagy in Immunity against Mycobacterium Tuberculosis: Role of CGAS and STING1.

Mycobacterium tuberculosis (M. tb) is a significant intracellular pathogen responsible for numerous infectious disease-related deaths worldwide. It uses ESX-1 T7SS to damage phagosomes and to enter the cytosol of host cells after phagocytosis. During infection, M. tb and host mitochondria release dsDNA, which activates the CGAS-STING1 pathway. This pathway leads to the production of type I interferons and proinflammatory cytokines and activates autophagy, which targets and degrades bacteria within autophagosomes. However, the role of type I IFNs in immunity against M. tb is controversial. While previous research has suggested a protective role, recent findings from cgas-sting1 knockout mouse studies have contradicted this. Additionally, a study using knockout mice and non-human primate models uncovered a new mechanism by which neutrophils recruited to lung infections form neutrophil extracellular traps. Activating plasmacytoid dendritic cells causes them to produce type I IFNs, which interfere with the function of interstitial macrophages and increase the likelihood of tuberculosis. Notably, M. tb uses its virulence proteins to disrupt the CGAS-STING1 signaling pathway leading to enhanced pathogenesis. Investigating the CGAS-STING1 pathway can help develop new ways to fight tuberculosis.

The Interplay between KSHV Infection and DNA-Sensing Pathways.

During viral infection, the innate immune system utilizes a variety of specific intracellular sensors to detect virus-derived nucleic acids and activate a series of cellular signaling cascades that produce type I IFNs and proinflammatory cytokines and chemokines. Kaposi's sarcoma-associated herpesvirus (KSHV) is an oncogenic double-stranded DNA virus that has been associated with a variety of human malignancies, including Kaposi's sarcoma, primary effusion lymphoma, and multicentric Castleman disease. Infection with KSHV activates various DNA sensors, including cGAS, STING, IFI16, and DExD/H-box helicases. Activation of these DNA sensors induces the innate immune response to antagonize the virus. To counteract this, KSHV has developed countless strategies to evade or inhibit DNA sensing and facilitate its own infection. This review summarizes the major DNA-triggered sensing signaling pathways and details the current knowledge of DNA-sensing mechanisms involved in KSHV infection, as well as how KSHV evades antiviral signaling pathways to successfully establish latent infection and undergo lytic reactivation.

Suppression of Type I Interferon Signaling in Myeloid Cells by Autoantibodies in Severe COVID-19 Patients.

PURPOSE: Auto-antibodies (auto-abs) to type I interferons (IFNs) have been identified in patients with life-threatening coronavirus disease 2019 (COVID-19), suggesting that the presence of auto-abs may be a risk factor for disease severity. We therefore investigated the mechanism underlying COVID-19 exacerbation induced by auto-abs to type I IFNs. METHODS: We evaluated plasma from 123 patients with COVID-19 to measure auto-abs to type I IFNs. We performed single-cell RNA sequencing (scRNA-seq) of peripheral blood mononuclear cells from the patients with auto-abs and conducted epitope mapping of the auto-abs. RESULTS: Three of 19 severe and 4 of 42 critical COVID-19 patients had neutralizing auto-abs to type I IFNs. Patients with auto-abs to type I IFNs showed no characteristic clinical features. scRNA-seq from 38 patients with COVID-19 revealed that IFN signaling in conventional dendritic cells and canonical monocytes was attenuated, and SARS-CoV-2-specific BCR repertoires were decreased in patients with auto-abs. Furthermore, auto-abs to IFN-α2 from COVID-19 patients with auto-abs recognized characteristic epitopes of IFN-α2, which binds to the receptor. CONCLUSION: Auto-abs to type I IFN found in COVID-19 patients inhibited IFN signaling in dendritic cells and monocytes by blocking the binding of type I IFN to its receptor. The failure to properly induce production of an antibody to SARS-CoV-2 may be a causative factor of COVID-19 severity.

IFNα Subtypes in HIV Infection and Immunity.

Type I interferons (IFN), immediately triggered following most viral infections, play a pivotal role in direct antiviral immunity and act as a bridge between innate and adaptive immune responses. However, numerous viruses have evolved evasion strategies against IFN responses, prompting the exploration of therapeutic alternatives for viral infections. Within the type I IFN family, 12 IFNα subtypes exist, all binding to the same receptor but displaying significant variations in their biological activities. Currently, clinical treatments for chronic virus infections predominantly rely on a single IFNα subtype (IFNα2a/b). However, the efficacy of this therapeutic treatment is relatively limited, particularly in the context of Human Immunodeficiency Virus (HIV) infection. Recent investigations have delved into alternative IFNα subtypes, identifying certain subtypes as highly potent, and their antiviral and immunomodulatory properties have been extensively characterized. This review consolidates recent findings on the roles of individual IFNα subtypes during HIV and Simian Immunodeficiency Virus (SIV) infections. It encompasses their induction in the context of HIV/SIV infection, their antiretroviral activity, and the diverse regulation of the immune response against HIV by distinct IFNα subtypes. These insights may pave the way for innovative strategies in HIV cure or functional cure studies.

Evolutionary and functional conservation of IRF7 in the Tibetan frog Nanorana parkeri.

BACKGROUND: The production of interferons (IFNs) is essential for the control of viral infections, and interferon regulatory factor 7 (IRF7) is considered as a vital regulator for the transcription of type I IFNs. Amphibians appear to possess a highly expanded type I IFN repertoire, consisting of intron-containing genes as observed in fish, and intronless genes as in other higher vertebrates. However, the knowledge on transcriptional regulatory mechanism of these two types of type I IFN genes is rather scarce in amphibians. METHODS AND RESULTS: A IRF7 gene named as Np-IRF7 was identified in Tibetan frog (Nanorana parkeri), and bioinformatic analysis revealed that the predicted protein of Np-IRF7 contains several important structural features known in IRF7. Expression analysis showed that Np-IRF7 gene was widely expressed and rapidly induced by poly(I:C) in different organs/tissues. Interestingly, luciferase reporter assay revealed that intronless IFN promoters were more effectively activated than intron-containing IFN promoter in Np-IRF7-transfected cells. Moreover, the overexpression of Np-IRF7 could induce the expression of ISGs and suppress the replication of FV3 in A6 cells. CONCLUSION: Np-IRF7 is indeed the ortholog of known IRF7, and IRF7 is structurally conserved in different lineages of vertebrates. Np-IRF7 played distinct roles in the activation of intron-containing and intronless type I IFN promoters, thus inducing the expression of interferon-stimulated antiviral effectors and providing a protection against ranavirus infection. The present research thus contributes to a better understanding of regulatory function of IRF7 in the IFN-mediated antiviral response of anuran amphibians.

GAS-STING: a classical DNA recognition pathways to tumor therapy.

Cyclic GMP-AMP synthetase (cGAS), recognized as the primary DNA sensor within cells, possesses the capability to identify foreign DNA molecules along with free DNA fragments. This identification process facilitates the production of type I IFNs through the activator of the interferon gene (STING) which induces the phosphorylation of downstream transcription factors. This action characterizes the most archetypal biological functionality of the cGAS-STING pathway. When treated with anti-tumor agents, cells experience DNA damage that triggers activation of the cGAS-STING pathway, culminating in the expression of type I IFNs and associated downstream interferon-stimulated genes. cGAS-STING is one of the important innate immune pathways,the role of type I IFNs in the articulation between innate immunity and T-cell antitumour immunity.type I IFNs promote the recruitment and activation of inflammatory cells (including NK cells) at the tumor site.Type I IFNs also can promote the activation and maturation of dendritic cel(DC), improve the antigen presentation of CD4+T lymphocytes, and enhance the cross-presentation of CD8+T lymphocytes to upregulating anti-tumor responses. This review discussed the cGAS-STING signaling and its mechanism and biological function in traditional tumor therapy and immunotherapy.

Polyamine metabolism controls B-to-Z DNA transition to orchestrate DNA sensor cGAS activity.

Cyclic guanosine monophosphate (GMP)-AMP (cGAMP) synthase (cGAS) is a universal double-stranded DNA (dsDNA) sensor that recognizes foreign and self-DNA in the cytoplasm and initiates innate immune responses and has been implicated in various infectious and non-infectious contexts. cGAS binds to the backbone of dsDNA and generates the second messenger, cGAMP, which activates the stimulator of interferon genes (STING). Here, we show that the endogenous polyamines spermine and spermidine attenuated cGAS activity and innate immune responses. Mechanistically, spermine and spermidine induced the transition of B-form DNA to Z-form DNA (Z-DNA), thereby decreasing its binding affinity with cGAS. Spermidine/spermine N1-acetyltransferase 1 (SAT1), the rate-limiting enzyme in polyamine catabolism that decreases the cellular concentrations of spermine and spermidine, enhanced cGAS activation by inhibiting cellular Z-DNA accumulation; SAT1 deficiency promoted herpes simplex virus 1 (HSV-1) replication in vivo. The results indicate that spermine and spermidine induce dsDNA to adopt the Z-form conformation and that SAT1-mediated polyamine metabolism orchestrates cGAS activity.

Regulation of type I IFN responses by deubiquitinating enzyme A in inflammatory bowel diseases.

The development of Inflammatory bowel disease (IBD) is driven by excessive production of pro-inflammatory cytokines including TNF-α, IL-12, and IL-23. This notion is supported by the remarkable clinical success of biologics targeting these cytokines. Recognition of cell wall components derived from intestinal bacteria by Toll-like receptors (TLRs) induces the production of these pro-inflammatory cytokines by macrophages and dendritic cells in human IBD and experimental colitis model. Although sensing of bacterial nucleic acids by endosomal TLRs, specifically TLR3, TLR7, and TLR9 leads to robust production of type I IFNs, it remains debatable whether TLR-mediated type I IFN responses are pathogenic or protective in IBD patients. Additionally, recent studies identified deubiquitinating enzyme A (DUBA) as a novel negative regulator of TLR-mediated type I IFN responses. In light of these observations and their potential applications, in this review, we summarize recent findings on the roles of type I IFN responses and DUBA-mediated negative regulation of these responses in human IBD and experimental colitis model.

Heterogeneity and functions of the 13 IFN-α subtypes - lucky for some?

Type I IFNs are critical for host responses to viral infection and are also implicated in the pathogenesis of multiple autoimmune diseases. Multiple subtypes exist within the type I IFN family, in particular 13 distinct IFN-α genes, which signal through the same heterodimer receptor that is ubiquitously expressed by mammalian cells. Both evolutionary genetic studies and functional antiviral assays strongly suggest differential functions and activity between the 13 IFN-α subtypes, yet we still lack a clear understanding of these different roles. This review summarizes the evidence from studies describing differential functions of IFN-α subtypes and highlights potential reasons for discrepancies between the reports. We examine both acute and chronic viral infection, as well as autoimmunity, and integrate a more recent awareness of the importance of anti-IFN-α autoantibodies in shaping the type I IFN responses in these different conditions.

Transcriptomic Analysis of Mycobacterial Infected Macrophages Reveals a High MOI Specific Type I IFN Signaling.

Macrophage (MΦ) infection models are important tools for studying host-mycobacterial interactions. Although the multiplicity of infection (MOI) is an important experimental variable, the selection of MOI in mycobacterial infection experiments is largely empirical, without reference to solid experimental data. To provide relevant data, we used RNA-seq to analyze the gene expression profiles of MΦs 4 or 24 h after infection with Mycobacterium marinum (M. m) at MOIs ranging from 0.1 to 50. Analysis of differentially expressed genes (DEGs) showed that different MOIs are linked to distinct transcriptomic changes and only 10% of DEGs were shared by MΦ infected at all MOIs. KEGG pathway enrichment analysis revealed that type I interferon (IFN)-related pathways were inoculant dose-dependent and enriched only at high MOIs, whereas TNF pathways were inoculant dose-independent and enriched at all MOIs. Protein-protein interaction (PPI) network alignment showed that different MOIs had distinct key node genes. By fluorescence-activated cell sorting and follow-up RT-PCR analysis, we could separate infected MΦs from uninfected MΦs and found phagocytosis of mycobacteria to be the determinant factor for type I IFN production. The distinct transcriptional regulation of RAW264.7 MΦ genes at different MOIs was also seen with Mycobacterium tuberculosis (M.tb) infections and primary MΦ infection models. In summary, transcriptional profiling of mycobacterial infected MΦs revealed that different MOIs activate distinct immune pathways and the type I IFN pathway is activated only at high MOIs. This study should provide guidance for selecting the MOI most appropriate for different research questions.

Type I Interferon Orchestrates Demand-Adapted Monopoiesis during Influenza A Virus Infection via STAT1-Mediated Upregulation of Macrophage Colony-Stimulating Factor Receptor Expression.

Whether and how a local virus infection affects the hematopoietic system in the bone marrow is largely unknown, unlike with systemic infection. In this study, we showed that influenza A virus (IAV) infection leads to demand-adapted monopoiesis in the bone marrow. The beta interferon (IFN-ß) promoter stimulator 1 (IPS-1)-type I IFN-IFN-α receptor 1 (IFNAR1) axis-mediated signaling was found to induce the emergency expansion of the granulocyte-monocyte progenitor (GMP) population and upregulate the expression of the macrophage colony-stimulating factor receptor (M-CSFR) on bipotent GMPs and monocyte progenitors via the signal transducer and activator of transcription 1 (STAT1), leading to a scaled-back proportion of granulocyte progenitors. To further address the influence of demand-adapted monopoiesis on IAV-induced secondary bacterial infection, IAV-infected wild-type (WT) and Stat1-/- mice were challenged with Streptococcus pneumoniae. Compared with WT mice, Stat1-/- mice did not demonstrate demand-adapted monopoiesis, had more infiltrating granulocytes, and were able to effectively eliminate the bacterial infection. IMPORTANCE Our findings show that influenza A virus infection induces type I interferon (IFN)-mediated emergency hematopoiesis to expand the GMP population in the bone marrow. The type I IFN-STAT1 axis was identified as being involved in mediating the viral-infection-driven demand-adapted monopoiesis by upregulating M-CSFR expression in the GMP population. As secondary bacterial infections often manifest during a viral infection and can lead to severe or even fatal clinical complications, we further assessed the impact of the observed monopoiesis on bacterial clearance. Our results suggest that the resulting decrease in the proportion of granulocytes may play a role in diminishing the IAV-infected host's ability to effectively clear secondary bacterial infection. Our findings not only provide a more complete picture of the modulatory functions of type I IFN but also highlight the need for a more comprehensive understanding of potential changes in hematopoiesis during local infections to better inform clinical interventions.

Role of Innate Interferon Responses at the Ocular Surface in Herpes Simplex Virus-1-Induced Herpetic Stromal Keratitis.

Herpes simplex virus type 1 (HSV-1) is a highly successful pathogen that primarily infects epithelial cells of the orofacial mucosa. After initial lytic replication, HSV-1 enters sensory neurons and undergoes lifelong latency in the trigeminal ganglion (TG). Reactivation from latency occurs throughout the host's life and is more common in people with a compromised immune system. HSV-1 causes various diseases depending on the site of lytic HSV-1 replication. These include herpes labialis, herpetic stromal keratitis (HSK), meningitis, and herpes simplex encephalitis (HSE). HSK is an immunopathological condition and is usually the consequence of HSV-1 reactivation, anterograde transport to the corneal surface, lytic replication in the epithelial cells, and activation of the host's innate and adaptive immune responses in the cornea. HSV-1 is recognized by cell surface, endosomal, and cytoplasmic pattern recognition receptors (PRRs) and activates innate immune responses that include interferons (IFNs), chemokine and cytokine production, as well as the recruitment of inflammatory cells to the site of replication. In the cornea, HSV-1 replication promotes type I (IFN-α/ß) and type III (IFN-λ) IFN production. This review summarizes our current understanding of HSV-1 recognition by PRRs and innate IFN-mediated antiviral immunity during HSV-1 infection of the cornea. We also discuss the immunopathogenesis of HSK, current HSK therapeutics and challenges, proposed experimental approaches, and benefits of promoting local IFN-λ responses.

Type I interferon autoantibodies in hospitalized patients with Middle East respiratory syndrome and association with outcomes and treatment effect of interferon beta-1b in MIRACLE clinical trial.

Background: Type I interferons (IFNs) are essential antiviral cytokines induced upon respiratory exposure to coronaviruses. Defects in type I IFN signaling can result in severe disease upon exposure to respiratory viral infection and are associated with worse clinical outcomes. Neutralizing autoantibodies (auto-Abs) to type I IFNs were reported as a risk factor for life-threatening COVID-19, but their presence has not been evaluated in patients with severe Middle East respiratory syndrome (MERS). Methods: We evaluated the prevalence of type I IFN auto-Abs in a cohort of hospitalized patients with MERS who were enrolled in a placebo-controlled clinical trial for treatment with IFN-ß1b and lopinavir-ritonavir (MIRACLE trial). Samples were tested for type I IFN auto-Abs using a multiplex particle-based assay. Results: Among the 62 enrolled patients, 15 (24.2%) were positive for immunoglobulin G auto-Abs for at least one subtype of type I IFNs. Auto-Abs positive patients were not different from auto-Abs negative patients in age, sex, or comorbidities. However, the majority (93.3%) of patients who were auto-Abs positive were critically ill and admitted to the ICU at the time of enrollment compared to 66% in the auto-Abs negative patients. The effect of treatment with IFN-ß1b and lopinavir-ritonavir did not significantly differ between the two groups. Conclusion: This study demonstrates the presence of type I IFN auto-Abs in hospitalized patients with MERS.

A hnRNPA2B1 agonist effectively inhibits HBV and SARS-CoV-2 omicron in vivo.

The twenty-first century has already recorded more than ten major epidemics or pandemics of viral disease, including the devastating COVID-19. Novel effective antivirals with broad-spectrum coverage are urgently needed. Herein, we reported a novel broad-spectrum antiviral compound PAC5. Oral administration of PAC5 eliminated HBV cccDNA and reduced the large antigen load in distinct mouse models of HBV infection. Strikingly, oral administration of PAC5 in a hamster model of SARS-CoV-2 omicron (BA.1) infection significantly decreases viral loads and attenuates lung inflammation. Mechanistically, PAC5 binds to a pocket near Asp49 in the RNA recognition motif of hnRNPA2B1. PAC5-bound hnRNPA2B1 is extensively activated and translocated to the cytoplasm where it initiates the TBK1-IRF3 pathway, leading to the production of type I IFNs with antiviral activity. Our results indicate that PAC5 is a novel small-molecule agonist of hnRNPA2B1, which may have a role in dealing with emerging infectious diseases now and in the future.

How Different Pathologies Are Affected by IFIT Expression.

The type-I interferon (IFN) system represents the first line of defense against viral pathogens. Recognition of the virus initiates complex signaling pathways that result in the transcriptional induction of IFNs, which are then secreted. Secreted IFNs stimulate nearby cells and result in the production of numerous proinflammatory cytokines and antiviral factors. Of particular note, IFN-induced tetratricopeptide repeat (IFIT) proteins have been thoroughly studied because of their antiviral activity against different viral pathogens. Although classically studied as an antiviral protein, IFIT expression has recently been investigated in the context of nonviral pathologies, such as cancer and sepsis. In oral squamous cell carcinoma (OSCC), IFIT1 and IFIT3 promote metastasis, while IFIT2 exhibits the opposite effect. The role of IFIT proteins during bacterial/fungal sepsis is still under investigation, with studies showing conflicting roles for IFIT2 in disease severity. In the setting of viral sepsis, IFIT proteins play a key role in clearing viral infection. As a result, many viral pathogens, such as SARS-CoV-2, employ mechanisms to inhibit the type-I IFN system and promote viral replication. In cancers that are characterized by upregulated IFIT proteins, medications that decrease IFIT expression may reduce metastasis and improve survival rates. Likewise, in cases of viral sepsis, therapeutics that increase IFIT expression may improve viral clearance and reduce the risk of septic shock. By understanding the effect of IFIT proteins in different pathologies, novel therapeutics can be developed to halt disease progression.

Immunogenicity of small-cell lung cancer associates with STING pathway activation and is enhanced by ATR and TOP1 inhibition.

INTRODUCTION: The activation of STING (stimulator of interferon genes) pathway enhances antitumor immunity in small-cell lung cancer (SCLC), while the DNA damage induced by non-cGAMP-based agonists is a potent inducer of STING activity. Here, we investigate the intrinsic expression of STING in cancer cells and evaluate the value of the combination of ATR and TOP1 inhibitors in enhancing antitumor immunity. METHODS: STING expression was assessed at mRNA and protein levels in SCLC and normal lung tissues. Transcriptomic subsets of SCLC were identified based on STING-related genes. Distinct mutation and immunogenomic profiles of these subsets were determined. The direct antitumor efficacy and the potential of enhancing antitumor immunity of the strategy using the ATR-TOP1-inhibitor combination were tested in SCLC cell lines. RESULTS: The intrinsic expression of STING was significantly reduced in SCLC compared to normal lung tissues (p < 0.0001). Three STING-related SCLC subtypes were identified in which the STING-high subtype was associated with (1) high immune infiltration, (2) high expression of genes related to MHC and immune checkpoints, and (3) high EMT and ferroptosis score. On the contrary, the STING-low subtype was enriched with pathways related to DNA damage response (DDR) and cell cycle progression. The association between the DDR pathway activity and the STING-IFN innate immune response was verified by in vitro experiments in which the inhibition of ATR and TOP1 triggered the expression of genes encoding type I IFN signaling and pro-inflammatory cytokines/chemokines in a STING-low SCLC cell line. CONCLUSION: Our study verifies that activation of the STING-IFN response by ATR and TOP1 inhibitors might be a therapeutic strategy to improve the response to immune checkpoint therapy in STING-low SCLC. Furthermore, the combinations of ATR and TOP1 inhibitors can augment tumor inflammation in STING-low SCLC.

Sex-dependent differences in type I IFN-induced natural killer cell activation.

Natural killer (NK) cells are important antiviral effector cells and also involved in tumor clearance. NK cells express IFNAR, rendering them responsive to Type I IFNs. To evaluate Type I IFN-mediated modulation of NK cell functions, individual Type I IFNs subtypes were assessed for their ability to activate NK cells. Different Type I IFN subtypes displayed a broad range in the capacity to induce and modulate NK cell activation and degranulation, measured by CD69 and CD107a expression in response to leukemia cell line K562. When including biological sex as a variable in the analysis, transwell co-cultures of NK cells with either male- or female-derived PBMCs or pDCs stimulated with the TLR7/8 agonist CL097 showed that NK cells were more activated by CL097-stimulated cells derived from females. These sex-specific differences were linked to higher CL097-induced IFNα production by pDCs derived from females, indicating an extrinsic sex-specific effect of Type I IFNs on NK cell function. Interestingly, in addition to the extrinsic effect, we also observed NK cell-intrinsic sex differences, as female NK cells displayed higher activation levels after IFNα-stimulation and after co-culture with CL097-stimulated pDCs, suggesting higher activation of IFNα-signaling transduction in female NK cells. Taken together, the results from these studies identify both extrinsic and intrinsic sex-specific differences in Type I IFN-dependent NK cell functions, contributing to a better understanding of sex-specific differences in innate immunity.