III Interferons Research Articles

Abstract 964: Induction of immunogenic cell death and interferon signaling by carboplatin and the ATR inhibitor M6620 may contribute to anti-tumor activity of M6620-carboplatin-avelumab triplet combination in MC38 tumor model

Abstract Ataxia-telangiectasia mutated and Rad3-related kinase (ATR) is one of the key players in the DNA damage response (DDR) to replication stress. M6620 is a potent and selective ATR inhibitor which has been shown to sensitize cancer cells to the lethal effects of DNA-damaging chemotherapeutic agents. We previously demonstrated that M6620 significantly enhances the anti-tumor efficacy of dual combinations of platinum-based chemotherapy and avelumab, an FDA-approved anti-PD-L1 monoclonal antibody, in syngeneic tumor models in vivo. The purpose of this study was to further elucidate the potential mechanisms of action that drive the therapeutic benefit of this triplet combination therapy. Treatment with certain chemotherapeutic agents has been reported to promote immunogenic cell death (ICD) in tumor cells. We utilized in vitro assays to test the expression of markers of immunogenic cell death in murine MC38 colorectal carcinoma cells. After 72 hours of treatment, carboplatin and M6620 dual combination treatment significantly increased the levels of Calreticulin, HMGB1, and ATP relative to treatment with either single agent. In addition, gene expression profiling was used to determine pathways differentially induced by the drug treatments. RNA-sequencing (RNA-seq) analysis was performed on MC38 cells treated in vitro with carboplatin, M6620, or dual combination therapy. Comparison of gene expression between treatment groups at 48 hours post-treatment revealed that the dual combination therapy significantly increased expression of genes regulated by type I/II interferons (IFN), including CCl5, CXCL10, CXCL11 and ISG15. RNA-seq gene expression analysis was also performed on MC38 tumor samples from mice treated with carboplatin, M6620, avelumab, carboplatin/avelumab dual therapy, or carboplatin/M6620/avelumab triple combination therapy. Triple combination therapy induced a more robust upregulation of IFN-inducible genes at day 3 compared with monotherapies or carboplatin/avelumab dual combination, consistent with in vitro observations that M6620 and carboplatin dual combination therapy enhanced expression of genes regulated by IFNs. Taken together, these data suggest that potentiation of immunogenic cell death and induction of pro-inflammatory responses by M6620, in the context of chemotherapy-induced DNA damage, may contribute to the enhanced anti-tumor efficacy observed with carboplatin, M6620, and avelumab triple combination therapy. Citation Format: Marat Alimzhanov, Ashwin George, Parantu Shah, Astrid Zimmerman, Andreas Schroeder, Martin Falk, Christiane Amendt, Joern-Peter Halle, Andree Blaukat, Frank Zenke, Patricia Soulard. Induction of immunogenic cell death and interferon signaling by carboplatin and the ATR inhibitor M6620 may contribute to anti-tumor activity of M6620-carboplatin-avelumab triplet combination in MC38 tumor model [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 964.

Abstract 5615: Anti-human CD47 antibody Hu5F9 inhibits MC38 tumor growth in hCD47/hSIRPα double knock-in mice

Abstract Introduction: CD47 is broadly expressed on all normal cells or tissues, such as hematopoietic cells, T cells, B cells, monocytes, thymocytes, and platelets as well as other cell lineages, but has reduced expression on Rh erythrocytes. Signal regulatory proteins (SIRP) are a multigene family of immune receptors encoded by a cluster of genes. The most conserved member, SIRPα, binds to CD47 to engage the “don't eat me” signal and avoid macrophage phagocytosis. Notably, CD47 is often highly expressed on various tumors, and is an important immune checkpoint inhibiting macrophage phagocytosis. Therefore, blockade of CD47/SIRPα signalling by an anti-CD47 antibody offers a promising therapeutic approach by unleashing the phagocytosis activity of macrophages against tumor cells. It is well known that the antitumor efficacy of anti-human CD47 antibodies is largely macrophage-dependent in immunocompromised mice (NOD-SCID or NOG®/NSG™) engrafted with xenograft tumors (PDX or cell line derived xenografts). However, the therapeutic effects of anti-CD47 antibodies in immunocompetent mice engrafted with syngeneic tumors have also shown the involvement of cytosolic DNA sensors, dendritic cells, cytotoxic T cells, and type I/II interferons (IFNs). A preclinical animal model with a competent immune system, and humanized CD47/SIRPα is needed to fully recapitulate the mode of action of anti-CD47 therapeutic antibodies. Methods: We developed the human CD47 and SIRPα double knock-in mouse model (CD47/SIRPα HuGEMM™) via CRISPR/Cas9 targeting. We have also engineered mouse syngeneic MC38 cells to express human CD47 (hCD47-MC38 HuCELL™). We engrafted the hCD47-MC38 HuCELL into CD47/SIRPα HuGEMM mice to evaluate the efficacy of the anti-human CD47 antibody Hu5F9. Results: FACS analysis verified that the expression patterns of chimeric h/m CD47 and h/m SIRPα in HuGEMM mice are consistent with the mouse proteins in wild-type C57BL/6 mice. We have also confirmed that an anti-human CD47 antibody (Hu5F9-hIgG4) can bind to human CD47 expressed on hCD47-MC38 HuCELL tumor cells. Finally, we demonstrated robust antitumor effects of an anti-human CD47 antibody by treating the HuGEMM mice bearing hCD47-MC38 tumors with Hu5F9 monotherapy or in combination with anti-PD-1 antibody. Conclusions: Collectively, our hCD47/hSIRPα double knock-in humanized model, along with hCD47-MC38 HuCELL, provides a powerful preclinical platform to evaluate the mode of action, and in vivo efficacy of human-specific anti-CD47 antibodies. It may also serve as a model for in vivo evaluation of human SIRPα antibodies, as well as for proof-of-concept of combination therapies with other immune checkpoint inhibitors or immune modulators. Citation Format: Daniel Xianfei He, Chenpan Nie, Lei Zheng, Ruilin Sun, Annie Xiaoyu An, Henry Q. x. Li, Davy Xuesong Ouyang. Anti-human CD47 antibody Hu5F9 inhibits MC38 tumor growth in hCD47/hSIRPα double knock-in mice [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5615.

Interferon-λ at the Center of the Storm

Type I and III interferons (IFNs) drive effective antiviral functions but differentially affect tissue homeostasis. Using mouse models of severe inflammation, Broggi et al. and Major et al. report in Science that type III IFNs disrupt epithelial cell proliferation and differentiation in the lung.

Interferon-Dependent and Respiratory Virus-Specific Interference in Dual Infections of Airway Epithelia

Many respiratory viruses cocirculate in the population and multiple infections are commonly reported. The clinical impact of coinfection is unclear and may vary depending on the viral couples involved. Using three-dimensional reconstituted human airway epithelia and clinical viral strains, we investigated the interaction between influenza virus (Flu), respiratory syncytial virus (RSV) and rhinovirus (RV). We showed that Flu and RSV interfere with RV replication, whereas RV does not interfere with either of these viruses. We then experimentally demonstrated that, when present, the interference is not related to a block of viral entry but rather to type I and type III interferon (IFN), the front-line antiviral defense of the respiratory mucosa. Consistent with this observation, we highlighted the differential sensitivity of each virus to IFNs, with RV being the only virus significantly inhibited by IFN-λ and the most sensitive to IFN-α. Finally, as type III IFN is of therapeutic interest due to its low proinflammatory profile, we also assessed and confirmed an inhibitory effect of IFN-λ in the context of persistent RV infections. The present work provides mechanistic clues concerning innate immunity involvement during respiratory virus interactions and confirms that IFN-λ is a promising candidate in the treatment of RV infections.

Innate Immunity in Hepatitis C Virus Infection.

Activation and viral control of the innate immune response are hallmarks of hepatitis C virus (HCV) infection and are major determinants of spontaneous clearance or progression to chronic infection and liver disease. In this review, we provide a contemporary overview of how HCV is sensed by the host cell to trigger innate immune activation and the mechanisms deployed by the virus to evade this response. Type I and III interferons (IFNs) are crucial mediators of antiviral innate immunity against HCV, and we specifically highlight the importance of IFN-λ host genetics for the outcome of HCV infection. Last, we focus on the proinflammatory responses elicited by HCV infection and describe our current understanding of how interleukin (IL)-1β signaling and cross talk between the IL-1β and IFN signaling pathways lead to sustained inflammation and increased risk of liver pathology.

Suppressing PARylation by 2',5'-oligoadenylate synthetase 1 inhibits DNA damage-induced cell death.

High expression of 2',5'-oligoadenylate synthetase 1 (OAS1), which adds AMP residues in 2',5' linkage to a variety of substrates, is observed in many cancers as a part of the interferon-related DNA damage resistance signature (IRDS). Poly(ADP-ribose) (PAR) is rapidly synthesized from NAD+ at sites of DNA damage to facilitate repair, but excessive PAR synthesis due to extensive DNA damage results in cell death by energy depletion and/or activation of PAR-dependent programmed cell death pathways. We find that OAS1 adds AMP residues in 2',5' linkage to PAR, inhibiting its synthesis invitro and reducing its accumulation in cells. Increased OAS1 expression substantially improves cell viability following DNA-damaging treatments that stimulate PAR synthesis during DNA repair. We conclude that high expression of OAS1 in cancer cells promotes their ability to survive DNA damage by attenuating PAR synthesis and thus preventing cell death.

Subversion of Host Innate Immunity by Human Papillomavirus Oncoproteins.

The growth of human papillomavirus (HPV)-transformed cells depends on the ability of the viral oncoproteins E6 and E7, especially those from high-risk HPV16/18, to manipulate the signaling pathways involved in cell proliferation, cell death, and innate immunity. Emerging evidence indicates that E6/E7 inhibition reactivates the host innate immune response, reversing what until then was an unresponsive cellular state suitable for viral persistence and tumorigenesis. Given that the disruption of distinct mechanisms of immune evasion is an attractive strategy for cancer therapy, the race is on to gain a better understanding of E6/E7-induced immune escape and cancer progression. Here, we review recent literature on the interplay between E6/E7 and the innate immune signaling pathways cGAS/STING/TBK1, RIG-I/MAVS/TBK1, and Toll-like receptors (TLRs). The overall emerging picture is that E6 and E7 have evolved broad-spectrum mechanisms allowing for the simultaneous depletion of multiple rather than single innate immunity effectors. The cGAS/STING/TBK1 pathway appears to be the most heavily impacted, whereas the RIG-I/MAVS/TBK1, still partially functional in HPV-transformed cells, can be activated by the powerful RIG-I agonist M8, triggering the massive production of type I and III interferons (IFNs), which potentiates chemotherapy-mediated cell killing. Overall, the identification of novel therapeutic targets to restore the innate immune response in HPV-transformed cells could transform the way HPV-associated cancers are treated.

Foot and mouth disease virus undergoes non-progressive replication in mice peritoneal macrophages and induces M1 polarization

Despite the fact that macrophages link the innate and adaptive arms of immunity, it’s role in the early infection of foot and mouth disease virus (FMDV) is largely unknown. Recently, depletion of macrophages in vivo after vaccination has shown to drastically diminish the protection against FMDV challenge in mouse model. Even the ability of macrophages to reduce or resist FMDV infection is not known hitherto. Therefore, we examined the replication ability of FMDV in mice peritoneal macrophages and the responsiveness in terms of macrophage polarization and cytokine production. Negative strand specific RT-PCR indicated replication of FMDV RNA in macrophages. Absolute quantitation of FMDV transcripts, immunofluorescence studies and titre of the infectious progeny virus revealed that replication peaked at 12 hpi and significantly declined by 18 hpi indicating non-progressive replication in the infected macrophages. Further, significant up regulation of inducible nitric oxide synthase by 8 –12 hpi and increase of M1 specific CD11c + cells by 42.6 % after infection showed that FMDV induce M1 polarization. A significant up regulation of TNFα and IL12 transcripts at 8 hpi supported that M1 macrophages were functional. Further, we studied the expression of Type I to III interferons (IFN) and other antiviral molecules. The results indicate a marked up regulation of Type I IFNα and β by 9.2 and 11.2 fold, respectively at 8 hpi. Of the four IFN stimulated genes (ISG), viperin showed a significant up regulation by 286-fold at 12 hpi in the mice macrophages. In conclusion, the results suggest that replication of FMDV in mice peritoneal macrophages is non-progressive with up regulation of Type I IFN and ISGs. Further, FMDV induces M1 polarization in murine peritoneal macrophages.

A pull-down assay using DNA/RNA-conjugated beads with a customized competition strategy: An effective approach to identify DNA/RNA binding proteins

Innate immune response is insisted upon detection of foreign intracellular DNA or RNA derived from viruses and bacteria. This reaction is important to initiate an effective protective response for the host cells. This crucial step is induced by cytosolic nucleic acids sensors/binding proteins, which triggers the production of type I or type III interferons (IFNs) and proinflammatory cytokines such as Interleukin 6 (IL-6). The identification of these cytosolic DNA or RNA sensors is a key step in understanding the signaling pathways triggered by those pathogens. Here we describe an effective approach to identify potential known/novel DNA or RNA sensors: a pull-down assay using DNA/RNA-conjugated beads with a customized competition strategy, which conferred a more effective and efficient way to determine the interaction between DNA/RNA and the sensor protein(s), therefore greatly improves the progress to investigate potential novel cytosolic DNA or RNA sensors/ binding proteins •The customized method makes the traditional pull-down assays more effective and efficient to identify DNA/RNA binding protein(s).•With the competitor of your choice, the method provides specific information about the competitive binding between DNA/RNA and binding proteins.

FOXO3a regulates rhinovirus-induced innate immune responses in airway epithelial cells

Forkhead transcription factor class O (FOXO)3a, which plays a critical role in a wide variety of cellular processes, was also found to regulate cell-type-specific antiviral responses. Airway epithelial cells express FOXO3a and play an important role in clearing rhinovirus (RV) by mounting antiviral type I and type III interferon (IFN) responses. To elucidate the role of FOXO3a in regulating antiviral responses, we generated airway epithelial cell-specific Foxo3a knockout (Scga1b1-Foxo3a−/−) mice and a stable FOXO3a knockout human airway epithelial cell line. Compared to wild-type, Scga1b1-Foxo3a−/− mice show reduced IFN-α, IFN-β, IFN-λ2/3 in response to challenge with RV or double-stranded (ds)RNA mimic, Poly Inosinic-polycytidylic acid (Poly I:C) indicating defective dsRNA receptor signaling. RV-infected Scga1b1-Foxo3a−/− mice also show viral persistence, enhanced lung inflammation and elevated pro-inflammatory cytokine levels. FOXO3a K/O airway epithelial cells show attenuated IFN responses to RV infection and this was associated with conformational change in mitochondrial antiviral signaling protein (MAVS) but not with a reduction in the expression of dsRNA receptors under unstimulated conditions. Pretreatment with MitoTEMPO, a mitochondrial-specific antioxidant corrects MAVS conformation and restores antiviral IFN responses to subsequent RV infection in FOXO3a K/O cells. Inhibition of oxidative stress also reduces pro-inflammatory cytokine responses to RV in FOXO3a K/O cells. Together, our results indicate that FOXO3a plays a critical role in regulating antiviral responses as well as limiting pro-inflammatory cytokine expression. Based on these results, we conclude that FOXO3a contributes to optimal viral clearance and prevents excessive lung inflammation following RV infection.

In vitro production of synthetic viral RNAs and their delivery into mammalian cells and the application of viral RNAs in the study of innate interferon responses.

Mammalian cells express different types of RNA molecules that can be classified as protein coding RNAs (mRNA) and non-coding RNAs (ncRNAs) the latter of which have housekeeping and regulatory functions in cells. Cellular RNAs are not recognized by cellular pattern recognition receptors (PRRs) and innate immunity is not activated. RNA viruses encode and express RNA molecules that usually differ from cell-specific RNAs and they include for instance 5'capped and 5'mono- and triphosphorylated RNAs, small viral RNAs and viral RNA-protein complexes called vRNPs. These molecules are recognized by certain members of Toll-like receptor (TLR) and RIG-I-like receptor (RLR) families leading to activation of innate immune responses and the production of antiviral cytokines, such as type I and type III interferons (IFNs). Virus-specific ssRNA and dsRNA molecules that mimic the viral genomic RNAs or their replication intermediates can efficiently be produced by bacteriophage T7 DNA-dependent RNA polymerase and bacteriophage phi6 RNA-dependent RNA polymerase, respectively. These molecules can then be delivered into mammalian cells and the mechanisms of activation of innate immune responses can be studied. In addition, synthetic viral dsRNAs can be processed to small interfering RNAs (siRNAs) by a Dicer enzyme to produce a swarm of antiviral siRNAs. Here we describe the biology of RNAs, their in vitro production and delivery into mammalian cells as well as how these molecules can be used to inhibit virus replication and to study the mechanisms of activation of the innate immune system.

2608. Restriction of Rhinovirus Infection Depends on Virus Sensing and Early IFN Induction

BackgroundHuman rhinovirus (RV) infections are ubiquitous, underestimated, and costly. RV causes 3–12 infections per year per individual with a wide range of clinical presentations from mild upper respiratory infections to severe viral pneumonias (1). The virus-host interactions that control RV infection are poorly understood. Thus, there are no vaccines or antiviral medications available. RV infection begins at the airway epithelial surface where the virus first encounters the host cell immune defenses, including type I and III interferon (IFN). IFN actions are critical for defense against RV infection wherein interferon-stimulated genes (ISG)s direct antiviral actions to limit RV infection.MethodsWe hypothesized that the timing of IFN induction is a critical determinant of RV restriction by host innate immune defenses in the human respiratory tract. Thus, an immortalized bronchial epithelial cell line was infected with RV-14 with multiplicity of infection (MOI) ranging from 0.1 – 10 and under conditions of pre/post infection treatment with IFN-β or IFN-λ. Host and viral RNA, protein, and RV infectious particle levels were analyzed.ResultsWe found that RV infection induces IFN-β and IFN-λ production and subsequent ISG induction, including expression of IFIT-1, OAS1, and MX1. RV-14 infection induced IFN-β and IFN-λ in a dose-dependent manner, with a maximum fold increase of IFN expression at 48hours post infection. ISGs were induced in a similar pattern to IFNs. Viral titers increased significantly over the first 24hours post infection and then plateaued through 96hours. IFN-β and IFN-λ pre- and posttreatment conditions significantly decreased maximum viral titers achieved but with continued viral plateaus 24–96 hours post infection.ConclusionOur observations demonstrate that RV induces innate immune activation and the production of type I and III IFN during acute infection of airway cells. Sustained viral titer plateaus, despite antiviral ISG induction, suggests viral blocking of IFN pathway mechanisms that can be overcome by early IFN induction to significantly restrict RV viral replication.1. Royston L, Tapparel C. 2016. Rhinoviruses and Respiratory Enteroviruses: Not as Simple as ABC. Viruses 8.Disclosures All authors: No reported disclosures.

975. Roles of Type I and III Interferon in Severe Pathogenesis of Human Metapneumovirus

BackgroundHuman metapneumovirus (HMPV) is a leading cause of respiratory tract infection in children and adults. However, mechanisms of pathogenesis are not fully understood.MethodsWe tested HMPV clinical and laboratory isolates in an established C57BL/6 mouse model and measured weight loss, airway function, and viral titers. Immune responses were determined using cytokine quantitation and flow cytometry.ResultsHMPV clinical isolates induced variable disease severity ranging from mild to fatal disease. Laboratory strain TN/94-49 did not cause weight loss, but mice infected with clinical isolate C2-202 showed dramatic weight loss and 40% mortality within 5 days post-infection (Figure 1). These findings were confirmed in other inbred mouse strains. C2-202-infected mice also suffered from impaired pulmonary function post-recovery. Lung viral titer did not correlate with disease severity, suggesting immune-mediated pathogenesis. C2-202-infected mice exhibited increased production of type I and III interferons (IFN) and pro-inflammatory cytokines, and lung neutrophil infiltration. However, neutrophil depletion or inflammasome inactivation did not reduce disease. Stat1/Stat2 double knockout (KO) mice lacking type I and III IFN signaling exhibited reduced weight loss but increased lung viral titer after C2-202 infection (Figure 2). Type I IFN receptor (IFNAR) KO mice infected with C2-202 had reduced weight loss but unchanged lung viral titer (Figure 3), while the addition of type III IFN blockade to C2-202-infected IFNAR mice had no effect on disease but increased lung viral titer (Figure 4).ConclusionThese results suggest that severe disease caused by virulent HMPV was due to exuberant IFN response. Moreover, type I IFN was primarily associated with disease, while type III IFN was associated with viral clearance. These data suggest that IFN signaling plays an important role in HMPV pathogenesis, and thus serves as a potential therapeutic target. Disclosures All Authors: No reported Disclosures.

Respiratory syncytial virus nonstructural proteins 1 and 2: Exceptional disrupters of innate immune responses.

Human respiratory syncytial virus (RSV) is the most important cause of acute lower respiratory tract disease in infants worldwide. As a first line of defense against respiratory infections, innate immune responses, including the production of type I and III interferons (IFNs), play an important role. Upon infection with RSV, multiple pattern recognition receptors (PRRs) can recognize RSV-derived pathogen-associated molecular patterns (PAMPs) and mount innate immune responses. Retinoic-acid-inducible gene-I (RIG-I) and nucleotide-binding oligomerization domain-containing protein 2 (NOD2) have been identified as important innate receptors to mount type I IFNs during RSV infection. However, type I IFN levels remain surprisingly low during RSV infection despite strong viral replication. The poor induction of type I IFNs can be attributed to the cooperative activity of 2 unique, nonstructural (NS) proteins of RSV, i.e., NS1 and NS2. These viral proteins have been shown to suppress both the production and signaling of type I and III IFNs by counteracting a plethora of key host innate signaling proteins. Moreover, increasing numbers of IFN-stimulated genes (ISGs) are being identified as targets of the NS proteins in recent years, highlighting an underexplored protein family in the identification of NS target proteins. To understand the diverse effector functions of NS1 and NS2, Goswami and colleagues proposed the hypothesis of the NS degradasome (NSD) complex, a multiprotein complex made up of, at least, NS1 and NS2. Furthermore, the crystal structure of NS1 was resolved recently and, remarkably, identified NS1 as a structural paralogue of the RSV matrix protein. Unfortunately, no structural data on NS2 have been published so far. In this review, we briefly describe the PRRs that mount innate immune responses upon RSV infection and provide an overview of the various effector functions of NS1 and NS2. Furthermore, we discuss the ubiquitination effector functions of NS1 and NS2, which are in line with the hypothesis that the NSD shares features with the canonical 26S proteasome.

817 Identification of Biomarkers and Mechanistic Insight for Upadacitinib in Crohn’s Disease: Serum Inflammatory Mediator Analysis From the Phase 2b CELEST Study

INTRODUCTION: We report pharmacodynamic profiling of serum inflammatory mediators in CELEST (patients) pts with the aim of understanding UPA mechanism of action in CD and linking biomarkers to intestinal inflammation, abdominal pain (AP), and stool frequency (SF). METHODS: The CELEST Ph2 study evaluated the efficacy/ safety of multiple dosing regimens of UPA vs PBO in adults with moderately to severely active CD and inadequate response or intolerance to immunomodulators and/or TNF inhibitors.1,2 Serum samples from baseline (BL) visit and end of induction period (Wk16) were analyzed by OLINK® inflammation panel (92 proteins) and Singulex immunoassay for interleukin-17A(IL-17A), IL-17F, IL-22, and IL-23p19. Protein level changes (BL–Wk16) were analyzed by an ANCOVA model where BL protein level was adjusted as a covariate and treatment effect was considered significant when a false discovery rate < 0.10 was observed after multiplicity adjustment on F-test P-values. Spearman rank correlation coefficients(rs) determined relationships between BL levels or BL–Wk16 level changes of serum biomarkers (also including serum C-reactive protein and fecal calprotectin) and improvements in Simple Endoscopic Score for CD (SES-CD), AP, and SF. RESULTS: Paired BL and Wk16 serum samples were available from 104 pts (UPA 3 mg twice daily [BID], n = 18; 6 mg BID, n = 19; 12 mg BID, n = 13; 24 mg BID, n = 18; 24 mg once daily, n = 18; and PBO, n = 18). UPA treatment significantly reduced expression of serum pro-inflammatory mediators (Table). that are associated with immune cell migration, type I/II interferon (IFN) responses, T cell activation, T helper cell-1 (Th1) responses, and CD8+ T cell responses. UPA treatment did not significantly modulate expression of IL-23/Th17-related serum cytokines in CD. Correlations were observed for BL–Wk16 changes in serum proteins and SES-CD, AP, or SF. Importantly, oncostatin-M reductions correlated with SES-CD (rs = 0.35, P = 0.02) and SF improvement (rs = 0.33, P = 0.04), suggesting similar drivers of these aspects of disease. AP improvement correlated with reductions in CSF-1 (rs = 0.37, P = 0.01), TNFRSF9 (rs = 0.34, P = 0.03), IL-12B (rs = 0.31, P = 0.07), CXCL9 (rs = 0.31, P = 0.07), and CD8A (rs = 0.31, P = 0.09), which are all associated with the Th1/CD8/IFN-γ pathway. CONCLUSION: This analysis of inflammatory mediators in serum of CD pts from CELEST identifies potential pharmacodynamic biomarkers and mechanistic insights for the JAK-1 inhibitor, UPA.

Differential interferon gene expression in bronchiolitis caused by respiratory syncytial virus-A genotype ON1.

Bronchiolitis severity is determined by a complex interaction among viral replication and antiviral immunity. The current respiratory syncytial virus (RSV)-A, genotype ON1 demonstrated a high replicative capacity but seemed to be clinically less severe than the previously circulating RSV-A, NA1. To learn insights about ON1 innate immune response, we analyzed expression levels of type I/III interferon (IFN)-related genes in the respiratory mucosa of infants with RSV bronchiolitis. We enrolled RSV-positive bronchiolitis patients over 12 epidemic seasons at a university hospital in Rome. From nasopharyngeal washings' cells (46 positive to NA1, 47 to ON1 and 28 to RSV-B, genotype BA), the mRNA copy number of the type III IFN receptor (IFNLR1 and IL10RB subunits), and of the type I/III IFN-stimulated genes, MxA and ISG56, was calculated using the threshold cycle relative quantification method with respect to an invariant gene. Expression levels of type III IFN receptor subunits genes positively correlated to each other and did not differ in infants infected with different RSV genotypes. The ISGs levels also positively correlated between them but differed among groups. MxA levels were significantly higher in NA1-infected infants than in those with ON1 and BA; ISG56 expression was slightly higher in NA1 than in the other strains. Interestingly, a moderate negative correlation existed between viral load and both ISGs values in ON1-infected infants only. The reduced ISG levels elicited during infections with ON1 (and BA) may cause a weaker control of RSV replication and/or an inadequate host immune response which may impact the risk of respiratory sequelae.

Why Is IFN-λ Less Inflammatory? One IRF Decides

Type I and type III interferons (IFNs) activate similar antiviral transcriptional programs, but the type I IFN response is more inflammatory. In this issue of Immunity, Forero etal. find that selective induction of the transcription factor IRF1 promotes proinflammatory chemokine expression downstream of type I IFN signaling.

Differential Activation of the Transcription Factor IRF1 Underlies the Distinct Immune Responses Elicited by Type I and Type III Interferons

Type I and III interferons (IFNs) activate similar downstream signaling cascades, but unlike type I IFNs, type III IFNs (IFNλ) do not elicit strong inflammatory responses invivo. Here, we examined the molecular mechanisms underlying this disparity. Type I and III IFNs displayed kinetic differences in expression ofIFN-stimulated genes and proinflammatory responses, with type I IFNs preferentially stimulating expression of the transcription factor IRF1. Type III IFNs failed to induce IRF1 expression because of low IFNλ receptor abundance and insufficient STAT1 activation on epithelial cells and thus did not activate the IRF1 proinflammatory gene program. Rather, IFNλ stimulation preferentially induced factors implicated in tissue repair. Our findings suggest that IFN receptor compartmentalization and abundance confer a spatiotemporal division of labor where type III IFNs control viral spread at the site of the infection while restricting tissue damage; the transient induction of inflammatory responses by type I IFNs recruits immune effectors to promote protective immunity.

IFN-λ3 as a host immune response in acute hepatitis E virus infection

Background and aimHepatitis E virus (HEV) is mainly transmitted orally, either waterborne or zoonotic foodborne. Intestinal viruses such as rotavirus are known to induce type III interferon (IFN) in the gastrointestinal (GI) tract where type III IFN dominantly functions in comparison with type I IFN. Therefore, the aim of this study is to investigate the significance of type III IFN (IFN-λ3) in acute hepatitis E. MethodsIFN-λ3 and HEV RNA levels in the sera of patients with acute HEV infection and in the supernatant of HEV-inoculated cells were measured, using an in-house high-sensitivity method and reverse transcription-polymerase chain reaction, respectively. ResultsHigh serum IFN-λ3 levels were found in the early phase of acute HEV infection, which normalized after resolution. Interestingly, serum IFN-λ3 levels correlated well with serum HEV RNA titers in the same sera, both of which showed the peak before the robust increase of transaminases. In vitro experiments demonstrated that HEV replicated well in the cells with little IFN-λ3 induction (Caco-2, A549) and recombinant IFN-λ3 inhibited HEV replication in a dose-dependent manner. In contrast, in HT-29 cells, a colon cancer cell line, HEV poorly replicated and induced IFN-λ3 in a titer-dependent manner. ConclusionsThese clinical and experimental observations suggest that HEV induced IFN-λ3 as a host innate immune response, which may play a protective role against HEV.

Signaling Pathways of Type I and Type III Interferons and Targeted Therapies in Systemic Lupus Erythematosus

Type I and type III interferons (IFNs) share several properties in common, including the induction of signaling pathways, the activation of gene transcripts, and immune responses, against viral infection. Recent advances in the understanding of the molecular basis of innate and adaptive immunity have led to the re-examination of the role of these IFNs in autoimmune diseases. To date, a variety of IFN-regulated genes, termed IFN signature genes, have been identified. The expressions of these genes significantly increase in systemic lupus erythematosus (SLE), highlighting the role of type I and type III IFNs in the pathogenesis of SLE. In this review, we first discussed the signaling pathways and the immunoregulatory roles of type I and type III IFNs. Next, we discussed the roles of these IFNs in the pathogenesis of autoimmune diseases, including SLE. In SLE, IFN-stimulated genes induced by IFN signaling contribute to a positive feedback loop of autoimmunity, resulting in perpetual autoimmune inflammation. Based on this, we discussed the use of several specific IFN blocking strategies using anti-IFN-α antibodies, anti-IFN-α receptor antibodies, and IFN-α-kinoid or downstream small molecules, which intervene in Janus kinase (JAK)-signal transducer and activator of transcription (STAT) pathways, in clinical trials for SLE patients. Hopefully, the development of novel regimens targeting IFN signaling pathways will shed light on promising future therapeutic applications for SLE patients.