IFNAR1 Research Articles

Anifrolumab in systemic lupus erythematosus: a profile of its use

Anifrolumab (Saphnelo™), an intravenously administered immunoglobulin G1 kappa monoclonal antibody targeting the type 1 interferon alpha receptor, is effective and generally well tolerated as an add-on to standard therapy in adults with moderate to severe systemic lupus erythematosus (SLE). Anifrolumab reduces overall disease activity in patients with moderate to severe SLE despite receiving stable standard-of-care treatment, based on the totality of evidence from randomized, double-blind, placebo-controlled, phase 2b and 3 clinical trials. Anifrolumab also has corticosteroid-sparing effects and reduces skin disease severity. Therapeutic benefits of anifrolumab appear to be sustained over longer-term treatment. With respect to tolerability, anifrolumab recipients are at increased risk of infections (including herpes zoster). Additional long-term data are being collected in an ongoing extension of the phase 3 trials.

Sequence-Specific Features of Short Double-Strand, Blunt-End RNAs Have RIG-I- and Type 1 Interferon-Dependent or -Independent Anti-Viral Effects.

Pathogen-associated molecular patterns, including cytoplasmic DNA and double-strand (ds)RNA trigger the induction of interferon (IFN) and antiviral states protecting cells and organisms from pathogens. Here we discovered that the transfection of human airway cell lines or non-transformed fibroblasts with 24mer dsRNA mimicking the cellular micro-RNA (miR)29b-1* gives strong anti-viral effects against human adenovirus type 5 (AdV-C5), influenza A virus X31 (H3N2), and SARS-CoV-2. These anti-viral effects required blunt-end complementary RNA strands and were not elicited by corresponding single-strand RNAs. dsRNA miR-29b-1* but not randomized miR-29b-1* mimics induced IFN-stimulated gene expression, and downregulated cell adhesion and cell cycle genes, as indicated by transcriptomics and IFN-I responsive Mx1-promoter activity assays. The inhibition of AdV-C5 infection with miR-29b-1* mimic depended on the IFN-alpha receptor 2 (IFNAR2) and the RNA-helicase retinoic acid-inducible gene I (RIG-I) but not cytoplasmic RNA sensors MDA5 and ZNFX1 or MyD88/TRIF adaptors. The antiviral effects of miR29b-1* were independent of a central AUAU-motif inducing dsRNA bending, as mimics with disrupted AUAU-motif were anti-viral in normal but not RIG-I knock-out (KO) or IFNAR2-KO cells. The screening of a library of scrambled short dsRNA sequences identified also anti-viral mimics functioning independently of RIG-I and IFNAR2, thus exemplifying the diverse anti-viral mechanisms of short blunt-end dsRNAs.

Immunomodulatory and direct activities of ropeginterferon alfa-2b on cancer cells in mouse models of leukemia.

Although ropeginterferon alfa‐2b has recently been clinically applied to myeloproliferative neoplasms with promising results, its antitumor mechanism has not been thoroughly investigated. Using a leukemia model developed in immunocompetent mice, we evaluated the direct cytotoxic effects and indirect effects induced by ropeginterferon alfa‐2b in tumor cells. Ropeginterferon alfa‐2b therapy significantly prolonged the survival of mice bearing leukemia cells and led to long‐term remission in some mice. Alternatively, conventional interferon‐alpha treatment slightly extended the survival and all mice died. When ropeginterferon alfa‐2b was administered to interferon‐alpha receptor 1–knockout mice after the development of leukemia to verify the direct effect on the tumor, the survival of these mice was slightly prolonged; nevertheless, all of them died. In vivo CD4+ or CD8+ T‐cell depletion resulted in a significant loss of therapeutic efficacy in mice. These results indicate that the host adoptive immunostimulatory effect of ropeginterferon alfa‐2b is the dominant mechanism through which tumor cells are suppressed. Moreover, mice in long‐term remission did not develop leukemia, even after tumor rechallenge. Rejection of rechallenge tumors was canceled only when both CD4+ and CD8+ T cells were removed in vivo, which indicates that each T‐cell group functions independently in immunological memory. We show that ropeginterferon alfa‐2b induces excellent antitumor immunomodulation in hosts. Our finding serves in devising therapeutic strategies with ropeginterferon alfa‐2b.

Targeting the Annexin A1-FPR2/ALX pathway for host-directed therapy in dengue disease.

Host immune responses contribute to dengue's pathogenesis and severity, yet the possibility that failure in endogenous inflammation resolution pathways could characterise the disease has not been contemplated. The pro-resolving protein Annexin A1 (AnxA1) is known to counterbalance overexuberant inflammation and mast cell (MC) activation. We hypothesised that inadequate AnxA1 engagement underlies the cytokine storm and vascular pathologies associated with dengue disease. Levels of AnxA1 were examined in the plasma of dengue patients and infected mice. Immunocompetent, interferon (alpha and beta) receptor one knockout (KO), AnxA1 KO, and formyl peptide receptor 2 (FPR2) KO mice were infected with dengue virus (DENV) and treated with the AnxA1 mimetic peptide Ac2-26 for analysis. In addition, the effect of Ac2-26 on DENV-induced MC degranulation was assessed in vitro and in vivo. We observed that circulating levels of AnxA1 were reduced in dengue patients and DENV-infected mice. Whilst the absence of AnxA1 or its receptor FPR2 aggravated illness in infected mice, treatment with AnxA1 agonistic peptide attenuated disease manifestationsatteanuated the symptoms of the disease. Both clinical outcomes were attributed to modulation of DENV-mediated viral load-independent MC degranulation. We have thereby identified that altered levels of the pro-resolving mediator AnxA1 are of pathological relevance in DENV infection, suggesting FPR2/ALX agonists as a therapeutic target for dengue disease.

Genetic variants of interferon lambda-related genes and chronic kidney disease susceptibility in the Korean population.

BackgroundChronic kidney disease (CKD) is a common condition leading to renal dysfunction and is closely related to increased cardiovascular and mortality risk. CKD is an important public health issue, and recent genetic studies have verified common CKD susceptibility variants. This research examines the interrelationship between candidate genes polymorphisms of interferon lambda (IFNL) induction, its signaling pathway, and CKD.MethodsSeventy-five patients with advanced CKD and 312 healthy subjects (as controls) participated in this research. A replication set composed of 172 patients with advanced CKD and 365 controls was used for additional analysis. The genotype of single nucleotide polymorphisms (SNPs) was determined by the Axiom Genome-Wide Human Assay and SNaPshot assay.ResultsThe SNP of IFNL3 was significantly associated with CKD in the codominant (p = 0.02) and dominant models (p = 0.02). In addition, the SNPs of IFNL2 were significantly associated with CKD in the dominant model (p = 0.03), and the SNP of interferon alpha receptor 2 (IFNAR2) was significantly associated with CKD in the log-additive model (p = 0.03). Concerning rs148543092, in the IFNL3 gene, a significant association was observed after pooling the original and replication sets.ConclusionThese results indicate that SNPs in the IFNL induction and signal pathway may be associated with CKD risk in the Korean population. Finally, our results also show that the IFNL3 gene variant may be associated with CKD risk.

Abstract P2-11-14: Pathology AI features and immune biomarkers of postpartum involution: Implications for postpartum breast cancer

Abstract Breast cancer risk is increased in parous women in the period following childbirth, peaking at five years. After weaning or in the absence of lactation, the breast undergoes postpartum involution (PPI), a process characterized by epithelial cell death and tissue remodeling that returns the mammary gland to a baseline state, and has previously been associated with increased inflammation and breast cancer development and progression in preclinical models. Features of PPI and determinants of related inflammation have been incompletely characterized among women. In this study, we performed visual and AI analysis of 725 digitized H&E images of normal breast tissues donated to the Komen Tissue Bank (KTB) by parous and nulliparous women ≤45 years, and further investigated the immune microenvironment in specimens from 23 women (also from KTB, 13 parous, 10 nulliparous) using NanoString RNA immuno oncology (IO) 360 panels and immune protein biomarkers by NanoString GeoMx Digital Spatial Profiling (DSP). We found that recently parous women (≤5 years of a last birth) had higher numbers of terminal duct lobular units (TDLUs), immune cells, and plasma cells as compared with nulliparous women, and that beyond 5 years of a last birth, TDLU numbers were still higher in parous women, but the immune and plasma cell content of the breast of parous and nulliparous women were not significantly different. Our gene expression analysis revealed differential expression of genes that clustered according to parity status, with interferon stimulated genes; STAT1 and IFITM1, and interferon alpha receptor 1 (IFNAR1), and MHC class II genes (HLA-DMA, HLA-DPA,HLA-DPB1, and HLA-DRA) elevated in parous women. Our DSP analysis revealed that immune biomarkers with immunosuppressive functions, including ARG1, VISTA, CTLA-4, and CD68 were significantly higher in parous women as compared with nulliparous women. Breast tissues of parous women show lobular expansion and alterations in immune cell components compared with nulliparous tissues. Further research is needed to assess how these alterations may impact risk. Citation Format: Joshua Were Ogony, Thomas De Bel, Derek Radisky, Jeroen VanderLaak, Mark Sherman. Pathology AI features and immune biomarkers of postpartum involution: Implications for postpartum breast cancer [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P2-11-14.

Curcumin assists anti-EV71 activity of IFN-\u03b1 by inhibiting IFNAR1 reduction in SH-SY5Y cells

Background and aimEnterovirus 71(EV71) can cause severe hand, foot, and mouth disease (HFMD) with brain tissue involvement. Few effective anti-EV71 drugs are presently available in clinical practice. Interferon-α (IFN-α) was ineffective while Curcumin was effective in restricting EV71 replication in non-neuronal cells. Ubiquitin–proteasome-mediated degradation of interferon-alpha receptor 1 (IFNAR1) protein contributes to IFN-α resistance. Current study aimed to determine synergistic inhibition of EV71 by Curcumin and IFN-α in human neuroblastoma SH-SY5Y cells.MethodsSH-SY5Y cells were infected with mock-/Curcumin-pre-incubated EV71 or transfected with plasmid containing interferon-stimulated response element (ISRE) or mRNA containing viral internal ribosomal entry site (IRES) following by post-treatment with Curcumin with or without IFN-α. Supernatant IFN-α/β was detected by ELISA. ISRE, IRSE, proteasome and deubiquitinating activity were measured by luciferase assay. EV71 RNA and viral protein or IFNAR1 were determined by qPCR and western blot, respectively.ResultsEV71 flailed to completely block IFN-α/β production but inhibited IFN-α signal. Curcumin only slightly inhibited EV71 proliferation without modulating virus attachment and internalization. However, Curcumin addition restored IFN-α-mediated ISRE activity thus significantly inhibiting EV71 replication. Furthermore, EV71 also reduced IFNAR1 protein with proteasome-dependence in SH-SY5Y cells, which can be reversed by Curcumin addition with the evidence that it lowered proteasome activity.ConclusionThese data demonstrate that Curcumin assists anti-EV71 activity of IFN-α by inhibiting IFNAR1 reduction via ubiquitin–proteasome disruption in SH-SY5Y cells.

Long non-coding RNA prostate cancer-associated transcript 6 inhibited gefitinib sensitivity of non-small cell lung cancer by serving as a competing endogenous RNA of miR-326 to up-regulate interferon-alpha receptor 2

ABSTRACT The critical roles of lncRNAs in drug resistance of malignancies have been widely recognized. This investigation aims to study the function of lncRNA PCAT6 in the resistance of non-small cell lung cancer (NSCLC) to gefitinib. In our study, we demonstrated that prostate cancer-associated transcript 6 (PCAT6) was upregulated in gefitinib-resistant NSCLC. PCAT6 knockdown inhibited gefitinib resistance of NSCLC, as indicated by decreased IC50 value, proliferation, and metastasis, and increased cell apoptosis. Besides, PCAT6 could directly target miR-326 in gefitinib-resistant NSCLC cells and augment NSCLC resistance to gefitinib by serving as ceRNA of miR-326. Furthermore, interferon-alpha receptor 2 (IFNAR2) was validated as a downstream target of miR-326 and miR-326 reduced resistance to gefitinib by inhibiting IFNAR2 expression. Our investigation identified that PCAT6 enhanced gefitinib resistance of NSCLC via miR-326/IFNAR2 axis, which might offer a new therapeutic strategy against gefitinib resistance of NSCLC patients.

Antiviral Toll-like Receptor Signaling in Non-Parenchymal Liver Cells Is Restricted to TLR3.

The role of non-parenchymal liver cells as part of the hepatic, innate immune system in the defense against hepatotropic viruses is not well understood. Here, primary human Kupffer cells, liver sinusoidal endothelial cells and hepatic stellate cells were isolated from liver tissue obtained after tumor resections or liver transplantations. Cells were stimulated with Toll-like receptor 1–9 ligands for 6–24 h. Non-parenchymal liver cells expressed and secreted inflammatory cytokines (IL6, TNF and IL10). Toll-like receptor- and cell type-specific downstream signals included the phosphorylation of NF-κB, AKT, JNK, p38 and ERK1/2. However, only supernatants of TLR3-activated Kupffer cells, liver sinusoidal endothelial cells and hepatic stellate cells contained type I and type III interferons and mediated an antiviral activity in the interferon-sensitive subgenomic hepatitis C virus replicon system. The antiviral effect could not be neutralized by antibodies against IFNA, IFNB nor IFNL, but could be abrogated using an interferon alpha receptor 2-specific neutralization. Interestingly, TLR3 responsiveness was enhanced in liver sinusoidal endothelial cells isolated from hepatitis C virus-positive donors, compared to uninfected controls. In conclusion, non-parenchymal liver cells are potent activators of the hepatic immune system by mediating inflammatory responses. Furthermore, liver sinusoidal endothelial cells were identified to be hyperresponsive to viral stimuli in chronic hepatitis C virus infection.

Review: Protective Immunity and Immunopathology of Ehrlichiosis.

Human monocytic ehrlichiosis, a tick transmitted infection, ranges in severity from apparently subclinical to a fatal toxic shock-like fatal disease. Models in immunocompetent mice range from an abortive infection to uniformly lethal depending on the infecting Ehrlichia species, dose of inoculum, and route of inoculation. Effective immunity is mediated by CD4+ T lymphocytes and gamma interferon. Lethal infection occurs with early overproduction of proinflammatory cytokines and overproduction of TNF alpha and IL-10 by CD8+ T lymphocytes. Furthermore, fatal ehrlichiosis is associated with signaling via TLR 9/MyD88 with upregulation of several inflammasome complexes and secretion of IL-1 beta, IL-1 alpha, and IL-18 by hepatic mononuclear cells, suggesting activation of canonical and noncanonical inflammasome pathways, a deleterious role for IL-18, and the protective role for caspase 1. Autophagy promotes ehrlichial infection, and MyD88 signaling hinders ehrlichial infection by inhibiting autophagy induction and flux. Activation of caspase 11 during infection of hepatocytes by the lethal ehrlichial species after interferon alpha receptor signaling results in the production of inflammasome-dependent IL-1 beta, extracellular secretion of HMGB1, and pyroptosis. The high level of HMGB1 in lethal ehrlichiosis suggests a role in toxic shock. Studies of primary bone marrow-derived macrophages infected by highly avirulent or mildly avirulent ehrlichiae reveal divergent M1 and M2 macrophage polarization that links with generation of pathogenic CD8 T cells, neutrophils, and excessive inflammation or with strong expansion of protective Th1 and NKT cells, resolution of inflammation and clearance of infection, respectively.

OASL1-Mediated Inhibition of Type I IFN Reduces Influenza A Infection-Induced Airway Inflammation by Regulating ILC2s.

PurposeThree observations drove this study. First, 2′-5′-oligoadenylate synthetase-like protein (OASL) is a negative regulator of type I interferon (IFN). Second, type I IFN plays a central role during virus infections and the pathogenesis of various diseases, including asthma. Third, influenza A virus (IAV) causes non-eosinophilic asthma. To evaluate the potential relationships between OASL, type I IFN, and pulmonary innate immune cells in IAV-induced acute airway inflammation by using Oasl1-/- mice.MethodsAsthma was induced in wild-type (WT) and Oasl1-/- mice with IAV or ovalbumin (OVA). Airway hyperreactivity (AHR) and immune cell infiltration in the bronchoalveolar lavage (BAL) fluids were measured. The immune cells in the lungs were analyzed by flow cytometry. To investigate the ability of type I IFN to shape the response of lung type 2 innate lymphoid cells (ILC2s), IFN-α was treated intratracheally. Plasmacytoid dendritic cells (pDCs) sorted from bone marrow and ILC2s sorted from lungs of naive mice were co-cultured with/without interferon-alpha receptor subunit 1 (IFNAR-1)-blocking antibodies.ResultsIn the IAV-induced asthma model, Oasl1-/- mice developed greater AHR and immune cell infiltration in the BAL fluids than WT mice. This was not observed in OVA-induced asthma, a standard model of allergen-induced asthma. The lungs of infected Oasl1-/- mice also had elevated DC numbers and Ifna expression and depressed IAV-induced ILC2 responses, namely, proliferation and type 2 cytokine and amphiregulin production. Intratracheal administration of type I IFN in naïve mice suppressed lung ILC2 production of type 2 cytokines and amphiregulin. Co-culture of ILC2s with pDCs showed that pDCs inhibit the function of ILC2s by secreting type I IFN.ConclusionsOASL1 may impede the IAV-induced acute airway inflammation that drives AHR by inhibiting IAV-induced type I IFN production from lung DCs, thereby preserving the functions of lung ILC2s, including their amphiregulin production.

BRD9 regulates interferon-stimulated genes during macrophage activation via cooperation with BET protein BRD4

Macrophages induce a number of inflammatory response genes in response to stimulation with microbial ligands. In response to endotoxin Lipid A, a gene-activation cascade of primary followed by secondary-response genes is induced. Epigenetic state is an important regulator of the kinetics, specificity, and mechanism of gene activation of these two classes. In particular, SWI/SNF chromatin-remodeling complexes are required for the induction of secondary-response genes, but not primary-response genes, which generally exhibit open chromatin. Here, we show that a recently discovered variant of the SWI/SNF complex, the noncanonical BAF complex (ncBAF), regulates secondary-response genes in the interferon (IFN) response pathway. Inhibition of bromodomain-containing protein 9 (BRD9), a subunit of the ncBAF complex, with BRD9 bromodomain inhibitors (BRD9i) or a degrader (dBRD9) led to reduction in a number of interferon-stimulated genes (ISGs) following stimulation with endotoxin lipid A. BRD9-dependent genes overlapped highly with a subset of genes differentially regulated by BET protein inhibition with JQ1 following endotoxin stimulation. We find that the BET protein BRD4 is cobound with BRD9 in unstimulated macrophages and corecruited upon stimulation to ISG promoters along with STAT1, STAT2, and IRF9, components of the ISGF3 complex activated downstream of IFN-alpha receptor stimulation. In the presence of BRD9i or dBRD9, STAT1-, STAT2-, and IRF9-binding is reduced, in some cases with reduced binding of BRD4. These results demonstrate a specific role for BRD9 and the ncBAF complex in ISG activation and identify an activity for BRD9 inhibitors and degraders in dampening endotoxin- and IFN-dependent gene expression.

Association study of promoter polymorphisms of interferon alpha and beta receptor subunit 1 (IFNAR1) gene and therapeutic response to interferon-beta in patients with multiple sclerosis.

Multiple sclerosis (MS) is an autoimmune disease described by inflammatory neuronal losses and resultant failures. The disease could abate by interferon-beta (IFN-β) therapy in MS patients. However, the drug response productivity is changeable between patients, and the accurate mechanism of action of the IFN-β is not obvious. The present study aims to investigate the role of interferon alpha and beta receptor subunit 1 (IFNAR1) promoter polymorphisms towards IFN-β treatment response in MS patients. The subjects herein were separated into either responder (n = 57) or non-responder (n = 43) groups according to IFN-β treatment and Expanded Disability Status Scale score. The Sanger sequencing method was used for genotyping. Among nearly 64 Single Nucleotide Polymorphisms (SNPs), we found a significant association between the rs2850015 polymorphism and the responders and non-responders to IFN-β treatment in the recessive model of inheritance (P = 0.02). The results also revealed a significant change in the two groups of responders and non-responders to the treatment for rs36158718 as an Insertion/Deletion (INDEL) (P = 0.02). Moreover, bioinformatic analyses predicted a remarkable role for both rs2850015 and rs36158718 related to the changes of binding affinity of transcription factors and alterations in their alleles. The present study results suggest that the genetic heterogeneity in the promoter region of IFNAR1 could affect the response to IFN-β. However, further studies with a larger sample size are needed to further demonstrate this relationship.

Novel drug for COVID-19: Virafin a therapeutic breakthrough

Background: Pegylated Interferon Alpha-2b is synthesized via the use of PEG (polyethylene glycol). It is formulated via attaching a 12kDa mono methoxy polyethylene glycol moiety to the recombinant human Interferon Alpha-2b protein. Pegylated Interferon Alpha-2b acts via binding to the (JAK/STAT) Janus kinase signal transducer and activator of the transcription interferon receptor. The Pegylated Interferon Alpha-2b /Virafin binds to the (IFNAR1/2) alpha interferon receptor 1 and 2. Pegylated Interferon Alpha-2b better known as Virafin has been approved by the Drugs Controller General of India. This therapeutic agent is currently under a multicentric phase 3 trial with very promising results being reported. A single subcutaneous dose of Virafin has been shown to decrease the need for oxygen therapy in patients. This reduction in the need for oxygen therapy is a vital factor needed to provide relief to the Indian medical system in light of the recent oxygen shortages faced due to India’s worst wave of COVID-19 cases since the onset of the global pandemic. Pegylated Interferon Alpha-2b/ Virafin confers enhanced viral clearance and bolsters the immune response to induce a quicker recovery in patients with mild to moderate symptoms.
 Conclusion: It is of paramount importance that further research on Virafin is undertaken as it can hinder the progression of COVID-19, reduce pressure on the inundated health systems, and save countless lives.

Naringin prevents cyclophosphamide-induced hepatotoxicity in rats by attenuating oxidative stress, fibrosis, and inflammation

Cyclophosphamide (CYCP), a synthetic alkylating antineoplastic, disrupts both cancerous and non-cancerous cells to cause cancer regression and multi organotoxicity respectively. CYCP-induced hepatotoxicity is rare but possible. Evidence has shown that naringin has several beneficial potentials against oxidative stress, inflammation, and fibrosis. This study examined the chemoprotective potentials of naringin on exited radical scavenging, hepatic integrity, oxidative stress, fibrosis, and inflammation in CYCP-mediated hepatotoxicity. Rats were pre-treated orally by gavage for fourteen consecutive days with three doses (50, 100, and 200 mg/kg) of naringin before single CYCP (200 mg/kg, i.p.) administration. Subsequently, the rats were euthanized; blood and liver were removed, and assessed for serum and hepatic enzymes, oxidative stress, inflammation, and gene expression dynamics. Naringin concentrations required for 50% scavenging hydroxyl radical and 2,2′-azinobis-(3-ethylbenzothiazoline-6-sulphonic acid) radical cation were 0.32 mg/mL and 0.39 mg/mL, respectively. Pretreatment with naringin significantly (p < 0.05) abolish CYCP-induced changes in the activities of serum and hepatic ALT, AST, GGT, ALP, and LDH. Pretreatment with naringin remarkably (p < 0.05) reversed CYCP-mediated increases in hepatic levels of malondialdehyde, hydroperoxide, and nitric oxide; reverse CYCP-induced decreases in the hepatic glutathione levels, activities of catalase, glutathione peroxidase, and glutathione reductase; and also attenuated CYCP-induced upregulation of expression of hepatic chemokine (C-C motif) ligand 2 (CCL2), interferon alpha1 (IFN-α1), interleukine-1β, interleukine-1 receptor, and transforming growth factor beta 1 (TGF-β1). Taken together, different doses of naringin can prevent CYCP-induced oxidants generation, hepatocytes dysfunctions, oxidative stress as well as inflammatory perturbations in rats when pre-administered for as few as 14 days.

Alcohol Consumption Accumulation of Monocyte Derived Macrophages in Female Mice Liver Is Interferon Alpha Receptor Dependent

Monocytes develop in the bone marrow from the hematopoietic stem cells and represent heterogeneous phagocyte cells in the circulation. In homeostatic and inflammatory conditions, after recruitment into tissues, monocytes differentiate into macrophages and dendritic cells. Alcohol use causes about 3.3 million worldwide deaths per year, which is about 5.9% of all deaths. In the United States and Europe, alcohol use disorders represent the fifth leading cause of death. Females are more susceptible to alcoholic liver injury in both humans and mice. Strikingly, we still do not know how much of this difference in tissue injury is due to the differential effect of alcohol and its toxic metabolites on a) parenchymal or resident cells and/or b) immune response to alcohol. Therefore, we used a model of chronic alcohol exposure in mice to investigate the dynamics of monocytes, an innate immune cell type showed to be critical in alcoholic liver injury, by using immunophenotypic characterization. Our data reveal a sex-dimorphism of alcohol response of hepatic monocytes in female mice that is interferon receptor alpha dependent. This dimorphism could shed light on potential cellular mechanism(s) to explain the susceptibility of females to alcoholic immunopathogenesis and suggests an additional targetable pathway for alcoholic liver injury in females.

Abstract PR-007: Targeting FASN improves type I interferon responses in irradiated glioblastoma

Abstract Glioblastoma (GBM) is a devastating primary brain cancer with a median survival of 11-15 months. Radiotherapy (RT), the standard of care for GBM, increases adjuvanticity of cancer cells, by at least generating cancer cell intrinsic type I interferon (IFN-I) responses to jumpstart antitumor immunity. However, RT can also exacerbate potent immune inhibitory mechanisms to facilitate immune evasion. Among which, metabolic reprogramming of irradiated GBM represents an emerging mechanism of immune resistance. Notably, increased de novo lipid synthesis by the fatty acid synthase (FASN) is a hallmark of GBM that is emerging as a mechanisms of therapy resistance and immune escape. Because FASN was found to impair IFN-I in antiviral immunity, we hypothesize that FASN is preventing RT-induced IFN-I responses to promote GBM survival and evade immune recognition. To test this hypothesis, we first defined RT-induced metabolic changes by seahorse assay and metabolomics in the GL261 murine GBM model. We observed an increase in mitochondrial respiration, glycolysis and in lipid metabolism-related pathways in 10 gray (Gy) irradiated GL261 cells. Consistent with this last observation, we found upregulation of FASN expression by western blot and intracellular neutral lipid accumulation by BODIPY staining and transmission electron microscopy (TEM) in irradiated GL261 cells (10 Gy). Confirming the role of FASN, RT-induced accumulation of neutral lipids was reverted when GL261 cells were incubated with a pharmacologic FASN inhibitor. Next, to ask whether FASN was impairing IFN-I in irradiated GBM, GL261 cells were engineered to express an inducible shRNA silencing FASN (GL261shFASN) or its non-silencing control (GL261shNS). As expected, irradiation of GL261shNS cells enhanced the secretion of IFN-I cytokines. Strikingly, we found that FASN knockdown spontaneously induced IFN-beta and CXCL10 secretion, and this effect was even more pronounced when cells were irradiated. Finally, GL261shNS and GL261shFASN cells were orthotopically implanted in mice and IFN-I signaling was blocked using an antibody against the interferon alpha/beta receptor (IFNAR). FASN knockdown alone was sufficient to increase the median survival of mice bearing GL261shFASN tumors by 51 days vs. 35 days for GL261shNS tumors. Moreover, 33% of mice bearing GL261 tumors unable to express FASN remained tumor free for up to 100 days, while none of the GL261shNS-tumor bearing animal survived. This significant survival prolongation of mice bearing GL261shFASN tumor was completely abrogated when IFNAR was blocked. Overall, our findings suggest that FASN-mediated lipogenesis prevents RT-induced cancer cell intrinsic IFN-I to promote GBM survival. Consequently, it is possible that FASN acts as an immuno-metabolic checkpoint capable to regulate the immune system upon metabolic cues generated by RT. Citation Format: Mara De Martino, Camille Daviaud, Claire Vanpouille-Box. Targeting FASN improves type I interferon responses in irradiated glioblastoma [abstract]. In: Proceedings of the AACR Virtual Special Conference on Radiation Science and Medicine; 2021 Mar 2-3. Philadelphia (PA): AACR; Clin Cancer Res 2021;27(8_Suppl):Abstract nr PR-007.

Association between type I interferon receptor 1 single nucleotide polymorphisms and susceptibility to tuberculosis

Objective To investigate the relationship between single nucleotide polymorphism (SNP) of type I interferon receptor 1 (IFNAR1) and susceptibility to tuberculosis in the Han population in southern China. Methods A case-control study was used, with 1 533 patients with active tuberculosis (including 1 432 tuberculosis and 101 extrapulmonary tuberculosis) as the case group, and 1 445 healthy people as the control group. Interferon (IFN) gene rs72552343, rs2834191, rs1012334, rs17875752, rs2843710, rs1041868 genotypes were detected by MassARRAY time-of flight mass spectrometry. The differences in the frequency of SNP alleles between the two groups were analyzed, and the differences in the frequency of rs72552343 TCC / Del locus in patients with pulmonary tuberculosis and extrapulgative tuberculosis were further compared. Results MassARRAY time-of-flight mass spectrometry can effectively detect the genotypes of six SNP loci. Among the 6 SNP loci, it was found that only the rs72552343 TCC/Del locus allele frequency was significantly different between the active tuberculosis group and the control group, and the rs72552343 TCC/Del locus TCC allele frequency was significantly increased ( OR =0.46; 95% CI =0.31 -0.70; P =0.000 2), the frequency of the other five SNP alleles was not significantly different between the two groups. In addition, the frequency of TCC alleles at rs72552343 TCC / Del loci in patients with extrapulmonary tuberculosis was not significantly different from that in patients with tuberculosis ( OR =0.88; 95% CI =0.20-3.74; P =0.86). Conclusion The IFN gene rs72552343 TCC / Del site SNP is related to susceptibility to tuberculosis. The TCC allele is a susceptibility gene for tuberculosis, but individuals carrying the TCC allele have the same risk of active pulmonary tuberculosis as those with extrapulmonary tuberculosis. 摘要：目的 探究Ⅰ型干扰素受体 1 (typeⅠinterferon receptor 1, IFNAR1) 单核苷酸多态性 (single nucleotide poly⁃ morphism, SNP) 与结核病易感性的关系。 方法 应用病例-对照研究方法, 以深圳市第三人民医院 1 533 例活动性结核 病患者 (包括 1 432 例肺结核、101 例肺外结核) 作为病例组, 1 445 例健康人群作为对照组。运用 MassARRAY 飞行时间 质 谱 技 术 , 通 过 检 测 Ⅰ 型 干 扰 素 (interferon, IFN) 基 因 rs72552343、rs2834191、rs1012334、rs17875752、rs2843710、rs1041868 位点基因型, 分析两组间 SNP 等位基因频率差异, 同时进一步比较肺结核与肺外结核病患者 rs72552343 TCC/Del 位点等位基因频率差异。 结果 用 MassARRAY 飞行时间质谱技术可以有效地检测 6 个 SNP 位点基因型。在 6 个 SNP 位点中, 发现仅 rs72552343 TCC/Del 位点等位基因频率在活动性结核组和对照组中差异有统计学意义 ( P 0.05)。另外, 发现肺外结核病患者 rs72552343 TCC/Del 位点 TCC 等位基因的频率与肺结核病患者差异无统计学意 义 ( OR =0.88; 95% CI =0.20~3.74; P =0.86)。 结论 IFN 基因 rs72552343 TCC/Del 位点 SNP 与结核易感性相关, 其 TCC 等位基因为结核易感基因, 但携带 TCC 等位基因的个体患活 动性肺结核与肺外结核的风险一致。

The impact of TLR7 agonist R848 treatment on mast cell phenotype and activity

Bearing in mind that mast cell contribution to viral clearance is still not fully understood, in this study, we evaluated the effect of Toll-like receptor (TLR)7 viral single-stranded ribonucleic acid (ssRNA) mimic ligand, namely resiquimod (R)848, on mast cell phenotype and activity. We demonstrated that rat peritoneal mast cells exhibit surface and intracellular expression of ssRNA-specific TLR7 molecule, and that mimic ligand switches the self-expression of this receptor. We also detected other proteins associated with the cellular antiviral response: interferon-alpha receptor 1 (IFNAR1), interferon-gamma receptor 1 (IFNGR1), and major histocompatibility complex I (MHC I). Moreover, we showed that R848 caused the decrease of all molecule’s expression after prolonged incubation. Interestingly, we found that R848 induced the increase of high-affinity IgE receptor (FcεRI) expression. Finally, we documented that TLR7 ligand-stimulated mast cells synthesize/release interferon (IFN)-α and -β, tumor necrosis factor (TNF), and chemokines CCL3, CXCL8, as well as pro-inflammatory lipid mediators. Our findings confirm that mast cells may respond to TLR7 ligand by altering their phenotype and synthesizing mediators and could serve as active participants in the antiviral immune response.

Monophosphoryl lipid A-induced activation of plasmacytoid dendritic cells enhances the anti-cancer effects of anti-PD-L1 antibodies.

Monophosphoryl lipid A (MPLA) is a toll-like receptor 4 ligand that promotes immune activation in mice and humans, without undesired inflammation. Immunotherapy by the combining immune checkpoint blockade and MPLA has shown promising anti-cancer effects in both mice and humans. In this study, we explored how MPLA enhanced the anti-cancer effects of anti-PD-L1 antibodies (Abs). Anti-cancer immunity induced by the combination of anti-PD-L1 Abs and MPLA failed in CD4 and CD8 cell-depleted mice. Moreover, the combination treatment of anti-PD-L1 Abs and MPLA synergistically enhanced the activation of plasmacytoid dendritic cells (pDCs) in the mouse in vivo, while conventional DCs were not. In addition, mice treated with anti-PD-L1 Abs and MPLA were not protected from B16 melanoma by blockade of interferon-alpha receptor (IFNAR). The combination of anti-PD-L1 Abs and MPLA also promoted human peripheral blood pDC activation and induced IFN-α-dependent T cell activation. Therefore, these results demonstrate that MPLA enhances anti-PD-L1 Ab-mediated anti-cancer immunity through the activation and IFN-α production of pDCs.