No abstract available

Interferon has been used to treat chronic viral hepatitis and several malignancies. However, it may cause various neuropsychiatric adverse effects including parkinsonism. We report a rare case of interferon alpha-2a therapy-related parkinsonism in a 67-year-old man with metastatic papillary renal cell carcinoma and our experience of using Tc-99m-TRODAT-1 single photon emission computed tomography (SPECT) as a tool for evaluation of parkinsonism. Physicians should be alert to the possibility of interferon alpha-2a-related parkinsonism.

Interferon-alpha (IFN- α ) has immunoregulatory functions in autoimmune inflammatory diseases. The goal of this study was to determine occurrence and clinical consequences of IFN- α in neuromyelitis optica (NMO) patients. Thirty-six NMO and 41 multiple sclerosis (MS) patients from a population-based retrospective case series were included. Expanded Disability Status Scale (EDSS) score and MRI findings determined disease activity. Linear regression was used to assess the effects of the level of IFN- α on disability (EDSS). IFN- α was determined by sensitive ELISA assays. IFN- α was detectable in sera from 9/36 NMO patients, significantly more often than in the MS group (2/41) (P = 0.0197). A higher frequency of IFN- α was observed in NMO patients with acute relapse compared to NMO patients in remission (P < 0.001) and compared to the MS patients with relapse (P = 0.010). In NMO patients, the levels of IFN- α were significantly associated with EDSS (P = 0.0062). It may be concluded that IFN- α was detectable in a subgroup of NMO patients. Association of IFN- α levels with clinical disease activity and severity suggests a role for IFN- α in disease perpetuation and may provide a plausible explanation for a negative effect of IFN-1 treatment in NMO patients.

Bacillus anthracis, the etiologic agent of inhalational anthrax, is a facultative intracellular pathogen. Despite appropriate antimicrobial therapy, the mortality from inhalational anthrax approaches 45%, underscoring the need for better adjuvant therapies. The variable latency between exposure and development of disease suggests an important role for the host's innate immune response. Type I and Type II Interferons (IFN) are prominent members of the host innate immune response and are required for control of intracellular pathogens. We have previously described a protective role for exogenous Type I and Type II IFNs in attenuating intracellular B.anthracis germination and macrophage cell death in vitro.

Interferon (IFN) exhibits a potent antiviral activity in vitro and plays a major role in the early defense against viruses. Like IFN, the proinflammatory chemokine, interleukin (IL)-8, is induced by viruses and appears in circulation during viral infections. In an in vitro cytopathic effect assay for IFN, we found that IL-8 can inhibit IFN-alpha activity in a dose-dependent manner. This action was reversed by specific monoclonal antibodies to IL-8. The chemokine was able to attenuate the IFN-mediated inhibition of viral replication as determined by measuring infectious virus yield. IL-8 also diminished the ability of IFN to inhibit an early stage of viral replication since IL-8 attenuated the inhibition of the formation of viral proteins. It appeared that IL-8 interfered with a late rather than an early step of IFN-mediated pathway such as early gene expression. The IL-8 inhibitory action on IFN-alpha antiviral activity was associated with reduced 2',5'-A oligoadenylate synthetase activity, a pathway well correlative with the anti- encephalomyocarditis virus action of IFN-alpha. Understanding pathways that antagonize IFN action may lead to novel approaches to potentiate endogenous and therapeutic IFN.

The negative regulators in the interferon (IFN) signaling pathway inhibit intrahepatic immune response, resulting in suboptimal therapeutic response to IFNα treatment in chronic hepatitis B (CHB) patients. Identifying the key negative factors and elucidating the regulating mechanism are essential for improving anti-HBV (hepatitis B virus) efficacy of IFNα. From the Gene Expression Omnibus (GEO) database, we downloaded and analyzed gene expression profiles of CHB patients with different responses to IFNα (GSE54747), and found that innate immune status was associated with the IFNα-based therapeutic response in CHB patients. Through PCR array, we found higher baseline level of IFN-induced transmembrane protein 2 (IFITM2) mRNA and lower baseline level of IFNα mRNA in peripheral blood mononuclear cells (PBMCs) of CHB patients with suboptimal response to IFNα treatment. Increased IFITM2 protein was also found in the serum of IFNα nonresponsive patients. With further experiments, we found that overexpressing IFITM2 in Huh7 cells suppressed endogenous IFNα synthesis by inhibiting phosphorylation of extracellular signal-regulated kinase (ERK), TANK-binding kinase 1 (TBK1), and interferon regulatory factor 3 (IRF3); knocking out IFITM2 enhanced activation of the endogenous IFNα synthesis pathway, exhibiting better inhibition on HBV replication. We also found that IFITM2 protein was shuttled by exosomes to dendritic cells (DCs), the main source of endogenous IFNα. Exosome-mediated transport of IFITM2 inhibited synthesis of endogenous IFNα in DCs whereas the inhibitory effect was abolished when IFITM2 was knocked out. Furthermore, we demonstrated that both palmitoylation inhibitor and mutation on 70/71 sites of IFITM2 protein influenced its incorporation into exosomes. Mutated IFITM2 protein increased the effect of IFNα against HBV. Conclusion: Exosome-mediated transport of IFITM2 to DCs inhibits IFNα pathway activation and blocks anti-HBV efficacy of exogenous IFNα. The findings provide an explanation to the suboptimal response of CHB patients to IFNα treatment.

Pegylated interferon alpha 2 (a or b) plus ribavirin is the most effective treatment of chronic hepatitis C but a large proportion of patients do not respond to therapy. So, it is interesting to improve the treatment efficacy. Interferon alpha is a type I interferon composed of 12 different subtypes. Each subtype signals by the Jak-Stat pathway but modulations in the antiviral activity was previously described.

A new form of circuitry for gene regulation has been identified in which RNAs can crosstalk by competing for shared microRNAs (miRNAs). Such competing endogenous RNAs (ceRNAs) form a network via shared miRNA response elements (MREs) to antagonize miRNA function. We previously reported natural antisense RNA (AS) as an important modulator of interferon-α1 (IFN-α1) mRNA levels by promoting IFN-α1 mRNA stability. We show that IFN-α1 AS forms a ceRNA network with specific IFN-α AS (IFN-α7/-α8/-α10/-α14) and mRNA (IFN-α8/-α10/-α14/-α17) subtypes from the IFN-α gene (IFNA) family to antagonize miRNA-1270 (miR-1270), thereby modulating IFN-α1 mRNA levels. Bioinformatic analysis demonstrated that IFN-α1 AS harbors multiple miR-1270 MREs (MRE-1270s), whose presence was substantiated by miR-1270 overexpression and transfection of antimiR-1270. The antimiR-1270, complementary to the miR-1270 seed region, revealed that IFN-α1 AS likely shares the MRE-1270 with IFN-α1 mRNA and specific IFN-α AS and mRNA subtypes. Subsequent bioinformatic analysis for MRE-1270s showed that IFN-α1 AS and other RNA subtypes shared the 6-mer MRE-1270 site. Further MRE-mapping demonstrated that the total number of MRE-1270s in IFN-α1 AS accounted for approximately 30 % of the miR-1270 population. AntimiR-1270 transfection also caused specific de-repression of five cellular mRNAs, including that of CAPRIN1. These results suggest that IFN-α1 AS, together with specific IFN-α AS and mRNA subtypes, as well as the five cellular mRNAs, participate as competing molecules in the ceRNA network against miR-1270. This coordinated regulatory architecture suggests a vital function for the innate immune system in maintaining precise physiological type I IFN levels via post-transcriptional regulatory mechanisms.

Interferon alpha (IFNα) is a cytokine approved for the treatment of several types of cancer. However, the modest effect on overall survival and the high toxicity associated with the treatment has reduced the clinical use of this cytokine. In this study, we have developed a tumor model that reproduces this clinical setting. A high dose of an adeno-associated virus encoding IFNα (AAV-IFNα) was able to eradicate a liver metastases model of colon cancer but induced lethal pancytopenia. On the other hand, a safe dose of AAV-IFNα was not able to eliminate the liver metastases of colon cancer. In this IFNα-resistant tumor model, administration of an adeno-associated vector encoding apolipoprotein A-1 fused to IFNα was able to fully eradicate the tumor in 43% of mice without toxicity. This antitumor effect was limited by suboptimal long-term CD8+ T cell activation and the expansion of T regulatory cells. In contrast, IFNα upregulated suppressor molecules such as PD-1 and interleukin 10 on CD8+ T lymphocytes. In conclusion, we show that apolipoprotein A-1 fused to IFNα is a novel antitumor drug that differs from IFNα in the modulation of suppressor mechanisms of the immune response. These differential properties pave the way for rational combinations with other immunomodulatory drugs.

Depression is one of the major side effects of interferon alpha (IFN-α) treatment, but the molecular mechanism underlying IFN-α-induced depression remains unclear. Several studies have shown that the serotonin receptors 5-HTR1b and 5-HTR4 play key roles in the anti-depression effects associated with p11 (S100A10). We investigated the effects of IFN-α on the regulation of p11, 5-HTR1b and 5-HTR4 in mice and human neuroblastoma cells (SH-sy5y). We found that intraperitoneal injection with IFN-α in Balb/c mice resulted in an increased immobility in FST and TST, and potently lowered the protein levels of p11, 5-HTR1b and 5-HTR4 in the hippocampus or cingulate gyrus. IFN-α significantly down-regulated the protein levels of p11, 5-HTR1b and 5-HTR4 in SH-sy5y cells, in a time- and dose-dependent manner. Our study revealed that over-expression of p11 could prevent the IFN-α-induced down-regulation of 5-HTR1b and 5-HTR4. The results indicated that IFN-α treatment resulted in p11 down-regulation, which subsequently decreased 5-HTR1b and 5-HTR4 in vitro or in vivo. Our findings suggested that p11 might be a potential regulator on 5-HTR1b and 5-HTR4 as well as a predictor of or a therapeutic target for IFN-α-induced depression.

This paper describes the cloning of full length marsupial type I interferon (IFN) genes and their flanking regions using a genome walking approach and PCR primers based on previously isolated partial DNA sequences. We confirm that the two major classes of Tammar Wallaby type I IFN genes are homologous with the eutherian IFN-alpha and IFN-beta gene families. The wallaby IFN genes share a number of conserved features with their eutherian counterparts, including codons for cysteines at equivalent positions, implying similar secondary structures for the encoded proteins, and promoter regions with conserved putative regulatory motifs. Moreover, the wallaby genes have AT-rich elements in their flanking sequence corresponding to the mRNA 3'-untranslated regions, also implying that, as in eutherian mammals, rapid mRNA degradation plays a role in regulating expression of these genes. The complex nature of the type I IFN gene families in viviparous mammals (eutherians and marsupials) may reflect their recruitment into nonimmunological processes and this concept is discussed.

Interferon alpha (IFN-alpha) not only plays a key role in innate host immunity against infections but also is involved in the cellular functions of the central nervous system (CNS). In this study, we examined the impact of morphine on IFN-alpha expression in human neuronal cells (NT2-N). Similar to human immune cells, NT2-N cells also expressed IFN-alpha at both mRNA and protein levels. IFN-alpha expression in NT2-N cells, however, was inhibited by morphine. Naltrexone antagonized the inhibitory effect of morphine on IFN-alpha expression in NT2-N cells. The specific mu opioid receptor antagonist, Cys2, Tyr3, Arg5, Pen7-amide (CTAP), also blocked the morphine action on intracellular IFN-alpha expression. Investigation of the mechanisms involved in the morphine action showed that although morphine had little effect on the expression of key IFN regulatory factors (IRFs), morphine inhibited IFN-alpha promoter activation and suppressed the expression and phosphorylation of signal transducer and activator of transcription 1 (STAT1) in the neuronal cells. These findings provide direct in vitro evidence that opioids may impair neuronal cell-mediated innate protection in the CNS.

Hepatitis B virus (HBV) infection remains a major health problem worldwide, and the current antiviral therapy, including nucleoside analogs, cannot achieve life-long cure, and clarification of antiviral host immunity is necessary for eradication. Here, we found that a clathrin-binding membrane protein epsin3 (EPN3) negatively regulates the expression of HBV RNA. EPN3 expression was induced by transfection of an HBV replicon plasmid, and reduced HBV-RNA level in hepatic cell lines and murine livers hydrodynamically injected with the HBV replicon plasmid. Viral RNA reduction by EPN3 was dependent on transcription, and independent from epsilon structure of viral RNA. Viral RNA reduction by overexpression of p53 or IFN-α treatment, was attenuated by knockdown of EPN3, suggesting its role downstream of IFN-α and p53. Taken together, this study demonstrates the anti-HBV role of EPN3. The mechanism how it decreases HBV transcription is discussed.

Interferon (IFN) alpha is one of the central agents in immunotherapy for renal cell carcinoma (RCC). It acts by binding to the IFN-alpha receptor (IFNAR). We previously reported that increased tumor expression of IFNAR2 mRNA was associated with the metastatic potential and progression of RCC, as well as with a poor response of metastatic RCC to IFN-alpha therapy. This study investigated the influence of serum IFNAR2 in RCC patients.

Interferon (IFN) genes are among the earliest transcriptional responses to virus infection of mammalian cells. Although the regulation of the IFNbeta gene has been well characterized, the induction of the large family of IFNalpha genes has remained obscure. We report that the IFNalpha genes can be divided into two groups: an immediate-early response gene (IFNalpha4) which is induced rapidly and without the need for ongoing protein synthesis; and a set of genes that display delayed induction, consisting of at least IFNalpha2, 5, 6 and 8, which are induced more slowly and require cellular protein synthesis. One protein that must be synthesized for induction of the delayed gene set is IFN itself, presumably IFNalpha4 or IFNbeta, which stimulates the Jak-Stat pathway through the IFN receptor, resulting in activation of the transcription factor interferon-stimulated gene factor 3 (ISGF3). Among the IFN-stimulated genes induced through this positive feedback loop is the IFN regulatory factor (IRF) protein, IRF7. Induction of IRF7 protein in response to IFN and its subsequent activation by phosphorylation in response to virus-specific signals, involving two C-terminal serine residues, are required for induction of the delayed IFNalpha gene set.

Before an infection can be completely established, the host immediately turns on the innate immune system through activating the interferon (IFN)-mediated antiviral pathway. Kaposi's sarcoma-associated herpesvirus (KSHV) utilizes a unique antagonistic mechanism of type I IFN-mediated host antiviral immunity by incorporating four viral interferon regulatory factors (vIRF1-4). Herein, we characterized novel immune evasion strategies of vIRF4 to inhibit the IRF7-mediated IFN-α production. KSHV vIRF4 specifically interacts with IRF7, resulting in inhibition of IRF7 dimerization and ultimately suppresses IRF7-mediated activation of type I IFN. These results suggest that each of the KSHV vIRFs, including vIRF4, subvert IFN-mediated anti-viral response via different mechanisms. Therefore, it is indicated that KSHV vIRFs are indeed a crucial immunomodulatory component of their life cycles.

Tightly interlinked feedback regulators control the dynamics of intracellular responses elicited by the activation of signal transduction pathways. Interferon alpha (IFNα) orchestrates antiviral responses in hepatocytes, yet mechanisms that define pathway sensitization in response to prestimulation with different IFNα doses remained unresolved. We establish, based on quantitative measurements obtained for the hepatoma cell line Huh7.5, an ordinary differential equation model for IFNα signal transduction that comprises the feedback regulators STAT1, STAT2, IRF9, USP18, SOCS1, SOCS3, and IRF2. The model-based analysis shows that, mediated by the signaling proteins STAT2 and IRF9, prestimulation with a low IFNα dose hypersensitizes the pathway. In contrast, prestimulation with a high dose of IFNα leads to a dose-dependent desensitization, mediated by the negative regulators USP18 and SOCS1 that act at the receptor. The analysis of basal protein abundance in primary human hepatocytes reveals high heterogeneity in patient-specific amounts of STAT1, STAT2, IRF9, and USP18. The mathematical modeling approach shows that the basal amount of USP18 determines patient-specific pathway desensitization, while the abundance of STAT2 predicts the patient-specific IFNα signal response.

Aggregation is a common phenomenon in the field of protein therapeutics and can lead to function loss or immunogenic patient responses. Two strategies are currently used to reduce aggregation: (1) finding a suitable formulation, which is labor-intensive and requires large protein quantities, or (2) engineering the protein, which requires extensive knowledge about the protein aggregation pathway. We present a biophysical characterization of the oligomerization and aggregation processes by Interferon alpha-2a (IFNα-2a), a protein drug with antiviral and immunomodulatory properties. This study combines experimental high throughput screening with detailed investigations by small-angle X-ray scattering and analytical ultracentrifugation. Metropolis Monte Carlo simulations are used to gain insight into the underlying intermolecular interactions. IFNα-2a forms soluble oligomers that are controlled by a fast pH and concentration-dependent equilibrium. Close to the isoelectric point of 6, IFNα-2a forms insoluble aggregates which can be prevented by adding salt. We show that monomer attraction is driven mainly by molecular anisotropic dipole-dipole interactions that increase with increasing pH. Repulsion is due to monopole-monopole interactions and depends on the charge of IFNα-2a. The study highlights how combining multiple methods helps to systematically dissect the molecular mechanisms driving oligomer formation and to design ultimately efficient strategies for preventing detrimental protein aggregation.

Interferon alpha (IFNα) is used for the treatment of hepatitis C infection and whilst efficacious it is associated with multiple adverse events including reduced leukocyte, erythrocyte, and platelet counts, fatigue, and depression. These events are most likely caused by systemic exposure to interferon. We therefore hypothesise that targeting the therapeutic directly to the intended site of action in the liver would reduce exposure in blood and peripheral tissue and hence improve the safety and tolerability of IFNα therapy. We genetically fused IFN to a domain antibody (dAb) specific to a hepatocyte restricted antigen, asialoglycoprotein receptor (ASGPR). Our results show that the murine IFNα2 homolog (mIFNα2) fused to an ASGPR specific dAb, termed DOM26h-196-61, could be expressed in mammalian tissue culture systems and retains the desirable biophysical properties and activity of both fusion partners when measured in vitro. Furthermore a clear increase in in vivo targeting of the liver by mIFNα2-ASGPR dAb fusion protein, compared to that observed with either unfused mIFNα2 or mIFNα2 fused to an isotype control dAb VHD2 (which does not bind ASGPR) was demonstrated using microSPECT imaging. We suggest that these findings may be applicable in the development of a liver-targeted human IFN molecule with improved safety and patient compliance in comparison to the current standard of care, which could ultimately be used as a treatment for human hepatitis virus infections.

Ligand-specific negative regulation of cytokine-induced signaling relies on down regulation of the cytokine receptors. Down regulation of the IFNAR1 sub-unit of the Type I interferon (IFN) receptor proceeds via lysosomal receptor proteolysis, which is triggered by ubiquitination that depends on IFNAR1 serine phosphorylation. While IFN-inducible phosphorylation, ubiquitination, and degradation requires the catalytic activity of the Tyk2 Janus kinase, here we found the ligand- and Tyk2-independent pathway that promotes IFNAR1 phosphorylation, ubiquitination, and degradation when IFNAR1 is expressed at high levels. A major cellular kinase activity that is responsible for IFNAR1 phosphorylation in vitro does not depend on either ligand or Tyk2 activity. Inhibition of ligand-independent IFNAR1 degradation suppresses cell proliferation. We discuss the signaling events that might lead to ubiquitination and degradation of IFNAR1 via ligand-dependent and independent pathways and their potential physiologic significance.