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Influenza A virus ( IAV)-induced severe disease is characterized by infected lung epithelia, robust inflammatory responses and acute lung injury. Since type I interferon ( IFNαβ) and type III interferon ( IFNλ) are potent antiviral cytokines with immunomodulatory potential, we assessed their efficacy as IAV treatments. IFNλ treatment of IAV-infected Mx1-positive mice lowered viral load and protected from disease. IFNα treatment also restricted IAV replication but exacerbated disease. IFNα treatment increased pulmonary proinflammatory cytokine secretion, innate cell recruitment and epithelial cell death, unlike IFNλ-treatment. IFNλ lacked the direct stimulatory activity of IFNα on immune cells. In epithelia, both IFNs induced antiviral genes but no inflammatory cytokines. Similarly, human airway epithelia responded to both IFNα and IFNλ by induction of antiviral genes but not of cytokines, while hPBMCs responded only to IFNα. The restriction of both IFNλ responsiveness and productive IAV replication to pulmonary epithelia allows IFNλ to limit IAV spread through antiviral gene induction in relevant cells without overstimulating the immune system and driving immunopathology. We propose IFNλ as a non-inflammatory and hence superior treatment option for human IAV infection. [ABSTRACT FROM AUTHOR]

Hepatitis C virus (HCV) infection is one of the major causes of liver cancer worldwide and every year 3–4 million people become infected. Whereas some infected people eradicate the virus spontaneously, almost 85% do not and instead develop a chronic infection. Recently, it was shown that people with a functional IFNL4 gene have a lower chance of clearing the virus spontaneously or in response to treatment than people with a non-functional IFNL4 gene. Why it is a disadvantage to have a functional IFNL4 gene during HCV infection is currently not known although a causal relationship between the activity of the IFN λ 4 protein and poor HCV clearance has been demonstrated. IFN λ 4 belongs to the type III IFNs together with IFN λ 1, -2, and -3. However, it differs from the others not only by its low sequence similarity but also by its impaired secretion. This impairment is not due to a weak signal peptide, as swapping the signal peptides between IFN λ 3 and -4 had no effect on secretion. When we purified IFN λ 3 and -4, we found that IFN λ 4 is far more difficult to refold in vitro than IFN λ 3 suggesting that the poor secretion of IFN λ 4 could be due to an inherent problem of folding the protein. Because such a problem could also mean that IFN λ 4 could be far unstable than the other type III IFNs, we decided to compare the stability of recombinant IFN λ 3 and IFN λ 4. Our results demonstrate that IFN λ 4 like IFN λ 3 is surprisingly stable once it has folded properly. [ABSTRACT FROM AUTHOR]

Interferon lambda 4 (IFNL4) is a recently identified enigmatic member of the interferon lambda family. Genetic data suggest that the IFNL4 gene acts in a pro-viral and anti-inflammatory manner in patients. However, the protein is in vitro indistinguishable from the other members of the interferon lambda family. We have investigated the gene regulation of IFNL4 in detail and found that it differs radically from that of canonical antiviral interferons. Being induced by viral infection is a defining characteristic of interferons, but viral infection or overexpression of members of the interferon regulatory factor (IRF) family of transcription factors only leads to a minute induction of IFNL4. This behavior is evolutionarily conserved and can be reversed by inserting a functional IRF3 binding site into the IFNL4 promoter. Thus, the regulation of the IFNL4 gene is radically different and might explain some of the atypical phenotypes associated with the IFNL4 gene in humans. [ABSTRACT FROM AUTHOR]

Type I and type III interferons (IFNs) can promote adaptive immune responses in mice and improve vaccine-induced resistance to viral infections. The adjuvant effect of type III IFN (IFN-λ) specifically boosts mucosal immunity by an indirect mechanism, involving IFN-λ-induced production of thymic stromal lymphopoietin (TSLP), a cytokine that activates immune cells. To date, it remained unclear whether the previously described adjuvant effect of type I IFN (IFN-α/β) would also depend on TSLP and whether type I IFN stimulates different antibody subtypes. Here, we show that after infection with a live attenuated influenza virus, mice lacking functional type I IFN receptors failed to produce normal amounts of virus-specific IgG2c and IgA antibodies. In contrast, mice lacking functional IFN-λ receptors contained normal levels of virus-specific IgG2c but had reduced IgG1 and IgA antibody levels. When applied together with protein antigen, IFN-α stimulated the production of antigen-specific IgA and IgG2c to a greater extent than IgG1, irrespective of whether the mice expressed functional TSLP receptors and irrespective of whether the vaccine was applied by the intranasal or the intraperitoneal route. Taken together, these results demonstrate that the adjuvant activities of type I and type III IFNs are mechanistically distinct. IMPORTANCE Interferons can shape antiviral immune responses, but it is not well understood how they influence vaccine efficacy. We find that type I IFN preferentially promotes the production of antigen-specific IgG2c and IgA antibodies after infection with a live attenuated influenza virus or after immunization with influenza subunit vaccines. In contrast, type III IFN specifically enhances influenza virus-specific IgG1 and IgA production. The adjuvant effect of type I IFN was not dependent on TSLP, which is essential for the adjuvant effect of type III IFN. Type I IFN boosted vaccine-induced antibody production after immunization by the intranasal or the intraperitoneal route, whereas type III IFN exhibited its adjuvant activity only when the vaccine was delivered by the mucosal route. Our findings demonstrate that type I and type III IFNs trigger distinct pathways to enhance the efficacy of vaccines. This knowledge might be used to design more efficient vaccines against infectious diseases. [ABSTRACT FROM AUTHOR]

IFNL4 , a recently discovered member of the interferon λ family (IFN λ s or type III IFNs), is a pseudogene in a significant fraction of the human population. A single nucleotide polymorphism located in IFNL4 , which determines the ability to express IFN λ 4, has been correlated with poor spontaneous and treatment-induced clearance of hepatitis C virus (HCV) infections. We show that IFN λ 4 is an active type III IFN that induces a typical subset of ISGs, signals through the classical type III IFN receptor complex and is antiviral against HCV and coronaviruses. However, its secretion is impaired and this impairment is caused by a yet unknown molecular determinant, but appears to be partially caused by a weak signal peptide and inefficient N-linked glycosylation. This glycosylation is not required for antiviral activity and secretion of IFN λ 4, but seems to improve its processing. The impaired secretion of IFN λ 4 appears to be a recently acquired feature of primates. A single amino acid substitution in IFN λ 4 changing a proline at position 70 to a serine (P70S) alters its activity. We demonstrate that the IFN λ 4-S70 variant has a significantly lower antiviral activity compared to IFN λ 4-P70. Our subsequent genetic study on a cohort of patients infected with HCV shows that individuals, who encode IFN λ 4-S70, display lower hepatic ISG expression, better treatment response rates and better spontaneous clearance rates than patients encoding IFN λ 4-P70. This study provides important evidence supporting a role for active IFN λ 4 as the driver of high hepatic ISG expression as well as the cause of poor HCV clearance. [ABSTRACT FROM AUTHOR]

Inhibition of tumor necrosis factor alpha (TNFalpha) is a favorable way of treating several important diseases such as rheumatoid arthritis, Crohn disease, and psoriasis. Therefore, an extensive range of TNFalpha inhibitory proteins, most of them based upon an antibody scaffold, has been developed and used with variable success as therapeutics. We have developed a novel technology platform using C-type lectins as a vehicle for the creation of novel trimeric therapeutic proteins with increased avidity and unique properties as compared with current protein therapeutics. We chose human TNFalpha as a test target to validate this new technology because of the extensive experience available with protein-based TNFalpha antagonists. Here, we present a novel and highly specific TNFalpha antagonist developed using this technology. Furthermore, we have solved the three-dimensional structure of the antagonist-TNFalpha complex by x-ray crystallography, and this structure is presented here. The structure has given us a unique insight into how the selection procedure works at a molecular level. Surprisingly little change is observed in the C-type lectin-like domain structure outside of the randomized regions, whereas a substantial change is observed within the randomized loops. Thus, the overall integrity of the C-type lectin-like domain is maintained, whereas specificity and binding affinity are changed by the introduction of a number of specific contacts with TNFalpha.

Interferon-lambda (IFN-lambda) is an antiviral cytokine that signals through a distinct receptor complex, composed of the IFN-lambdaR1 and interleukin-10R2 (IL-10R2) receptor chains. We have determined the crystal structure of human IFN-lambda3 and characterized the interaction with its receptor complex through structure-based site-directed mutagenesis. The ability of IFN-lambda3 mutants to signal was determined by measuring the antiviral activity and induced STAT2 phosphorylation. In conclusion, our data show that, although IFN-lambda is functionally an interferon, it is clearly structurally related to members of the IL-10 family. In particular, we found an interesting similarity between IFN-lambda and IL-22, and we suggest that IFN-lambda and IL-22 possess parallel functions, protecting epithelial tissue against viral and bacterial infections, respectively.