Mapping Receptor Binding Sites in Interleukin (IL)-1 Receptor Antagonist and IL-1β by Site-directed Mutagenesis

Evans, Ron J.; Bray, Jeffery P.; Childs, John D.; Vigers, Guy; Brandhuber, Barbara J.; Skalicky, Jack J.; Thompson, Robert C.; Eisenberg, Stephen P.

doi:10.1074/jbc.270.19.11477

Cited by 102 publications

(63 citation statements)

References 27 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…Alternatively, the binding of IL-18 mutants to the cells may not be significantly different from WT binding, but signal transduction may be greater. The dissociation of binding affinities with biological activities of IL-1␤ mutants is well described (16,17). In the case of IL-1 mutants, receptor affinities are either unchanged or somewhat lower, but biological activity is decreased by 100-fold.…”

Section: Discussionmentioning

confidence: 85%

Site-specific mutations in the mature form of human IL-18 with enhanced biological activity and decreased neutralization by IL-18 binding protein

Kim

Azam

Yoon

et al. 2001

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

IL-18 can be considered a proinflammatory cytokine mediating disease as well as an immunostimulatory cytokine that is important for host defense against infection and cancer. The high-affinity, constitutively expressed, and circulating IL-18 binding protein (IL-18BP), which competes with cell surface receptors for IL-18 and neutralizes IL-18 activity, may act as a natural antiinflammatory as well as immunosuppressive molecule. In the present studies, the IL-18 precursor caspase-1 cleavage site was changed to a factor Xa site, and, after expression in Escherichia coli, mature IL-18 was generated by factor Xa cleavage. Mature IL-18 generated by factor Xa cleavage was fully active. Single point mutations in the mature IL-18 peptide were made, and the biological activities of the wild-type (WT) IL-18 were compared with those of the mutants.

show abstract

Section: Discussionmentioning

confidence: 85%

Site-specific mutations in the mature form of human IL-18 with enhanced biological activity and decreased neutralization by IL-18 binding protein

Kim

Azam

Yoon

et al. 2001

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

show abstract

“…IL-1β and IL-1Ra engage the same primary receptor, IL-1R1, but only IL-1β recruits IL-1RAcP and signals (21). Mutagenesis and structural studies have identified two disparate sites on IL-1β that participate in IL-1R1 binding, designated sites A and B, only one of which-site A-is formed with IL-1Ra (22)(23)(24). Site A of the ligand binds to the first two Ig domains, D1 and D2, whereas site B binds to the third Ig domain, D3 (23,24) (Fig.…”

Section: Resultsmentioning

confidence: 99%

Design of a superior cytokine antagonist for topical ophthalmic use

Hou¹,

Townson²,

Kovalchin³

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

IL-1 is a key inflammatory and immune mediator in many diseases, including dry-eye disease, and its inhibition is clinically efficacious in rheumatoid arthritis and cryopyrin-associated periodic syndromes. To treat ocular surface disease with a topical biotherapeutic, the uniqueness of the site necessitates consideration of the agent's size, target location, binding kinetics, and thermal stability. Here we chimerized two IL-1 receptor ligands, IL-1β and IL-1Ra, to create an optimized receptor antagonist, EBI-005, for topical ocular administration. EBI-005 binds its target, IL-1R1, 85-fold more tightly than IL-1Ra, and this increase translates to an ∼100-fold increase in potency in vivo. EBI-005 preserves the affinity bias of IL-1Ra for IL-1R1 over the decoy receptor (IL-1R2), and, surprisingly, is also more thermally stable than either parental molecule. This rationally designed antagonist represents a unique approach to therapeutic design that can potentially be exploited for other β-trefoil family proteins in the IL-1 and FGF families.T he IL-1 cytokines (IL-1α and IL-1β) are master mediators of inflammatory responses (1). IL-1β also regulates immune function through its role in T helper 17 (Th17) cell differentiation and maintenance (2, 3). IL-1 action has been implicated in numerous human diseases, including rheumatoid arthritis, MuckleWells syndrome, gout, type 2 diabetes, and stroke (4). Several natural mechanisms directly oppose the actions of IL-1, including a soluble and cell surface decoy receptor (IL-1R2), a natural antagonist (IL-1Ra), and a soluble signaling receptor (IL-1R1) (5). Therapeutics that block IL-1 based on these mechanisms have been developed (6-8).Recently, a nonoptimized formulation of anakinra (methionyl-IL-1Ra; Kineret) was shown to provide clinical benefit in dry-eye disease (DED) (9). Moderate to severe DED is a chronic inflammatory condition of the corneal surface that results in pain, discomfort, and epitheliopathy (as measured by fluorescein staining). Inability to maintain a proper tear film over the cornea (owing to a variety of etiologies) results in desiccating stress, which drives an inflammatory cascade (10, 11). IL-1 plays a central role in the initiation and maintenance of this cascade, as well as in the pain mediated by the corneal neural plexus. IL-1α and IL-1β protein are elevated in the lacrimal gland, tears, and the ocular surface in all forms of dry-eye disease (12), and their mRNA is increased in both humans and in rodent disease models (13,14). Genetic ablation of IL-1R1, the primary receptor for IL-1α and IL-1β, can block the development of corneal staining in a Sjögren syndrome corneal epitheliopathy model (15), and topically administered anakinra can improve surface epithliopathy in a mouse dry-eye model (14). IL-1β is essential for Th17 cell differentiation and maintenance, and Th17 cells are likely the main effector cells that induce epithelial damage (2, 3). Genetic and pharmacologic studies have shown that IL-1β mediates, and IL-1Ra blocks, normal, inflamm...

show abstract

“…The biological activities of IL-1β are tightly controlled by different natural inhibitors, including membrane-bound and soluble IL-1 receptors, and IL-1 receptor antagonist (IL-1Ra) [1]. IL-1Ra binds to IL-1 receptors without inducing intracellular signals and acts as a natural competitive IL-1 inhibitor [2]. Genetic IL-1Ra deficiency leads to excessive IL-1 signaling and exaggerated inflammatory responses in several animal models and in humans [1,[3][4][5][6][7][8][9].…”

Section: Introductionmentioning

confidence: 99%

Mice deficient in hepatocyte‐specific IL‐1Ra show delayed resolution of concanavalin A‐induced hepatitis

et al. 2012

View full text Add to dashboard Cite

Interleukin-1 receptor antagonist (IL-1Ra) is a specific IL-1 inhibitor that possesses antiinflammatory activities. Several studies in human and mouse suggested a protective role for IL-1Ra in liver inflammation, and we previously demonstrated that hepatocytes produce high levels of IL-1Ra in response to inflammatory challenge in vitro and in vivo. In the present study, we investigated the production and the biological function of hepatocytederived IL-1Ra in concanavalin A (ConA)-induced hepatitis in mice. We show that the injured liver produces large amounts of IL-1Ra and that secreted and intracellular IL-1Ra isoforms are produced with different kinetics during the course of hepatitis. By using hepatocyte-specific IL-1Ra-deficient mice (IL-1Ra H ), we demonstrate that hepatocytes represent the major cellular source of local IL-1Ra. Most interestingly, hepatic necrosis and inflammation were increased in IL-1Ra H as compared with wild-type mice during the late phase of the disease, leading to a delayed resolution of hepatitis in IL-1Ra H mice. In conclusion, our results show that the local production of IL-1Ra by hepatocytes contributes to the resolution of hepatitis.Keywords: Cytokine r IL-1 inhibitor r Inflammation r Liver r Necrosis Supporting Information available online IntroductionInterleukin-1β (IL-1β) is a prototypic pro-inflammatory cytokine playing an important role in acute and chronic inflammatory processes and in some autoimmune diseases [1]. The biological activities of IL-1β are tightly controlled by different natural inhibitors, including membrane-bound and soluble IL-1 receptors, Correspondence: Prof. Cem Gabay e-mail: cem.gabay@hcuge.ch and IL-1 receptor antagonist (IL-1Ra) [1]. IL-1Ra binds to IL-1 receptors without inducing intracellular signals and acts as a natural competitive IL-1 inhibitor [2]. Genetic IL-1Ra deficiency leads to excessive IL-1 signaling and exaggerated inflammatory responses in several animal models and in humans [1,[3][4][5][6][7][8][9].IL-1β has been proposed as an important mediator in the pathophysiology of hepatic diseases. Indeed, a significant increase of circulating IL-1β levels has been reported in patients with fulminant hepatic failure and chronic liver diseases [10][11][12][13]. Increased serum levels of IL-1β were also observed in an experimental model of immune-mediated hepatitis induced by concanavalin-A (ConA) injection in mice [14]. By using the same experimental model,www.eji-journal.eu Eur. J. Immunol. 2012. 42: 1294-1303 Innate immunity 1295 mice lacking functional caspase-1, an enzyme involved in IL-1β and IL-18 processing, were resistant to hepatitis [15]. Interestingly, Sekiyama et al. [12] observed elevated IL-1Ra levels in the circulation of patients with fulminant hepatic failure and acute hepatitis, and found that the IL-1Ra/IL-1β ratio was significantly increased in patients who survived. In liver biopsy specimens from patients with chronic hepatitis, IL-1Ra/IL-1β messenger ribonucleic acid (mRNA) ratio were inversely correlated with disease se...

show abstract

Mapping Receptor Binding Sites in Interleukin (IL)-1 Receptor Antagonist and IL-1β by Site-directed Mutagenesis

Cited by 102 publications

References 27 publications

Site-specific mutations in the mature form of human IL-18 with enhanced biological activity and decreased neutralization by IL-18 binding protein

Site-specific mutations in the mature form of human IL-18 with enhanced biological activity and decreased neutralization by IL-18 binding protein

Design of a superior cytokine antagonist for topical ophthalmic use

Mice deficient in hepatocyte‐specific IL‐1Ra show delayed resolution of concanavalin A‐induced hepatitis

Contact Info

Product

Resources

About