Immune Complex-mediated Inflammation Research Articles

Bilateral renal artery stenosis in pediatric patient with Post-Streptococcal Glomerulonephritis in children: A case report

The immune-complex mediated inflammation known as post-streptococcal glomerulonephritis (PSGN) was once thought to be one of the most common causes of acute nephritis in children. In this case, we present a 12-year-old girl experiencing symptoms of cough phlegm, congested breath, change of urine color, decreased urine production, as well swelling in the area face and legs after experiencing fever, pain swallowing, cough, and runny nose 3 weeks before admission. On physical examination, the pressure blood was 130/80 mmHg. In addition, the palpebral and extremity were edema bilateral, hyperemia of the tonsil and pharynx, enlarged tonsils T2/T3, and detritus were found. Laboratory and Imaging Tests: hematology; leukocytes 12,930, albumin 2.4 g/dl, on urinalysis, obtained urine cloudy, brownish, leukocyturia, hematuria, and proteinuria, there was also an increase in ASTO titers, abdominal ultrasound found bilateral renal artery stenosis and nephritis bilateral acute. The patient in this case presented with bilateral renal artery stenosis, which is a rare finding in PSGN cases. Management of PSGN include bed rest, low salt diet, fluid balance, as well supportive treatment with IVFD D5 ½ NS 15 drops/min (micro), Antibiotics injection of Ceftriaxone 1 gram/12 hours on the first day and continued with oral Erythromycin 4x500mg, injection prednisolone 3x1, injection Furosemide 1x1 amp, oral spironolactone and sublingual nifedipine, and hypoalbuminemia correction. Prognosis in patients with the given GNAPS governance optimally will give good results.

Confocal Real-Time Analysis of Cutaneous Platelet Recruitment during Immune Complex‒Mediated Inflammation

Platelets preserve vascular integrity during immune complex‒mediated skin inflammation by preventing neutrophil-provoked hemorrhage. However, the single-cell dynamics of this hemostatic process have never been studied in real-time. To monitor the onset of thrombocytopenia-associated hemorrhages and analyze platelet recruitment, we developed a confocal microscopy‒based video-imaging platform for the dorsal skinfold chamber in living mice. For ultrastructural analysis of recruited platelets, we correlated our imaging approach with serial block-face scanning electron microscopy. We found that bleeding events were transient and occurred preferentially at vascular sites, which were repeatedly penetrated by extravasating neutrophils. Hemorrhage only resumed when previously affected sites were again breached by yet another neutrophil. In non-thrombocytopenic mice, we observed that neutrophil extravasation provoked the recruitment of single platelets to the vessel wall, which required platelet immunoreceptor tyrosine-based activation motif receptors glycoprotein VI and C-type-lectin-like receptor 2. Recruited platelets were found to spread across the endothelial barrier and some even across the basement membrane while retaining their granules. Thus, by visualizing the spatiotemporal dynamics of thrombocytopenia-associated bleeding and platelet recruitment on a single-cell level and in real-time, we provide further insights into how platelets preserve vascular integrity during immune complex‒mediated skin inflammation.

A Comprehensive Review Study on Glomerulonephritis Associated With Post-streptococcal Infection.

Post-streptococcal glomerulonephritis (PSGN) is an immune-complex mediated inflammation that used to be considered one of the commonest causes of acute nephritis amongst children. PSGN is characterized by the proliferation of cellular elements called nephritogenic M type as a result of an immunologic mechanism following an infection of the skin (impetigo) or throat (pharyngitis) caused by nephritogenic strains of group A beta-hemolytic streptococci, a gram-positive bacteria that enters the body across pores in the skin or mucus epithelia and is responsible for more than 500,000 deaths annually due to multiple subsequence diseases such as rheumatic heart disease, rheumatic fever, PSGN, and other invasive infections. After the infection, the formation of an immune complex of antigen-antibody and complement system will take place and will deposit in the glomeruli where the injury occurs and leads to inflammation. The manifestations of PSGN can be explained by nephritic syndrome manifestation. PSGN is diagnosed by laboratory tests like microscopy and urinalysis. The imaging studies in PSGN could be used to assess the possible complications of PSGN such as pulmonary congestion and chronic kidney disease. The management of PSGN is symptomatic. If PSGN is not treated, the patient may develop chronic kidney disease. The main way to prevent PSGN is to treat group A streptococcal (GAS) infections by giving good coverage of antibiotic therapy to a patient who has primary GAS infections to prevent the development of the complication.

Therapeutic inhibition of spleen tyrosine kinase in inflammatory macrophages using PLGA nanoparticles for the treatment of non-alcoholic steatohepatitis

Non-alcoholic steatohepatitis (NASH) is the leading cause of cirrhosis worldwide and the most rapidly growing indication for liver transplantation. Macrophages are the important cellular component in the inflammatory milieu in NASH. Inflammatory and pro-fibrotic mediators produced by macrophages causes significant tissue injury in many inflammatory diseases. Therefore, inhibition of the inflammatory macrophages would be a promising approach to attenuate NASH. In this study, we studied the implication of SYK pathway in NASH, and investigated PLGA nanoparticles-based delivery of SYK pathway inhibitor as an effective and promising therapeutic approach for the treatment of NASH. We found positive correlation between SYK expression with the pathogenesis of NASH and alcoholic hepatitis in patients. Importantly, SYK expression was significantly induced in M1-differentiated inflammatory macrophages. To inhibit SYK pathway specifically, we used a small-molecule inhibitor R406 that blocks Fc-receptor signaling pathway and reduces immune complex-mediated inflammation. R406 dose-dependently inhibited nitric-oxide release and M1-specific markers in M1-differentiated macrophages. Thereafter, we synthesized PLGA nanoparticles to deliver R406 to increase the drug pharmacokinetics for the efficient treatment of NASH. We investigated the therapeutic efficacy of R406-PLGA in-vitro in differentiated macrophages, and in-vivo in Methionine-Choline-deficient (MCD)-diet induced NASH mouse model. R406-PLGA inhibited M1-specific differentiation markers in RAW and bone-marrow-derived macrophages. In-vivo, R406 and more strongly R406-PLGA ameliorated fibrosis, inflammation and steatosis in mice. R406 and more significantly R406-PLGA reduced ALT, AST, cholesterol and triglyceride plasma levels. These results suggest that delivery of SYK inhibitor using PLGA nanoparticles can be a potential therapeutic approach for the treatment of Non-alcoholic steatohepatitis.

The Impact of Vitamin D on the Immunopathophysiology, Disease Activity, and Extra-Musculoskeletal Manifestations of Systemic Lupus Erythematosus.

Over the past two decades it has been increasingly recognized that vitamin D, aside from its crucial involvement in calcium and phosphate homeostasis and the dynamics of the musculoskeletal system, exerts its influential impact on the immune system. The mechanistic roles that vitamin D plays regarding immune activation for combating infection, as well as pathologically and mediating autoimmune conditions, have been progressively unraveled. In vitro and in vivo models have demonstrated that the action of vitamin D on various immunocytes is not unidirectional. Rather, how vitamin D affects immunocyte functions depends on the context of the immune response, in the way that its suppressive or stimulatory action offers physiologically appropriate and immunologically advantageous outcomes. In this review, the relationship between various aspects of vitamin D, starting from its adequacy in circulation to its immunological functions, as well as its autoimmune conditions, in particular systemic lupus erythematosus (SLE), a prototype autoimmune condition characterized by immune-complex mediated inflammation, will be discussed. Concurring with other groups of investigators, our group found that vitamin D deficiency is highly prevalent in patients with SLE. Furthermore, the circulating vitamin D levels appear to be correlated with a higher disease activity of SLE as well as extra-musculoskeletal complications of SLE such as fatigue, cardiovascular risk, and cognitive impairment.

Quercitrin ameliorates the development of systemic lupus erythematosus-like disease in a chronic graft-versus-host murine model.

Systemic lupus erythematosus (SLE) is a serious disorder of immune regulation characterized by overproduction of autoantibodies, lupus nephritis, CD4+ T cell aberrant activation, and immune complex-mediated inflammation. The chronic graft vs. host disease (cGVHD) mouse model is a well-established model of SLE. Quercitrin is a natural compound found in Tartary buckwheat with a potential anti-inflammatory effect that is used to treat heart and vascular conditions. In our previous study, we determined that quercitrin is an immunosuppressant with beneficial effects in mouse models of immune diseases. We hypothesized that quercitrin could prevent lupus nephritis in the cGVHD mouse model by decreasing the production of autoantibodies and inflammatory cytokines, and reducing immune cell activation. cGVHD was induced by injecting DBA/2 spleen cells into the tail vein of BDF1 mice. The cGVHD mice exhibited significant proteinuria, which is a marker of nephritis. Quercitrin decreased the number of serum antibodies, CD4+ T cell activation, as well as the expression levels of T-bet, GATA-3, and selected cytokines. Moreover, quercitrin treatment decreased the expression of inflammatory genes and cytokines in the kidney, as well as in peritoneal macrophages. In addition, quercitrin inhibited LPS-induced cytokines as well as the phosphorylation of ERK, p38 MAPK, and JNK in Raw264.7 cells. Overall, quercitrin ameliorated the symptoms of lupus nephritis in the cGVHD mouse model, which may be due to the inhibition of CD4 T cell activation and anti-inflammatory effects on macrophages.

Functional state of the ovaries, uterus, thymus and lymph nodes in conditions of experimental immune complex-mediated inflammation and administration of the zero valent iron nanoparticles substance in mice

Functional state of the ovaries, uterus, thymus and lymph nodes in conditions of experimental immune complex-mediated inflammation and administration of the zero valent iron nanoparticles substance in mice

The many faces of Mac-1 in autoimmune disease.

Mac-1 (CD11b/CD18) is a β2 integrin classically regarded as a pro-inflammatory molecule because of its ability to promote phagocyte cytotoxic functions and enhance the function of several effector molecules such as FcγR, uPAR, and CD14. Nevertheless, recent reports have revealed that Mac-1 also plays significant immunoregulatory roles, and genetic variants in ITGAM, the gene that encodes CD11b, confer risk for the autoimmune disease systemic lupus erythematosus (SLE). This has renewed interest in the physiological roles of this integrin and raised new questions on how its seemingly opposing biological functions may be regulated. Here, we provide an overview of the CD18 integrins and how their activation may be regulated as this may shed light on how the opposing roles of Mac-1 may be elicited. We then discuss studies that exemplify Mac-1's pro-inflammatory versus regulatory roles particularly in the context of IgG immune complex-mediated inflammation. This includes a detailed examination of molecular mechanisms that could explain the risk-conferring effect of rs1143679, a single nucleotide non-synonymous Mac-1 polymorphism associated with SLE.

Single platelets seal neutrophil-induced vascular breaches via GPVI during immune-complex–mediated inflammation in mice

AbstractPlatelets protect vascular integrity during inflammation. Recent evidence suggests that this action is independent of thrombus formation and requires the engagement of glycoprotein VI (GPVI), but it remains unclear how platelets prevent inflammatory bleeding. We investigated whether platelets and GPVI act primarily by preventing detrimental effects of neutrophils using models of immune complex (IC)-mediated inflammation in mice immunodepleted in platelets and/or neutrophils or deficient in GPVI. Depletion of neutrophils prevented bleeding in thrombocytopenic and GPVI−/− mice during IC-mediated dermatitis. GPVI deficiency did not modify neutrophil recruitment, which was reduced by thrombocytopenia. Neutrophil cytotoxic activities were reduced in thrombocytopenic and GPVI−/− mice during IC-mediated inflammation. Intravital microscopy revealed that in this setting, intravascular binding sites for platelets were exposed by neutrophils, and GPVI supported the recruitment of individual platelets to these spots. Furthermore, the platelet secretory response accompanying IC-mediated inflammation was partly mediated by GPVI, and blocking of GPVI signaling impaired the vasculoprotective action of platelets. Together, our results show that GPVI plays a dual role in inflammation by enhancing neutrophil-damaging activities while supporting the activation and hemostatic adhesion of single platelets to neutrophil-induced vascular breaches.

Immune complex disease in guinea pig lungs: elicitation with pigeon serum.

Immune complex- and T cell-mediated reactions to organic antigens appear to contribute to the pathogenesis of hypersensitivity pneumonitis in humans. Because pigeon serum is one of the reagents used by clinicians to diagnose this disease, we assessed its potential to elicit immune complex-mediated pulmonary inflammation in guinea pigs. Animals were immunized with different concentrations of pigeon serum protein emulsified in complete Freund's adjuvant, and serums were collected at 4-day intervals after the booster injection. The largest amounts of tissue-fixing (IgG1) and complement-fixing (IgG2) antibodies to pigeon serum were detected in guinea pigs immunized with 1.0 mg of pigeon serum protein 20 to 24 days after the secondary immunization. Therefore, the responses of these animals and of recipients of serum from these animals to aerosol challenge with either homologous (pigeon serum) or heterologous (bovine gamma globulin) immunogen was investigated. Actively and passively immunized guinea pigs developed pulmonary inflammation only after exposure to aerosolized pigeon serum. However, lesions were not observed in the lungs of complement-deficient recipients of immune serum that had inhaled homologous immunogen. These observations suggest that such pigeon serum-elicited pulmonary inflammation in guinea pigs is a manifestation of a complement-dependent, humoral-immune mechanism of pathogenesis and thus is consistent with an immune complex disease.

Correction to “R406, an Orally Available Spleen Tyrosine Kinase Inhibitor Blocks Fc Receptor Signaling and Reduces Immune Complex-Mediated Inflammation”

Correction to “R406, an Orally Available Spleen Tyrosine Kinase Inhibitor Blocks Fc Receptor Signaling and Reduces Immune Complex-Mediated Inflammation”

Metabolomic investigation of Arthus reaction in a rat model using proton nuclear magnetic resonance (1H NMR) spectroscopy and rapid resolution liquid chromatography (RRLC)

Arthus reaction (AR), a type of unconventional immune complex-mediated inflammation, is likely accompanied by alterations in circulating metabolites. Here, a proton nuclear magnetic resonance ((1)H NMR) spectroscopy method coupled with a rapid resolution liquid chromatography (RRLC) method was developed to evaluate the systemic metabolic consequences of AR and characterize metabolic aberrations. Serum and urine samples from AR rats and normal controls were compared to determine whether there were significant alterations associated with AR. The partial least squares discriminant analysis (PLS-DA) models of metabolomic results demonstrated good intergroup separations between AR rats and normal controls. Multivariate statistical analysis revealed significant alterations in the levels of 34 metabolites, which were termed as the disease-associated biomarkers. Differential metabolites identified from the metabolomic analysis suggested that AR caused dysfunctions of kidney and liver accompanied with changes in widespread metabolic pathways including the tricarboxylic acid (TCA) cycle, gut microbiota metabolism, lipids and cell membranes metabolism, glucose metabolism, fatty acid β-oxidation, amino acids metabolism and ketogenesis. This study assessed and provided important metabolomic variations in serum and urine associated with AR and, therefore, demonstrated metabolomics as a powerful approach for the complete elucidation of the underlying pathophysiologic mechanisms of AR.

Interleukin-33 Primes Mast Cells for Activation by IgG Immune Complexes

Mast cells (MCs) are heterogeneous cells whose phenotype is modulated by signals received from the local microenvironment. Recent studies have identified the mesenchymal-derived cytokine IL-33 as a potent direct activator of MCs, as well as regulator of their effector phenotype, and have implicated this activity in the ability of mast cells to contribute to murine experimental arthritis. We explored the hypothesis that IL-33 enables participation of synovial MCs in murine K/BxN arthritis by promoting their activation by IgG immune complexes. Compared to wild-type (WT) control mice, transgenic animals lacking the IL-33 receptor ST2 exhibited impaired MC-dependent immune complex-induced vascular permeability (flare) and attenuated K/BxN arthritis. Whereas participation of MCs in this model is mediated by the activating IgG receptor FcγRIII, we pre-incubated bone marrow-derived MCs with IL-33 and found not only direct induction of cytokine release but also a marked increase in FcγRIII-driven production of critical arthritogenic mediators including IL-1β and CXCL2. This “priming” effect was associated with mRNA accumulation rather than altered expression of Fcγ receptors, could be mimicked by co-culture of WT but not ST2−/− MCs with synovial fibroblasts, and was blocked by antibodies against IL-33. In turn, WT but not ST2−/− MCs augmented fibroblast expression of IL-33, forming a positive feedback circuit. Together, these findings confirm a novel role for IL-33 as an amplifier of IgG immune complex-mediated inflammation and identify a potential MC-fibroblast amplification loop dependent on IL-33 and ST2.

Exacerbation of collagen induced arthritis by Fcγ receptor targeted collagen peptide due to enhanced inflammatory chemokine and cytokine production

Antibodies specific for bovine type II collagen (CII) and Fcγ receptors play a major role in collagen-induced arthritis (CIA), a mouse model of rheumatoid arthritis (RA). Our aim was to clarify the mechanism of immune complex-mediated inflammation and modulation of the disease. CII pre-immunized DBA/1 mice were intravenously boosted with extravidin coupled biotinylated monomeric CII-peptide epitope (ARGLTGRPGDA) and its complexes with biotinylated FcγRII/III specific single chain Fv (scFv) fragment. Disease scores were monitored, antibody titers and cytokines were determined by ELISA, and binding of complexes was detected by flow cytometry and immune histochemistry. Cytokine and chemokine secretion was monitored by protein profiler microarray. When intravenously administered into collagen-primed DBA/1 mice, both CII-peptide and its complex with 2.4G2 scFv significantly accelerated CIA and increased the severity of the disease, whereas the monomeric peptide and monomeric 2.4G2 scFv had no effect. FcγRII/III targeted CII-peptide complexes bound to marginal zone macrophages and dendritic cells, and significantly elevated the synthesis of peptide-specific IgG2a. Furthermore, CII-peptide containing complexes augmented the in vivo secretion of cytokines, including IL-10, IL-12, IL-17, IL-23, and chemokines (CXCL13, MIP-1, MIP-2). These data indicate that complexes formed by the CII-peptide epitope aggravate CIA by inducing the secretion of chemokines and the IL-12/23 family of pro-inflammatory cytokines. Taken together, these results suggest that the in vivo emerging immune complexes formed with autoantigen(s) may trigger the IL-12/23 dependent pathways, escalating the inflammation in RA. Thus blockade of these cytokines may be beneficial to downregulate immune complex-induced inflammation in autoimmune arthritis.

Hydrodynamic delivery of plasmid DNA encoding human FcγR-Ig dimers blocks immune-complex mediated inflammation in mice

Therapeutic use and function of recombinant molecules can be studied by the expression of foreign genes in mice. In this study, we have expressed human Fcgamma receptor –Ig fusion molecules (FcγR-Igs) in mice by administering FcγR-Ig plasmid DNAs hydrodynamically and compared their effectiveness to purified molecules in blocking immune-complex (IC) mediated inflammation in mice. The concentration of hydrodynamically expressed FcγR-Igs (CD16AF-Ig, CD32AR-Ig and CD32AH-Ig) reached a maximum of 130 μg/ml of blood within 24 h after plasmid DNA administration. The in vivo half-life of FcγR-Igs was found to be 9-16 days and Western blot analysis showed that the FcγR-Igs were expressed as a homodimer. The hydrodynamically expressed FcγR-Igs blocked 50-80% of IC-mediated inflammation up to 3 days in a reverse passive Arthus reaction model. Comparative analysis with purified molecules showed that hydrodynamically expressed FcγR-Igs are more efficient than purified molecules in blocking IC-mediated inflammation and had a higher half-life. In summary, these results suggest that the administration of a plasmid vector with a FcγR-Ig gene can be used to study the consequences of blocking IC-binding to FcγRs during the development of inflammatory diseases. This approach may have potential therapeutic value in treating IC-mediated inflammatory autoimmune diseases such as lupus, arthritis and autoimmune vasculitis.

CRESCENTIC GLOMERULONEPHRITIS AND LEUCOCYTOCLASTIC VASCULITIS ASSOCIATED WITH ACUTE EBV INFECTION

We report a case of leucocytoclastic vasculitis with crescentic glomerulonephritis in association with acute EBV infection. nep_1440 617..618 A 14 year old male presented to us with history of sudden onset of fever and skin rashes initially over the trunk and gradually involving the entire body including the face for 10 days. He was febrile and breathless with a blood pressure of 110/60 mmHg. He had conjunctival suffusion with a diffuse erythematous maculopapular rash. Systemic examination revealed hepatosplenomegaly with pulmonary congestion. His laboratory investigations revealed thrombocytopenia (87000/cmm) with acute renal failure (creatinine 1.7 mg/dL). Urinalysis showed 2+ proteinuria, haematuria and red blood cell casts with spot urine protein to creatinine ratio of 1.74. His blood and urine culture was negative and 2-D echocardiography was normal. Serum complement 3 and 4 levels were low and vasculitis work-up was negative. Serum for EBV capsid antigen (VCA) was highly positive with increasing titres. He had normal bone marrow studies and skin biopsy suggestive of leucocytoclastic vasculitis. He was treated with i.v. ceftriaxone and other supportive measures but he continued to have high grade fever with worsening azotemia (creatinine 5.2 mg/dL). Percutaneous kidney biopsy revealed severe proliferative glomerulonephritis with cellular crescents in more than 50% of the glomeruli (Fig. 1). He was treated with i.v. methyl prednisolone 500 mg once daily on 3 consecutive days followed by 1 mg/kg day p.o. Following this, his respiratory symptoms subsided, rashes started fading and his urine output improved. His serum creatinine dropped to 0.8 mg/dL with disappearance of proteinuria and haematuria. The finding of leucocytoclastic vasculitis on skin biopsy is suggestive of small-vessel vasculitis. The possible pathophysiological mechanism is either direct invasion of endothelial cells by EBV or by immune complex-mediated inflammation. Renal involvement in EBV infection is predominantly interstitial, 2 overt renal failure is uncommon 3 and crescentic glomerulonephritis has never been reported. Crescentic glomerulonephritis and small-vessel vasculitis in the presence of Ig M VCA antibodies in our patient may have resulted from an intense immune response to the EBV infection. Simultaneous decrease in C3 and C4 indicating classical complement activation along with prompt response to immunosuppressive therapy supports this possibility. Thus, we hypothesize that EBV infection should be included in the differential diagnosis while treating a patient with crescentic glomerulonephritis.

TNF receptor subtype 2 induction by viral dsRNA involves IP-10 and TNF-α in glomerular mesangial inflammation

the ubiquitously important immunoregulator tumor necrosis factor-α (TNF-α) mediates its effects via two distinct receptors, TNFR1 and TNFR2. Although the exact roles of these receptors are not yet completely understood and may differ depending on organ type, they are known to participate in the

Lipocalin-2 is involved in immune-complex mediated inflammation (102.23)

Abstract Lipocalin-2 (Lcn-2) is an innate immune protein whose expression is upregulated by several orders of magnitude in response to inflammation/injury/infection by variety of cells including neutrophils. However, the role of Lcn-2 induction in IC-induced inflammation is largely unknown. Here in, we have investigated the role of Lcn-2 during IC-mediated inflammation using Lcn-2 knock out (Lcn-2KO) and wild type (WT) mice. IC-mediated inflammation upregulated systemic Lcn-2 in WT mice when compared to naïve mice. The neutrophil chemoattractant KC was significantly elevated in WT, but not in Lcn-2KO mice, in response to IC-induced inflammation. Consistently Lcn-2KO mice showed about 50% reduced IC-mediated inflammation when compared to WT mice. Histopathological analysis of skin biopsy showed reduced inflammatory cell infiltration at inflamed site in Lcn-2KO mice relative to WT mice. Administration of recLcn-2 into Lcn-2KO mice resulted in inflammation similar to that of WT mice in response to IC. Lastly, we observed that neutralization of Lcn-2 via neutralizing mAb significantly reduced inflammation in WT mice. These results suggest that engagement of FcγRs expressed on local inflammatory cells with ICs leads to the induction of Lcn-2. Such IC-induced Lcn-2 expression may be essential for infiltration inflammatory cells at the site of inflammation. Our studies show that therapeutic targeting of Lcn-2 may be useful approach for treating IC-mediated inflammatory/autoimmune diseases

Suppression of inflammation in collagen-induced arthritis by administration of recombinant Fcγ receptors (54.17)

Abstract Recently we have shown that immune-complex mediated inflammation can be blocked by recombinant dimeric FcγR-Ig molecules in a mouse Arthus reaction model. In the present study, we have investigated whether administration of FcγR-Ig would inhibit the antibody-mediated hemolytic anemia, thrombocytopenia and arthritis in a mouse model. Systemic Administration of recombinant dimeric low affinity FcγRs such as human CD32A alleles (hCD32AR-Ig and hCD32AH-Ig) did not inhibit or prevent the antibody-induced hemolytic anemia and thrombocytopenia. Interestingly, we observed that after 48 hr of administration both hCD32AR-Ig and hCD32AH-Ig were able to inhibit 27% and 42% of the arthritic inflammation respectively, whereas the combination of hCD32AR-Ig and hCD32AH-Ig was able to inhibit 52% of arthritic inflammation. The severity of arthritis as assessed by inflammation and swelling in each digit and paw was significantly reduced. A single dose of hCD32A-Ig molecules was able to reduce the severity of arthritis up to 4 days, after which the disease returned and reached the maximum severity by day 7. The relapse of arthritis might be due to other mechanisms such as complement pathways or accelerated clearance of administered FcγR-Igs in arthritis mice. In conclusion, soluble dimeric FcγR-Igs could be of potential use in treating antibody-mediated autoimmune diseases such as arthritis but not for autoimmune diseases which affect blood cells such as hemolytic anemia and thrombocytopenia

Specific Inhibition of Syk Suppresses Leukocyte Immune Function and Alleviates Inflammation In Rodent Models of Rheumatoid Arthritis

AbstractAbstract 1727Genetic ablation of Syk in hematopoietic cells blocks various leukocyte immune functions, and protects mice from immune-complex mediated inflammation. These data have helped to identify Syk as an important therapeutic target for immune-mediated diseases. The next step is to test the hypothesis that low level, and specific, pharmacological inhibition of Syk retains the immunomodulatory potential observed with Syk genetic deficiency. With this goal in mind, we provide an update on the development of P505-15, a highly specific and potent small molecule Syk inhibitor which suppresses signaling and activation of primary human and rodent leukocyte immune function. The specificity of P505-15 was tested in a panel of 270 independent purified kinase assays at 300nM. At this concentration, Syk and 8 other kinases were inhibited by ≥ 80%. Subsequent analysis demonstrated a Syk IC50 of 1nM, whereas the next most potently inhibited kinase required an IC50 of 81nM. In a variety of cellular assays we observed potent inhibition of B cell receptor (BCR) induced Syk signaling, but not of Lyn, phorbol 12-myristate 13-acetate (PMA) induced protein kinase C, T cell receptor induced Zap70, or cytokine induced JAK1 (IL6), JAK2 (GM-CSF), or JAK1/3 (IL4) dependent STAT phosphorylation. Consistently, in Ba/F3 cell lines transformed by various kinases, P505-15 only inhibited proliferation of those cells transformed by Syk (IC50 = 0.12μM), and not by Zap70 or JAK family members (IC50 > 6μM). In human whole blood, P505-15 suppressed BCR-induced Syk signaling and cellular activation with IC50's of 0.383μM and 0.362μM, respectively. FceR1-induced basophil degranulation was similarly suppressed with an IC50 of 0.171μM. Importantly, Syk-independent signaling and cellular activation in human whole blood via PMA (B cell assays) or fMLP (basophil degranulation) was unaffected by this compound at 4μM and 1μM, respectively (the highest concentrations tested), again demonstrating its specificity of action. Oral administration of P505-15 in mice led to a reversible inhibition of Syk, with an IC50 of 0.282μM as determined by an ex vivo whole blood BCR stimulation assay. Finally, we tested the immunomodulatory potential of specific Syk inhibition in vivo using rodent models of rheumatoid arthritis. Oral administration of P505-15 resulted in statistically significant and dose-dependent anti-inflammatory activity in both the mouse collagen antibody-induced arthritis and rat collagen induced arthritis models. In each case, anti-inflammatory effects were achieved at sub-micromolar plasma concentrations in which Syk specificity was maintained. These data support the hypothesis that specific Syk inhibition can modulate immune function in vivo, and provide a therapeutic strategy for the treatment of human inflammatory disease by inhibition of this kinase. P505-15 is currently being evaluated in phase I clinical trials. Disclosures:Coffey:Portola Pharmaceuticals: Employment. Francis:Portola Pharmaceuticals: Employment. Inagaki:Portola Pharmaceuticals: Employment. Pak:Portola Pharmaceuticals: Employment. Delaney:Portola Pharmaceuticals: Employment. Betz:Portola Pharmaceuticals: Employment. Jia:Portola Pharmaceuticals: Employment. Xu:Portola Pharmaceuticals: Employment. Bauer:Portola Pharmaceuticals: Employment. Song:Portola Pharmaceuticals: Employment. Pandey:Portola Pharmaceuticals: Employment. Baker:Portola Pharmaceuticals: Employment. Hollenbach:Portola Pharmaceuticals: Employment. Phillips:Portola Pharmaceuticals: Employment. Sinha:Portola Pharmaceuticals: Employment.