C5a Receptor Antagonist Research Articles

Structural study of Ac‐Phe‐[Orn‐Pro‐dCha‐Trp‐Arg], a potent C5a receptor antagonist, by NMR

The cyclic hexapeptide Ac(0)-Phe(1)-[Orn(2)-Pro(3)-dCha(4)-Trp(5)-Arg(6)] (the square brackets denote cyclization) is a potent antagonist against C5a (the a-fragment of complement protein C5) binding to C5a receptor (C5aR) and an excellent candidate to become a therapeutic agent against diseases that involve unregulated activation of the complement system. We present the solution structure determination of this cyclic C5aR peptide antagonist (cC5aR-pa), using nuclear magnetic resonance (NMR) data and restrained molecular dynamics-based simulated annealing in torsion angle space with NMR-derived distance and torsion angle restraints. The calculated NMR ensemble of structures demonstrates the presence of a predominant conformation of a distorted type II' beta-turn in the segment Pro(3)-dCha(4)-Trp(5)-Arg(6). We critically examine the calculated structure with measured NMR parameters, such as nuclear Overhauser enhancement (NOE) connectivity patterns and intensities characteristic of specific structures, (3)J(H(N)-H(alpha)) scalar coupling constants, temperature coefficients for NH groups, and differences between observed chemical shifts and their random coil values. The raw NMR data are consistent with the presence of the type II' beta-turn, but also indicate the presence of conformational inter-conversion. The calculated three-dimensional coordinates for cC5aR-pa will form the basis for further computational studies and for the development of pharmacophore models.

Systemic blockade of complement C5a receptors reduces lipopolysacharride-induced responses in the paraventricular nucleus and the central amygdala

The complement anaphylatoxin C5a is a potent mediator of the innate immune response to infection. Recent evidence also reveals that C5a contributes to central nervous system effects in addition to its well-known peripheral functions. However, it is not known if C5a has a role in the activation of the hypothalamic-pituitary-adrenal (HPA) axis; a critical cascade that exemplifies neuroimmune interactions between the periphery and the brain. In the present study we examined if systemic pre-treatment with a C5a receptor antagonist, PMX53, can affect lipopolysaccharide-induced (LPS; 1 mg/kg, i.p.) activation of the HPA axis in the rat. Using Fos protein as a marker of neuronal activation, we found that systemic administration of PMX53 reduced the LPS-induced activation of paraventricular corticotropin-releasing factor (PVN CRF) and central amygdala cells. However, PMX53 did not alter LPS-induced responses in the bed nucleus of the stria terminalis, nucleus tractus solitarius and ventrolateral medulla. Our findings demonstrate that C5a may have a role in the activation of the HPA axis in response to systemic LPS.

A Protein Toxin from the Sea Anemone Phyllodiscus semoni Targets the Kidney and Causes a Severe Renal Injury with Predominant Glomerular Endothelial Damage

Envenomation by the sea anemone Phyllodiscus semoni causes fulminant dermatitis and, rarely, acute renal failure in humans. Here, we investigated whether the venom extracted from the nematocysts (PsTX-T) was nephrotoxic when administered intravenously in rats and whether PsTX-T induced activation of the complement system. Although small dose of PsTX-T induced acute tubular necrosis in rats resembling pathology seen in patients, kidneys displayed glomerular injury with glomerular endothelial damage, thrombus formation, mesangiolysis, and partial rupture of glomerular basement membrane, accompanied by severe tubular necrosis at 24 hours after administration of 0.03 mg of PsTX-T per animal, similar to the glomerular findings typical of severe hemolytic uremic syndrome. The early stage injury was accompanied by specific PsTX-T binding, massive complement C3b, and membrane attack complex deposition in glomeruli in the regions of injury and decreased glomerular expression of complement regulators. A pathogenic role for complement was confirmed by demonstrating that systemic complement inhibition reduced renal injury. The isolated nephrotoxic component, a 115-kd protein toxin (PsTX-115), was shown to cause identical renal pathology. The demonstration that PsTX-T and PsTX-115 were highly nephrotoxic acting via induction of complement activation suggests that inhibition of complement might be used to prevent acute renal damage following envenomation by P. semoni.

IgG1, IgG2a, complement and FcR in immune complex disease (90.16)

Abstract Immune complexes (IC) contribute to tissue damage in some diseases but prevent it in others. We hypothesize that this contrast occurs because different immunoglobulin (Ig) isotypes involved in the IC interact differently with both stimulatory and inhibitory FcγR and complement (C). To study this we compared IgG1 (which poorly activates C and stimulatory FcγR) to other Ig isotype-mediated IC processing using a model in which wild-type (WT) mice immunized with goat anti-mouse IgD antiserum (GaMD), produce a large, predominantly IgG1 Ab response, produce large amounts of IC, and eliminate them without developing renal disease. In GaMD-immunized IgG1-deficient mice, IgG2a (which activates C and interacts with stimulatory FcγR) predominates and mice develop severe, lethal IC-mediated renal disease that can be prevented by immunization with antigen-specific IgG1. Renal disease also develops in C3-deficient, FcγRIIb-deficient and anti-FcγRII/III mAb treated WT but not in FcRγ-deficient mice immunized with GaMD. Surprisingly, GaMD-immunized IgG1-deficient mice that are also lacking both FcRγ and C3 or lacking FcRγ and treated with a C5aR antagonist still develop renal disease. Together these findings suggest that IgG1 may protect against IC disease through a C- and FcγRIIb-dependent mechanism, while non-IgG1, presumably IgG2a, may mediate IC injury through a FcRγ- and either C3 or C5aR-independent mechanism.

A Study for an Allergic Inflammation Model Using Human Lungs and Its Pharmacological Application

The complement system, which plays an important role in inmate immunity, is considered to be important in the pathophysiology of allergic asthma. A patient with allergic asthma shows the reversible characteristic system of bronchoconstriction, increased mucus secretion, and complicated airway inflammation. Various cytokines secreted from Th2 cells contribute to the system. Cysteinyl-leukotrienes (CysLTs) are also considered to be one of the important mediators involved in asthmatic pathophysiology. However, the effects of a drug on humans may not be the same as those on animals due to species differences in complement-related molecules. In this series of experiments, we tried to establish a model in which the effects of a drug on the production of CysLTs from human lung preparations were evaluated following an anaphylactic reaction. CysLT production increased when the passively sensitized lung tissues were stimulated with anti-IgE antibody. The coaddition of anaphylatoxin, C5a, with the anti-IgE antibody potentiated CysLT production. The response to C3a was weaker when compared with that to C5a. In addition, increased production of CysLTs by adding serum at a specific ratio was dose dependently inhibited by nonpeptide C5a receptor antagonist, W-54011, or a novel complementary peptide inhibitor of C5a, acetyl peptide A. From these results, it is suggested that C5a potentiates cysLT production from human lung tissues and contributes to allergic inflammation like asthma, and thus acetylated peptide A and W-54011 are useful for suppressing allergic inflammation in the lungs.

Complement C5 Mediates Experimental Tubulointerstitial Fibrosis

Renal fibrosis is the final common pathway of most progressive renal diseases. C5 was recently identified as a risk factor for liver fibrosis. This study investigated the role of C5 in the development of renal tubulointerstitial fibrosis by (1) induction of renal fibrosis in wild-type and C5(-/-) mice by unilateral ureteral ligation (UUO) and (2) investigation of the effects of a C5a receptor antagonist (C5aRA) in UUO. In C5(-/-) mice, when compared with wild-type controls, markers of renal fibrosis (Sirius Red, type I collagen, fibronectin, alpha-smooth muscle actin, vimentin, and infiltrating macrophages) were significantly reduced on day 5 of UUO. On day 10, fibronectin mRNA and protein expression were still reduced in the C5(-/-) mice. Cortical mRNA of all PDGF isoforms and of TGF-beta(1) (i.e., central mediators of renal disease) were significantly reduced in C5(-/-) mice when compared with controls. Renal tubular cell expression of the C5aR was sparse in normal cortex but markedly upregulated after UUO. Treatment of wild-type UUO mice with C5aRA also led to a significant reduction of cortical Sirius Red staining, fibronectin protein expression, and PDGF-B mRNA expression on day 5. Neither genetic C5 deficiency nor C5aRA treatment caused any histologic changes in the nonobstructed kidneys. In cultured murine cortical tubular cells, C5a stimulated production of TGF-beta(1), and this was inhibited by C5aRA. Using a combined genetic and pharmacologic approach, C5, in particular C5a, is identified as a novel profibrotic factor in renal disease and as a potential new therapeutic target.

The role of complement C3 opsonization, C5a receptor, and CD14 inE. coli-induced up-regulation of granulocyte and monocyte CD11b/CD18 (CR3), phagocytosis, and oxidative burst in human whole blood

The relative role of complement and CD14 in Escherichia coli-induced leukocyte CD11b up-regulation, phagocytosis, and oxidative burst in human whole blood was examined. The highly specific thrombin inhibitor lepirudin was used as anticoagulant, as it does not affect complement activation. Complement inhibition at the level of C3 (anti-C2 and anti-factor D) and C5 (C5a receptor antagonist and anti-C5/C5a) efficiently inhibited CD11b up-regulation, phagocytosis, and oxidative burst in granulocytes. Monocyte activation was generally less complement-dependent, but when C3 activation was blocked, a pronounced inhibition of phagocytosis and oxidative burst was obtained. Only the combination of anti-C2 and antifactor D blocked E. coli C3 opsonization completely. Whole E. coli, disrupted E. coli, and the C3-convertase activator cobra venom factor up-regulated CD11b rapidly on both cell types, proportional to their complement activation potential in the fluid phase. In comparison, purified LPS at concentrations comparable with that present in the E. coli preparations did not activate complement. Oxidative burst was induced only by whole bacteria. Finally, the combination of complement inhibition and anti-CD14 completely blocked E. coli-induced granulocyte and monocyte CD11b up-regulation and quantitatively, virtually abolished phagocytosis. The results indicate that complement and CD14, despite differential effects on granulocytes and monocytes, are the two crucial, quantitative factors responsible for E. coli-induced CD11b, phagocytosis, and oxidative burst in both cell types.

Leukocyte Effects of C5a-Receptor Blockade During Simulated Extracorporeal Circulation

Distinct pathways of leukocyte activation during simulated cardiopulmonary bypass are mediated by the complement C5a anaphylatoxin. We hypothesized that a human C5a receptor antagonist would specifically inhibit the inflammatory response of neutrophils to simulated extracorporeal circulation, while preserving the C5b-9 pathway for innate immunity. An in vitro extracorporeal circuit recirculated fresh heparinized whole blood through a membrane oxygenator with and without addition of a small molecule human C5a receptor antagonist. Samples were periodically drawn over 90 minutes for complement and leukocyte activation studies. Addition of the C5a receptor antagonist to simulated extracorporeal circulation abrogated both neutrophil CD11b upregulation and interleukin 8 release (p < 0.01 for both), despite full generation of C3a and C5b-9; however, elastase release from neutrophils was unaffected. Although C5a receptor blockade only trended toward inhibiting monocyte CD11b upregulation (p = 0.09), circuit clearance of both monocytes (p = 0.04) and neutrophils (p = 0.01) was significantly decreased. In addition, the C5a receptor antagonist completely blocked both neutrophil-platelet and monocyte-platelet conjugate formation (p < 0.001 for both), without affecting platelet P-selectin expression. C5a receptor blockade during simulated extracorporeal circulation completely blocked neutrophil beta2 integrin upregulation and induction of plasma interleukin 8, suggesting an acute downregulatory effect on neutrophil chemotaxis-related pathways, while preserving terminal complement generation and neutrophil elastase release. Inhibition of leukocyte-platelet conjugate formation suggests a novel function for leukocyte adhesive receptors, possibly related to preservation of elastase generation.

The Unexpected Role of C3a-C3aR Axis in Maturation of Erythroid Cells - Implications for Pathogenesis of Hypoxia-Related Polycythemias.

Hypoxia leads both to secondary polycythemia and activation of the complement (C) cascade in bone marrow (BM). We reported that C is activated in BM during hypoxia by the IgM-dependent classical pathway (Blood 2006, 107:835–840). These observations prompted us to study the potential role of C cleavage fragments in erythropoiesis. We noticed that human and murine cells from the erythroid lineage express functional G-protein coupled C3aR and that C3 cleavage fragment - anaphylatoxin C3a enhances SDF-1-mediated chemotaxis of erythroid progenitors and induces in normal erythroblasts i) adhesion to fibronectin, ii) calcium flux and iii) phosphorylation of MAPKp42/44. To further explore a potential role of the C3a-C3aR axis in erythropoiesis we focused on C3aR−/ − mice. We noticed that these animals as compared to wild type (wt) littermates have i) lower hematocrits and ii) display delayed recovery of hematocrit values after sublethal irradiation. More importantly, we noticed that the number of clonogeneic BFU-E progenitors in BM of C3aR−/ − mice, but not CFU-GM and CFU-Meg, is reduced by ~65%. Thus, in order to learn more about the role of the C3a-C3aR axis in erythropoiesis we moved to an in vitro model and costimulated normal human CD34+ or murine SKL cells with C3a or desArgC3a in the presence of suboptimal doses of EpO and KL, but no effect on BFU-E colony formation was observed. We found, however, that C3a increases in early erythroid cells expression of mRNA for VEGF and MMP-2 - which are important for maturation/enucleation and egress of erythrocytes from the BM into peripheral blood. These data were subsequently confirmed by ELISA (VEGF) and zymography (MMP-2). Furthermore, we noticed that at the morphological level C3aR−/ − mice as compared to wt littermates had significantly reduced number of erytropoietic islands in BM. This suggests that in the absence of the C3aR, maturing erythroblasts do not traffic properly in the marrow microenvironment in BM and do not associate with C3 producing central macrophages in erythropoietic islands. In conclusion the C3a-C3aR axis modulates the interaction of erythroid progenitors with the hematopoietic environment and we conclude that it plays an important and underappreciated role in developmental trafficking of erythroid cells in BM. Thus C3a may enhance the erythropoietic effects of erythropoietin in polycythemias related to hypoxia. Furthermore, since hypoxia activates C, we postulate that C cleavage fragments could be involved in the pathogenesis of hypoxia-related polycythemias and C3aR antagonists could find a potential supportive role in controlling erythropoiesis in these patients.

Impaired Engraftment of Hematopoietic Stem/Progenitor Cells (HSPC) in Immunodefcient Mice Supports an Important Role of Complement System for Optimal Homing.

Recently we demonstrated that conditioning for transplantation (radio-chemotherapy) activates complement (C) in bone marrow (BM) and that the third complement component (C3) cleavage fragments (C3a and desArgC3a) increase responsiveness of hematopoietic stem/progenitor cells (HSPC) to stromal-derived factor (SDF)-1 gradient by enhancing the incorporation of CXCR4 into membrane lipid rafts – what enables its better interaction with small GTPases from the Rho/Rac family (Blood 2003, 101, 3784, Blood 2005, 105, 40–48). Based on these data we hypothesized that C could affect the homing/engraftment of HSPC. Thus we performed transplant experiments in several strains of immunodeficient animals. First, we noticed that lethally irradiated NOD/SCID mice engrafted worse with wt HSPC as compared to wt animals (~30% decrease in a presence of donor-derived clonogeneic CFU-GM in marrow cavities 24 hrs after transplantation). This impaired engraftment correlated with the lack of C activation in BM after conditioning for transplantation by lethal irradiation. The lack of C activation in NOD/SCID mice after conditioning for transplant could be explained by a lack of IgM antibodies that activate C by classical IgM-dependent pathway. Next, to learn more on the molecular mechanisms of C cascade activation during conditioning for transplantation and the role of the C3a-C3aR axis in engraftment of HSPC we studied engraftment i) of wild type (wt) murine HSPC in immunodeficient mice (C3−/− and C3aR−/−) and ii) murine HSPC derived from C3aR−/− or C3−/− deficient mice in wild type littermates. The engraftment of HSPC was evaluated by i) recovery of peripheral blood cell counts in transplanted animals, ii) number of CFU-S colonies and iii) number of clonogeneic progenitors in marrow cavities 16 day after transplantation. We noticed that both C3−/− and C3aR−/− mice had impaired engraftment with wt HSPC. At the same time HSPC from C3aR−/− mice but not C3−/− animals showed poor engraftment in wt recipients. This suggests that i) C3aR expressed on HSPC interacts with C3a generated during C-activation in BM environment and ii) that this interaction is important for optimal homing of HSPC. To support further this notion, human CD34+ cells were exposed to nontoxic doses of C3aR antagonist SB290157 and transplanted into NOD/SCID mice. Subsequently, 24 hrs after transplantation cells were isolated from the marrow cavities and stimulated to grow human CFU-GM colonies. By employing this assay we noticed reduced engraftment of human CD34+ cells (~30%, p<0.0001) as compared to mice transplanted with control CD34+ cells unexposed to SB290157. These data allow for the following conclusions: i) C is activated in BM during conditioning for transplantation by irradiation ii) C is activated after exposure of a natural neoepitope in damaged marrow which is recognized by natural IgM activating C via the classical pathway, iii) C3 cleavage product C3a binds to C3aR on transplanted HSPC and increases incorporation of CXCR4 into membrane lipid rafts enhancing responsiveness of HSPC to an SDF-1 gradient, and finally iv) a proper interplay between the C system and SDF-1-CXCR axis ensures optimal homing of HSPC.

C5 Modulates Airway Hyperreactivity and Pulmonary Eosinophilia during Enhanced Respiratory Syncytial Virus Disease by Decreasing C3a Receptor Expression

Enhanced respiratory syncytial virus disease, a serious pulmonary disorder that affected recipients of an inactivated vaccine against respiratory syncytial virus in the 1960s, has delayed the development of vaccines against the virus. The enhanced disease was characterized by immune complex-mediated airway hyperreactivity and a severe pneumonia associated with pulmonary eosinophilia. In this paper, we show that complement factors contribute to enhanced-disease phenotypes. Mice with a targeted disruption of complement component C5 affected by the enhanced disease displayed enhanced airway reactivity, lung eosinophilia, and mucus production compared to wild-type mice and C5-deficient mice reconstituted with C5. C3aR expression in bronchial epithelial and smooth muscle cells in the lungs of C5-deficient mice was enhanced compared to that in wild-type and reconstituted rodents. Treatment of C5-deficient mice with a C3aR antagonist significantly attenuated airway reactivity, eosinophilia, and mucus production. These results indicate that C5 plays a crucial role in modulating the enhanced-disease phenotype, by affecting expression of C3aR in the lungs. These findings reveal a novel autoregulatory mechanism for the complement cascade that affects the innate and adaptive immune responses.

A Novel C5a Receptor-Tissue Factor Cross-Talk in Neutrophils Links Innate Immunity to Coagulation Pathways

Neutrophils and complement are key sentinels of innate immunity and mediators of acute inflammation. Recent studies have suggested that inflammatory processes modulate thrombogenic pathways. To date, the potential cross-talk between innate immunity and thrombosis and the precise molecular pathway by which complement and neutrophils trigger the coagulation process have remained elusive. In this study, we demonstrate that antiphospholipid Ab-induced complement activation and downstream signaling via C5a receptors in neutrophils leads to the induction of tissue factor (TF), a key initiating component of the blood coagulation cascade. TF expression by neutrophils was associated with an enhanced procoagulant activity, as verified by a modified prothrombin time assay inhibited by anti-TF mAb. Inhibition studies using the complement inhibitor compstatin revealed that complement activation is triggered by antiphospholipid syndrome (APS) IgG and leads to the induction of a TF-dependent coagulant activity. Blockade studies using a selective C5a receptor antagonist and stimulation of neutrophils with recombinant human C5a demonstrated that C5a, and its receptor C5aR, mediate the expression of TF in neutrophils and thereby significantly enhance the procoagulant activity of neutrophils exposed to APS serum. These results identify a novel cross-talk between the complement and coagulation cascades that can potentially be exploited therapeutically in the treatment of APS and other complement-associated thrombotic diseases.

A C5a receptor antagonist provides greater protection than an NSAID in a skin pouch model of sodium urate induced inflammation

A C5a receptor antagonist provides greater protection than an NSAID in a skin pouch model of sodium urate induced inflammation

Peptidomimetic C5a receptor antagonists with hydrophobic substitutions at the C-terminus: Increased receptor specificity and in vivo activity

A new class of peptidomimetic C5a receptor antagonists characterized by C-terminal amino acids with hydrophobic side chains is presented. Systematic optimization of the first hits led to JPE1375 ( 36), which was intensively characterized in vitro and in vivo. Compound 36 exhibits high microsomal stability and receptor specificity and is highly active in an immune complex mediated peritonitis model (reverse passive Arthus reaction) in mice.

Therapeutic activity of C5a receptor antagonists in a rat model of neurodegeneration

The complement system is thought to be involved in the pathogenesis of numerous neurological diseases, although its precise role remains controversial. In this study we used orally active C5a receptor antagonists (PMX53 and PMX205) developed in our laboratories in a rat model of 3-nitropropionic acid (3-NP) -induced Huntington's disease. Administration of the C5a antagonists (10 mg/kg/day, oral) either 48 h pre- or 48 h post-toxin significantly reduced body weight loss, anorexia, and behavioral and motor deficits associated with 3-NP intoxication. Striatal lesion size, apoptosis, neutrophil infiltration, and hemorrhage were also significantly reduced in C5a antagonist-treated rats. Immunohistochemical analysis demonstrated marked deposition of C3 and C9, and up-regulation of C5a receptors on neuronal cells at the time of lesion formation. Inhibition of prostaglandins or TNF-alpha with ibuprofen or infliximab had no effect in this model. The C5a antagonists did not affect 3-NP-induced cell death when added directly to rat striatal neuronal cultures, indicating a secondary mechanism of action in vivo. Our findings demonstrate for the first time that complement activation in the brain, particularly C5a, is a key event in the pathogenesis of this disease model, and suggest a future role for inhibitors of C5a in the treatment of neurodegenerative diseases.

Complement Component C3 Mediates Inflammatory Injury Following Focal Cerebral Ischemia

The complement cascade has been implicated in ischemia/reperfusion injury, and recent studies have shown that complement inhibition is a promising treatment option for acute stroke. The development of clinically useful therapies has been hindered, however, by insufficient understanding of which complement subcomponents contribute to post-ischemic injury. To address this issue, we subjected mice deficient in selected complement proteins (C1q, C3, C5) to transient focal cerebral ischemia. Of the strains investigated, only C3-/- mice were protected, as demonstrated by 34% reductions in both infarct volume (P<0.01) and neurological deficit score (P<0.05). C3-deficient mice also manifested decreased granulocyte infiltration (P<0.02) and reduced oxidative stress (P<0.05). Finally, administration of a C3a-receptor antagonist resulted in commensurate neurological improvement and stroke volume reduction (P<0.05). Together, these results establish C3 activation as the key constituent in complement-related inflammatory tissue injury following stroke and suggest a C3a anaphylatoxin-mediated mechanism.

Blockade of the Complement C5a Receptor Reduces Incisional Allodynia, Edema, and Cytokine Expression

Activation of the complement system is one component of the inflammatory response. Various components of the complement system participate in killing foreign organisms, recruiting immune cells, enhancing edema, and stimulating cytokine formation. Complement-mediated enhancement of the inflammation surrounding surgical incisions may increase pain. In these studies, the authors used a murine hind paw incisional model to study the role of the complement C5a receptor in supporting incisional inflammation. At baseline and at various time points after incision, they measured the effects of a highly selective C5a receptor antagonist on nociceptive thresholds, edema formation, and cytokine production in the skin surrounding the incision. They also measured changes in C5a receptor expression near the incisions. The once-daily injection of the C5a receptor antagonist AcF-[OPdChaWR] reduced mechanical allodynia and edema in the incised hind paw. A multiplexed cytokine assay revealed that 8 of the 18 cytokines examined showed significant increases in skin tissue abundance after incision. Distinct time courses for the patterns of elevation were seen, though some degree of resolution occurred for all cytokines within 96 h. For 7 of these 8 cytokines, the C5a receptor antagonist reduced the enhancement of expression. In addition, the authors found that the C5a receptor messenger RNA level increased 15-fold in the skin surrounding the incisions within 24 h and then slowly declined. The tissue directly surrounding incisions in mouse hind paws undergoes large changes in the content of specific cytokines in addition to demonstrating edema and nociceptive sensitization. By blocking the receptor for one component of the complement system, C5a, all of these changes can be reduced. Complement receptor inhibitors may constitute a novel group of compounds useful in reducing the pain and swelling of surgical incisions.

Complement inhibitors selectively attenuate injury following administration of cobra venom factor to rats

Systemic activation of complement is a pathophysiological response common to severe disturbances such as hemorrhagic shock, major burn injury and sepsis. Intravenous infusion of cobra venom factor (CVF) has been used as an animal model of acute respiratory distress syndrome (ARDS), and reliably and selectively induces rapid intravascular activation of the complement system, leading to acute organ damage. In the present study, we have used different complement inhibitors to investigate the roles of complement products in CVF-induced responses in the rat. Rats were treated with either a C5a receptor antagonist (C5aRA, AcF-[OP( d-Cha)WR], 1 mg/kg i.v. or 10 mg/kg p.o.), a C3a receptor antagonist (C3aRA, N(2)-[(2,2-diphenylethoxy)acetyl]- l-arginine, 0.1 mg/kg i.v.) or a convertase inhibitor, rosmarinic acid (RMA, 10 mg/kg i.v.), prior to CVF-induced complement challenge. Intravenous CVF resulted in hallmark events evident in the development of ARDS, including systemic neutropenia followed by neutrophil migration to the lung and bronchoalveolar vascular leakage, blood pressure alterations, and an increase in TNFα levels in both serum and bronchoalveolar lavage fluid. These hemodynamic changes were differentially inhibited by antagonism of C5a receptors, C3a receptors or by inhibition of the entire complement cascade using RMA. This evidence strongly implicates complement factors in the development of lung injury associated with systemic complement activation and identifies complement inhibition as a potential therapeutic target for acute syndromes such as ARDS and other severe systemic shock states mediated by activation of complement.

Recent developments in C5/C5a inhibitors

Complement factor 5a (C5a) is formed upon complement system activation in response to infection, injury or disease. Whilst C5a is a potent mediator of immune and inflammatory processes, excessive production or inadequate regulation of C5a has been implicated in the pathogenesis of numerous immuno-inflammatory diseases, predominantly through experimental studies utilising animal models of disease. Both acute and chronic conditions may benefit from C5a inhibition, including rheumatoid arthritis, inflammatory bowel disease, asthma, psoriasis, haemorrhagic shock and neurodegenerative conditions. The potentially broad clinical application for treatments that inhibit the activity of C5a at C5a receptors and the large global market for anti-inflammatory therapeutics have made C5a and the C5a receptor attractive targets for academic and commercial drug development programmes. In the past 5 years, interest in C5a as a drug target has grown substantially, and this activity has resulted in a collection of patents and scientific papers reporting novel C5a and C5a receptor inhibitors and antagonists, and generated a secondary stream of patent applications broadly claiming the use of C5/C5a inhibitors as a method of treating various immune and inflammatory conditions. This paper will review the physiology and pathophysiology of C5a and discuss the development of C5a and C5a receptor inhibitors in light of the recent scientific and patent literature.

Complement: a novel factor in basal and ischemia-induced neurogenesis

Through its involvement in inflammation, opsonization, and cytolysis, the complement protects against infectious agents. Although most of the complement proteins are synthesized in the central nervous system (CNS), the role of the complement system in the normal or ischemic CNS remains unclear. Here we demonstrate for the first time that neural progenitor cells and immature neurons express receptors for complement fragments C3a and C5a (C3a receptor (C3aR) and C5a receptor). Mice that are deficient in complement factor C3 (C3(-/-)) lack C3a and are unable to generate C5a through proteolytic cleavage of C5 by C5-convertase. Intriguingly, basal neurogenesis is decreased both in C3(-/-) mice and in mice lacking C3aR or mice treated with a C3aR antagonist. The C3(-/-) mice had impaired ischemia-induced neurogenesis both in the subventricular zone, the main source of neural progenitor cells in adult brain, and in the ischemic region, despite normal proliferative response and larger infarct volumes. Thus, in the adult mammalian CNS, complement activation products promote both basal and ischemia-induced neurogenesis.