-
Brain, Behavior, and Immunity May 2024Complement is dysregulated in the brain in Alzheimer's Disease and in mouse models of Alzheimer's disease. Each of the complement derived effectors, opsonins,...
Complement is dysregulated in the brain in Alzheimer's Disease and in mouse models of Alzheimer's disease. Each of the complement derived effectors, opsonins, anaphylatoxins and membrane attack complex (MAC), have been implicated as drivers of disease but their relative contributions remain unclarified. Here we have focussed on the MAC, a lytic and pro-inflammatory effector, in the App mouse amyloidopathy model. To test the role of MAC, we back-crossed to generate App mice deficient in C7, an essential MAC component. C7 deficiency ablated MAC formation, reduced synapse loss and amyloid load and improved cognition compared to complement-sufficient App mice at 8-10 months age. Adding back C7 caused increased MAC formation in brain and an acute loss of synapses in C7-deficient App mice. To explore whether C7 was a viable therapeutic target, a C7-blocking monoclonal antibody was administered systemically for one month in App mice aged 8-9 months. Treatment reduced brain MAC and amyloid deposition, increased synapse density and improved cognitive performance compared to isotype control-treated App mice. The findings implicate MAC as a driver of pathology and highlight the potential for complement inhibition at the level of MAC as a therapy in Alzheimer's disease.
Topics: Mice; Animals; Alzheimer Disease; Amyloid beta-Peptides; Amyloid beta-Protein Precursor; Cognitive Dysfunction; Mice, Transgenic; Plaque, Amyloid; Brain; Cognition; Complement Activation; Disease Models, Animal
PubMed: 38485063
DOI: 10.1016/j.bbi.2024.03.017 -
Frontiers in Immunology 2024Single nucleotide variations (SNVs) are specific genetic variations that commonly occur in a population and often do not manifest phenotypically. However, depending on...
INTRODUCTION
Single nucleotide variations (SNVs) are specific genetic variations that commonly occur in a population and often do not manifest phenotypically. However, depending on their location and the type of nucleotide exchanged, an SNV can alter or inhibit the function of the gene in which it occurs. Immunoglobulin G (IgG) receptor genes have exhibited several polymorphisms, including rs1801274, which is found in the FcgRIIa gene. The replacement of A with T results in a Histidine (H) to Arginine (R) substitution, altering the affinity of the IgG receptor for IgG subtypes and C-reactive protein (CRP). In this study, we analyzed rs1801274 and its functional implications concerning L. Infantum uptake and cytokine production.
METHODS
We genotyped 201 individuals from an endemic area for visceral leishmaniasis to assess the presence of rs1801274 using Taqman probes for a candidate gene study. Additionally, we included seventy individuals from a non-endemic area for a functional study. Subsequently, we isolated and cultivated one-week adherent mononuclear cells (AMCs) derived from the peripheral blood of participants residing in the non-endemic region in the presence of L. infantum promastigotes, with and without antigen-specific IgG and/or CRP. We analyzed the rate of phagocytosis and the production of nitric oxide (NO), tumor necrosis factor (TNF)-a, interleukin (IL)-10, IL-12 p70, IL-1b, IL- 6, and IL-8 in the culture supernatants.
RESULTS AND DISCUSSION
In participants from the endemic region, the A/G (H/R isoform) heterozygous genotype was significantly associated with susceptibility to the disease. Furthermore, SNVs induced a change in the phagocytosis rate in an opsonin-dependent manner. Opsonization with IgG increased the production of IL-10, TNF-a, and IL-6 in AMCs with the H/R isoform, followed by a decrease in NO production. The results presented here suggest that the rs1801274 polymorphism is linked to a higher susceptibility to visceral leishmaniasis.
Topics: Humans; Leishmaniasis, Visceral; Leishmania infantum; Receptors, IgG; Interleukin-12; Tumor Necrosis Factor-alpha; Nucleotides; Protein Isoforms; Genetic Variation; Immunoglobulin G
PubMed: 38455048
DOI: 10.3389/fimmu.2024.1343602 -
The Journal of Biological Chemistry Apr 2024The introduction of a therapeutic anti-C5 antibody into clinical practice in 2007 inspired a surge into the development of complement-targeted therapies. This has led to...
The introduction of a therapeutic anti-C5 antibody into clinical practice in 2007 inspired a surge into the development of complement-targeted therapies. This has led to the recent approval of a C3 inhibitory peptide, an antibody directed against C1s and a full pipeline of several complement inhibitors in preclinical and clinical development. However, no inhibitor is available that efficiently inhibits all three complement initiation pathways and targets host cell surface markers as well as complement opsonins. To overcome this, we engineered a novel fusion protein combining selected domains of the three natural complement regulatory proteins decay accelerating factor, factor H and complement receptor 1. Such a triple fusion complement inhibitor (TriFu) was recombinantly expressed and purified alongside multiple variants and its building blocks. We analyzed these proteins for ligand binding affinity and decay acceleration activity by surface plasmon resonance. Additionally, we tested complement inhibition in several in vitro/ex vivo assays using standard classical and alternative pathway restricted hemolysis assays next to hemolysis assays with paroxysmal nocturnal hemoglobinuria erythrocytes. A novel in vitro model of the alternative pathway disease C3 glomerulopathy was established to evaluate the potential of the inhibitors to stop C3 deposition on endothelial cells. Next to the novel engineered triple fusion variants which inactivate complement convertases in an enzyme-like fashion, stoichiometric complement inhibitors targeting C3, C5, factor B, and factor D were tested as comparators. The triple fusion approach yielded a potent complement inhibitor that efficiently inhibits all three complement initiation pathways while targeting to surface markers.
Topics: Humans; Complement Factor H; Recombinant Fusion Proteins; Complement Activation; CD55 Antigens; Hemolysis; Complement Pathway, Alternative; Complement Inactivating Agents; Erythrocytes; Receptors, Complement 3b
PubMed: 38401844
DOI: 10.1016/j.jbc.2024.105784 -
Cancer Research Communications Feb 2024In normal cells, binding of the transmembrane protein CD47 to signal regulatory protein-α (SIRPα) on macrophages induces an antiphagocytic signal. Tumor cells hijack...
UNLABELLED
In normal cells, binding of the transmembrane protein CD47 to signal regulatory protein-α (SIRPα) on macrophages induces an antiphagocytic signal. Tumor cells hijack this pathway and overexpress CD47 to evade immune destruction. Macrophage antitumor activity can be restored by simultaneously blocking the CD47-SIRPα signaling axis and inducing a prophagocytic signal via tumor-opsonizing antibodies. We identified a novel, fully human mAb (BMS-986351) that binds SIRPα with high affinity. BMS-986351 demonstrated broad binding coverage across SIRPα polymorphisms and potently blocked CD47-SIRPα binding at the CD47 binding site in a dose-dependent manner. In vitro, BMS-986351 increased phagocytic activity against cell lines from solid tumors and hematologic malignancies, and this effect was markedly enhanced when BMS-986351 was combined with the opsonizing antibodies cetuximab and rituximab. A phase I dose-escalation/-expansion study of BMS-986351 for the treatment of advanced solid and hematologic malignancies is underway (NCT03783403).
SIGNIFICANCE
Increasing the phagocytotic capabilities of tumor-associated macrophages by modulating macrophage-tumor cell surface signaling via the CD47-SIRPα axis is a novel strategy. Molecules targeting CD47 have potential but its ubiquitous expression necessitates higher therapeutic doses to overcome potential antigen sink effects. The restricted expression pattern of SIRPα may limit toxicities and lower doses of the SIRPα antibody BMS-986351 may overcome target mediated drug disposition while maintaining the desired pharmacology.
Topics: Humans; CD47 Antigen; Receptors, Immunologic; Phagocytosis; Macrophages; Neoplasms; Antibodies, Neoplasm; Opsonin Proteins; Hematologic Neoplasms
PubMed: 38319147
DOI: 10.1158/2767-9764.CRC-23-0634 -
Frontiers in Cellular and Infection... 2023The complement receptor CR3, also known as integrin Mac-1 (CD11b/CD18), is one of the major phagocytic receptors on the surface of neutrophils and macrophages. We...
The complement receptor CR3, also known as integrin Mac-1 (CD11b/CD18), is one of the major phagocytic receptors on the surface of neutrophils and macrophages. We previously demonstrated that in its protein ligands, Mac-1 binds sequences enriched in basic and hydrophobic residues and strongly disfavors negatively charged sequences. The avoidance by Mac-1 of negatively charged surfaces suggests that the bacterial wall and bacterial capsule possessing net negative electrostatic charge may repel Mac-1 and that the cationic Mac-1 ligands can overcome this evasion by acting as opsonins. Indeed, we previously showed that opsonization of Gram-negative with several cationic peptides, including PF4 (Platelet Factor 4), strongly augmented phagocytosis by macrophages. Here, we investigated the effect of recombinant PF4 (rPF4) on phagocytosis of Gram-positive and examined its impact in a mouse model of peritonitis. Characterization of the interaction of rPF4 with nonencapsulated and encapsulated showed that rPF4 localizes on the bacterial surface, thus making it available for Mac-1. Furthermore, rPF4 did not have direct bactericidal and bacteriostatic activity and was not toxic to host cells. rPF4 enhanced phagocytosis of bioparticles by various primary and cultured Mac-1-expressing leukocytes by several folds. It also increased phagocytosis of live nonencapsulated and encapsulated bacteria. Notably, the augmentation of phagocytosis by rPF4 did not compromise the intracellular killing of by macrophages. Using a murine peritonitis model, we showed that treatment of infected mice with rPF4 caused a significant increase in the clearance of antibiotic-susceptible and its methicillin-resistant (MRSA) variant and markedly improved survival. These findings indicate that rPF4 binding to the bacterial surface circumvents its antiphagocytic properties, improving host defense against antibiotic-susceptible and antibiotic-resistant bacteria.
Topics: Animals; Mice; Methicillin-Resistant Staphylococcus aureus; Anti-Bacterial Agents; Platelet Factor 4; Staphylococcus aureus; Disease Models, Animal; Phagocytosis; Macrophage-1 Antigen; Immunologic Factors; Peritonitis
PubMed: 37868353
DOI: 10.3389/fcimb.2023.1217103 -
Frontiers in Immunology 2023C-reactive protein (CRP) is one of the major members of the family of acute phase proteins (APP). Interest in this CRP was the result of a seminal discovery of its... (Review)
Review
C-reactive protein (CRP) is one of the major members of the family of acute phase proteins (APP). Interest in this CRP was the result of a seminal discovery of its pattern of response to pneumococcal infection in humans. CRP has the unique property of reacting with phosphocholine-containing substances, such as pneumococcal C-polysaccharide, in the presence of Ca. The attention regarding the origin of CRP and its multifunctionality has gripped researchers for several decades. The reason can be traced to the integrated evolution of CRP in the animal kingdom. CRP has been unequivocally listed as a key indicator of infectious and inflammatory diseases including autoimmune diseases. The first occurrence of CRP in the evolutionary ladder appeared in arthropods followed by molluscs and much later in the chordates. The biological significance of CRP has been established in the animal kingdom starting from invertebrates. Interestingly, the site of synthesis of CRP is mainly the liver in vertebrates, while in invertebrates it is located in diverse tissues. CRP is a multifunctional player in the scenario of innate immunity. CRP acts as an opsonin in the area of complement activation and phagocytosis. Interestingly, CRP upregulates and downregulates both cytokine production and chemotaxis. Considering various studies of CRP in humans and non-human animals, it has been logically proposed that CRP plays a common role in animals. CRP also interacts with Fcγ receptors and triggers the inflammatory response of macrophages. CRP in other animals such as primates, fish, echinoderms, arthropods, and molluscs has also been studied in some detail which establishes the evolutionary significance of CRP. In mammals, the increase in CRP levels is an induced response to inflammation or trauma; interestingly, in arthropods and molluscs, CRP is constitutively expressed and represents a major component of their hemolymph. Investigations into the primary structure of CRP from various species revealed the overall relatedness between vertebrate and invertebrate CRP. Invertebrates lack an acquired immune response; they are therefore dependent on the multifunctional role of CRP leading to the evolutionary success of the invertebrate phyla.
Topics: Animals; C-Reactive Protein; Inflammation; Invertebrates; Mammals; Opsonin Proteins; Phagocytosis; Humans
PubMed: 37860004
DOI: 10.3389/fimmu.2023.1238411 -
Frontiers in Immunology 2023Systemic amyloidosis is a progressive disorder characterized by the extracellular deposition of amyloid fibrils and accessory proteins in visceral organs and tissues....
INTRODUCTION
Systemic amyloidosis is a progressive disorder characterized by the extracellular deposition of amyloid fibrils and accessory proteins in visceral organs and tissues. Amyloid accumulation causes organ dysfunction and is not generally cleared by the immune system. Current treatment focuses on reducing amyloid precursor protein synthesis and slowing amyloid deposition. However, curative interventions will likely also require removal of preexisting amyloid deposits to restore organ function. Here we describe a prototypic pan-amyloid binding peptide-antibody fusion molecule (mIgp5) that enhances macrophage uptake of amyloid.
METHODS
The murine IgG1-IgG2a hybrid immunoglobulin with a pan amyloid-reactive peptide, p5, fused genetically to the N-terminal of the immunoglobulin light chain was synthesized in HEK293T/17 cells. The binding of the p5 peptide moiety was assayed using synthetic amyloid-like fibrils, human amyloid extracts and amyloid-laden tissues as substrates. Binding of radioiodinated mIgp5 with amyloid deposits was evaluated in a murine model of AA amyloidosis using small animal imaging and microautoradiography. The bioactivity of mIgp5 was assessed in complement fixation and phagocytosis assays in the presence of patient-derived amyloid extracts and synthetic amyloid fibrils as substrates and in the presence or absence of human serum.
RESULTS
Murine Igp5 exhibited highly potent binding to AL and ATTR amyloid extracts and diverse types of amyloid in formalin-fixed tissue sections. In the murine model of systemic AA amyloidosis, I-mIgp5 bound rapidly and specifically to amyloid deposits in all organs, including the heart, with no evidence of non-specific uptake in healthy tissues. The bioactivity of the immunoglobulin Fc domain was uncompromised in the context of mIgp5 and served as an effective opsonin. Macrophage-mediated uptake of amyloid extract and purified amyloid fibrils was enhanced by the addition of mIgp5. This effect was exaggerated in the presence of human serum coincident with deposition of complement C5b9.
CONCLUSION
Immunostimulatory, amyloid-clearing therapeutics can be developed by incorporating pan-amyloid-reactive peptides, such as p5, as a targeting moiety. The immunologic functionality of the IgG remains intact in the context of the fusion protein. These data highlight the potential use of peptide-antibody fusions as therapeutics for all types of systemic amyloidosis.
Topics: Mice; Animals; Humans; Disease Models, Animal; HEK293 Cells; Plaque, Amyloid; Amyloidosis; Amyloid; Amyloidogenic Proteins; Peptides; Immunoglobulin Light Chains
PubMed: 37854603
DOI: 10.3389/fimmu.2023.1275372 -
Microbiological Research Dec 2023The human complement system is an important part of the innate immune response in the fight against invasive bacteria. Complement responses can be activated... (Review)
Review
The human complement system is an important part of the innate immune response in the fight against invasive bacteria. Complement responses can be activated independently by the classical pathway, the lectin pathway, or the alternative pathway, each resulting in the formation of a C3 convertase that produces the anaphylatoxin C3a and the opsonin C3b by specifically cutting C3. Other important features of complement are the production of the chemotactic C5a peptide and the generation of the membrane attack complex to lyse intruding pathogens. Invasive pathogens like Staphylococcus aureus and several species of the genus Streptococcus have developed a variety of complement evasion strategies to resist complement activity thereby increasing their virulence and potential to cause disease. In this review, we focus on secreted complement evasion factors that assist the bacteria to avoid opsonization and terminal pathway lysis. We also briefly discuss the potential role of complement evasion factors for the development of vaccines and therapeutic interventions.
Topics: Humans; Gram-Positive Cocci; Immunity, Innate; Staphylococcal Infections; Immune System; Immune Evasion
PubMed: 37826985
DOI: 10.1016/j.micres.2023.127512 -
PeerJ 2023The complement system is an essential part of innate immunity. It is activated by invading pathogens causing inflammation, opsonization, and lysis complement...
The complement system is an essential part of innate immunity. It is activated by invading pathogens causing inflammation, opsonization, and lysis complement anaphylatoxins, complement opsonin's and membrane attack complex (MAC), respectively. However, in SARS-CoV-2 infection overactivation of complement system is causing cytokine storm leading to multiple organs damage. In this study, the René Thomas kinetic logic approach was used for the development of biological regulatory network (BRN) to model SARS-CoV-2 mediated complement system signalling pathways. Betweenness centrality analysis in cytoscape was adopted for the selection of the most biologically plausible states in state graph. Among the model results, in strongly connected components (SCCs) pro-inflammatory cytokines (PICyts) oscillatory behaviour between recurrent generation and downregulation was found as the main feature of SARS-CoV-2 infection. Diversion of trajectories from the SCCs leading toward hyper-inflammatory response was found in agreement with studies that overactive innate immunity response caused PICyts storm during SARS-CoV-2 infection. The complex of negative regulators FI, CR1 and DAF in the inhibition of complement peptide (C5a) and PICyts was found desirable to increase immune responses. In modelling role of MAC and PICyts in lowering of SARS-CoV-2 titre was found coherent with experimental studies. Intervention in upregulation of C5a and PICyts by C3 was found helpful in back-and-forth variation of signalling pattern linked with the levels of PICyts. Moreover, intervention in upregulation of PICyts by C5a was found productive in downregulation of all activating factors in the normal SCCs. However, the computational model predictions require experimental studies to be validated by exploring the activation role of C3 and C5a which could change levels of PICyts at various phases of SARS-CoV-2 infection.
Topics: Humans; Cytokines; COVID-19; SARS-CoV-2; Complement System Proteins; Complement Membrane Attack Complex
PubMed: 37744234
DOI: 10.7717/peerj.15794 -
BioRxiv : the Preprint Server For... Aug 2023The complement receptor CR3, also known as integrin Mac-1 (CD11b/CD18), is one of the major phagocytic receptors on the surface of neutrophils and macrophages. We...
The complement receptor CR3, also known as integrin Mac-1 (CD11b/CD18), is one of the major phagocytic receptors on the surface of neutrophils and macrophages. We previously demonstrated that in its protein ligands, Mac-1 binds sequences enriched in basic and hydrophobic residues and strongly disfavors negatively charged sequences. The avoidance by Mac-1 of negatively charged surfaces suggests that the bacterial wall and bacterial capsule possessing net negative electrostatic charge may repel Mac-1 and that the cationic Mac-1 ligands can overcome this evasion by acting as opsonins. Indeed, we previously showed that opsonization of Gram-negative with several cationic peptides, including PF4 (Platelet Factor 4), strongly augmented phagocytosis by macrophages. Here, we investigated the effect of recombinant PF4 (rPF4) on phagocytosis of Gram-positive in vitro and examined its impact in a mouse model of peritonitis. Characterization of the interaction of rPF4 with nonencapsulated and encapsulated showed that rPF4 localizes on the bacterial surface, thus making it available for Mac-1. Furthermore, rPF4 did not have direct bactericidal and bacteriostatic activity and was not toxic to host cells. rPF4 enhanced phagocytosis of bioparticles by various primary and cultured Mac-1-expressing leukocytes by several folds. It also increased phagocytosis of live nonencapsulated and encapsulated bacteria. Notably, the augmentation of phagocytosis by rPF4 did not compromise the intracellular killing of by macrophages. Using a murine peritonitis model, we showed that treatment of infected mice with rPF4 caused a significant increase in the clearance of antibiotic-susceptible and its methicillin-resistant (MRSA) variant and markedly improved survival. These findings indicate that rPF4 binding to the bacterial surface circumvents its antiphagocytic properties, improving host defense against antibiotic-susceptible and antibiotic-resistant bacteria.
PubMed: 37662328
DOI: 10.1101/2023.08.25.554865