IgG Fc Receptor Research Articles

Bovine IgG Prevents Experimental Infection With RSV and Facilitates Human T Cell Responses to RSV

Respiratory syncytial virus (RSV) infections represent a major burden of disease in infants and are the second most prevalent cause of death worldwide. Human milk immunoglobulins provide protection against RSV. However, many infants depend on processed bovine milk-based nutrition, which lacks intact immunoglobulins. We investigated the potential of bovine antibodies to neutralize human RSV and facilitate-cell immune activation. We show cow's milk IgG (bIgG) and Intravenous Immunoglobulin (IVIG) have a similar RSV neutralization capacity, even though bIgG has a lower pre-F to post-F binding ratio compared to human IVIG, with the majority of bIgG binding to pre-F. RSV is better neutralized with human IVIG. Consequently, we enriched RSV specific T cells by culturing human PBMC with a mixture of RSV peptides, and used these T cells to study the effect of bIgG and IVIG on the activation of pre-F-pecific T cells. bIgG facilitated in vitro T cell activation in a similar manner as IVIG. Moreover, bIgG was able to mediate T cell activation and internalization of pathogens, which are prerequisites for inducing an adaptive viral response. Using in vivo mouse experiments, we showed that bIgG is able to bind the murine activating IgG Fc Receptors (FcγR), but not the inhibiting FcγRII. Intranasal administration of the monoclonal antibody palivizumab, but also of bIgG and IVIG prevented RSV infection in mice. The concentration of bIgG needed to prevent infection was ~5-fold higher compared to IVIG. In conclusion, the data presented here indicate that functionally active bIgG facilitates adaptive antiviral T cell responses and prevents RSV infection in vitro and in vivo.

Mutations in the Reelin pathway are associated with abnormal expression of microglial IgG FC receptors in the cerebellar cortex.

Microglia are the immune cells of the central nervous system involved in a variety of developmental processes, such as regulation of cell death and survival, spatial patterning, and contribute to the development of Purkinje cells (PCs) during migration. Microglia express immunoglobulin G Fc receptors (FcgRs). In this report, we describe microglial FcgR expression and its relation to abnormal PC migration in the cerebellum during development. To detect microglial FcgR, the direct anti-IgG (secondary antisera) and high concentrations of Triton X-100 were applied as a method for labeling microglial cells without the use of any specific primary antiserum. By using Acp2-/- mice, which show an excessive PC migration into the molecular layer (ml), and 3 different types of mice with a null to alter the Reelin pathway (Reeler-, Dab1 (SCM)-, and Apoer mutant mice), we studied the location of PCs and the expression of FcgRs. Wild type littermates were used as controls in all studies. We show that the expression of microglial FcgRs was absent and PCs were ectopically located in the white matter in the cerebella of all mutant mice, except for the Acp2-/- mice (PCs were located in the ml). These results suggest a role for FcgRs in the Reelin signaling pathway, not in regulating PC migration, but rather in the adaptation to an environment with a relatively large number of ectopically located PCs. However, the exact correlation between the ectopic location of PCs and lack of FcgRs in Reeler, SCM, and Apoer-/- mice and the presence of FcgRs and directed PC location in the ml in Acp2-/- mice are yet to be determined.

The neonatal Fc receptor in cancer FcRn in cancer.

ABSTRACTSince the neonatal IgG Fc receptor (FcRn) was discovered, it was found to be involved in immunoglobulin recycling and biodistribution, immune complexes routing, antigen presentation, humoral immune response, and cancer immunosurveillance. The latest data show that FcRn plays a part in cancer pathophysiology. In various types of cancers, such as lung and colorectal cancer, FcRn has been described as an early marker for prognosis. Dysregulation of FcRn expression by cancer cells allows them to increase their metabolism, and this process could be exploited for passive targeting of cytotoxic drugs. However, the roles of this receptor depend on whether the studied cell population is the tumor tissue or the infiltrating cells, bringing forward the need for further studies.

Anti-domain 1 \u03b22 glycoprotein antibodies increase expression of tissue factor on monocytes and activate NK Cells and CD8+ cells in vitro

Backgroundβ2-Glycoprotein I (β2GPI) represents the major antigenic target for antiphospholipid antibodies (aPL), with domain 1 (D1) being identified as a risk factor for thrombosis and pregnancy complications in APS. We aimed to analyse the ability of aPL, and particularly anti-D1 β2GPI, to stimulate prothrombotic and proinflammatory activity of immune cells in vitro.MethodsPeripheral blood mononuclear cells (PBMCs) from 11 healthy individuals were incubated with: (1) “anti-D1(+)”—pooled plasma derived from patients suspected of having APS contained anticardiolipin antibodies (aCL), lupus anticoagulant (LA), anti-β2GPI and anti-D1 β2GPI; (2) “anti-D1(−)”—pooled plasma from patients suspected of having APS contained aCL, LA, anti-β2GPI, and negative for anti-D1 β2GPI; (3) “seronegative”—negative for aPL.ResultsThe presence of anti-D1(+) and anti-D1(−) plasma resulted in increased HLA-DR and CD11b on monocytes. While only anti-D1(+) plasma markedly increased the percentage and median fluorescence intensity (MFI) of CD142 (tissue factor, TF) on monocytes in comparison with those cultured with anti-D1(−) and seronegative plasma. Anti-D1(+) plasma resulted in increased percentage and MFI of activation marker CD69 on NK and T cytotoxic cells. Expression of IgG receptor FcγRIII(CD16) on monocytes and NK cells was down-regulated by the anti-D1(+) plasma.ConclusionsTaking together, our study shows the ability of patient-derived aPL to induce immune cell activation and TF expression on monocytes. For the first time, we demonstrated the influence of anti-D1 β2GPI on the activation status of monocytes, NK and cytotoxic T cells. Our findings further support a crucial role of D1 epitope in the promotion of thrombosis and obstetrical complications in APS.

Fcγ Receptor IIB Controls Skin Inflammation in an Active Model of Epidermolysis Bullosa Acquisita.

Epidermolysis bullosa acquisita (EBA) is an autoimmune skin blistering disease characterized by IgG autoantibodies (aAb) against type VII collagen (COL7). The mechanisms controlling the formation of such aAbs and their effector functions in the skin tissue are incompletely understood. Here, we assessed whether the inhibitory IgG Fc receptor, FcγRIIB, controls the development of autoimmune skin blistering disease in an active model of EBA. For this purpose, we immunized congenic EBA-susceptible B6.SJL-H2s (B6.s) and B6.s-Fcgr2b−/− mice with the immunodominant vWFA2 region of COL7. B6.s-Fcgr2b−/− mice developed a strong clinical phenotype with 15 ± 3.3% of affected body surface area at week 4. In contrast, the body surface area in B6.s mice was affected to a maximum of 5% at week 6 with almost no disease signs at week 4. Surprisingly, we already found strong but similar COL7-specific serum IgG1 and IgG2b aAb production at week 2. Further, aAb and C3b deposition in the skin of B6.s and B6.s-Fcgr2b−/− mice increased between weeks 2 and 6 after vWFA2 immunization. Importantly, neutrophil skin infiltration and activation was much stronger in B6s-Fcgr2b−/− than in B6.s mice and already present at week 2. Also, the early aAb response in B6.s-Fcgr2b−/− mice was more diverse than in wt B6.s mice. Reactive oxygen species (ROS) release from infiltrating neutrophils play a crucial role as mediator of skin inflammation in EBA. In line, sera from B6.s and B6.s-Fcgr2b−/− mice induced strong ROS release from bone marrow-neutrophils in vitro. In contrast to the antibody-transfer-induced EBA model, individual targeting of FcγRIII or FcγRIV decreased ROS release to 50%. Combined FcγR blocking abrogated ROS release from BM neutrophils. Also, ROS release induced by COL7-specific serum IgG aAbs was significantly higher using BM neutrophils from B6.s-Fcgr2b−/− than from B6.s mice. Together, our findings identified FcγRIIB as a suppressor of skin inflammation in the active EBA model through inhibition of early epitope spreading, protection from strong early neutrophil infiltration to and activation of neutrophils in the skin and suppression of FcγRIII activation by IgG1 aAbs which drive strong ROS release from neutrophils leading to tissue destruction at the dermal-epidermal junction.

Pharmacokinetic variability of therapeutic antibodies

Therapeutic antibodies have been increasingly used for the treatment of various diseases, including cancers and chronic inflammatory diseases. The pharmacokinetic interindividual variability of mAbs is large and influences, at least in part, the clinical response to antibody treatment. This variability is explained by a number of individual sources of variability, which are reviewed here. Some of them are major because they are frequently reported to greatly influence the interindividual variability; notably, increased body size, the presence of anti-drug antibodies, and high antigen mass are associated with decreased antibody concentrations. Other individual sources of variability are of less critical importance. They include sex, age, co-treatments, or genetic polymorphisms of IgG Fc receptors (FcgRs). The interindividual variability of antibody pharmacokinetics should be soundly described in order to design optimal dosing strategy.

Follicular Dendritic Cells Modulate Germinal Center B Cell Diversity through FcγRIIB

SUMMARYFollicular dendritic cells (FDCs), a rare and enigmatic stromal cell type in the B cell follicles of secondary lymphoid organs, store and present antigen to B cells. While essential for germinal center (GC) responses, their exact role during GC B cell selection remains unknown. FDCs upregulate the inhibitory IgG Fc receptor FcγRIIB during GC formation. We show that the stromal deficiency of FcγRIIB does not affect GC B cell frequencies compared to wild-type mice. However, in the absence of FcγRIIB on FDCs, GCs show aberrant B cell selection during autoreactive and selective foreign antigen responses. These GCs are more diverse as measured by the AidCreERT2 -confetti system and show the persistence of IgM+ clones with decreased numbers of IgH mutations. Our results show that FDCs can modulate GC B cell diversity by the upregulation of FcγRIIB. Permissive clonal selection and subsequent increased GC diversity may affect epitope spreading during autoimmunity and foreign responses.

P3113Functional deficiency in activating fcgamma receptor protects against aortic abdominal aneurysm in mice

Abstract Background and aim Abdominal aortic aneurysm (AAA) is a degenerative disorder characterized by a localized and permanent dilation of the aorta. Pathological features of AAA include proteolysis, vascular smooth muscle cells (VSMC) apoptosis, oxidative stress and inflammation. Previous studies have demonstrated the role of innate and adaptive immunity in the initiation and progression of AAA. However, the specific mechanisms of humoral, antibody-mediated, immune response elicited by self and non-self-antigens are not completely known. IgG Fc receptors (FcgammaR) play an important role in the initiation and regulation of many immunological and inflammatory processes, thus providing a link between humoral and cellular immune responses. In this work, we focus on IgG immune response against antigens exposed in the damaged vessel, analysing the specific role played by FcgammaR in the pathogenesis of AAA. Methods and results AAA was induced by aortic elastase perfusion in wild-type (WT) C57BL/6 mice (n=24 males, 12 weeks old). Compared with healthy aortas, AAA tissue showed IgG and IgM deposition and increased expression levels of activating (IA, IIIA and IVA) and inhibitory (IIB) FcgammaR (5-, 3-, 10, and 2-fold increases; p<0.005) at day 14 post-elastase perfusion. To explore the functional contribution of activating FcgammaR to AAA formation, parallel experiments were performed in mice deficient in gamma-chain (gamma-KO), the common signalling subunit of FcgammaRI, III and IV. Compared with WT mice, gamma-KO mice (n=21) exhibited lower IgG and IgM deposits (21±4% and 28±12% vs WT, respectively; p<0.02) and decreased AAA lesions with reduced aortic expansion (% vs WT: aortic wall thickness, 67±3%; aortic diameter, 55±3%; p<0.0001). AAA lesions from gamma-KO mice showed less disruption of elastin layers (Verhoeff-van Gieson staining) and reduced loss of medial VSMC (α-actin immunofluorescence). Inflammatory markers such as leukocyte content (CD68+ macrophages, Ly6G+ neutrophils, CD45R+ B cells, and CD3+ T lymphocytes) and the gene expression of chemokines (MCP-1, RANTES), adhesion molecules (ICAM-1), cytokines (TNFα, IFNg, IL-10) and metalloproteinases (MMP9) were all significantly lower in AAA lesions from gamma-KO mice than those from WT mice. In vitro, cross-linking of FcgammaR with fibrinogen-containing immune complexes triggers the gene expression of proinflammatory cytokines and chemokines in both VSMC and bone marrow-derived macrophages. Conclusion Activating FcgammaR participate in AAA formation by promoting inflammatory cell recruitment, and cytokine and matrix-degrading protease expression. Therefore, modulation of FcgammaR-dependent responses could be a promising therapeutic option for the treatment of AAA. Acknowledgement/Funding MINECO/FEDER (SAF2015-63696-R), ISCIII (FIS/FEDER PI17/01495), Spanish Society of Arteriosclerosis, La Caixa (HR17-00247), Conchita Rabago Foundation

Identification of key genes and pathways associated with different immune statuses of hepatitis B virus infection.

We aimed to identify key genes and pathways associated with different immune statuses of hepatitis B virus (HBV) infection. The gene expression and DNA methylation profiles were analysed in different immune statuses of HBV infection. Differentially expressed genes (DEGs) and differentially methylated genes (DMGs) were identified, followed by their functional and integrative analyses. The differential expression of IgG Fc receptors (FcγRs) in chronic HBV‐infected patients and immune cells during different stages of HBV infection was investigated. Toll‐like receptor (TLR) signalling pathway (including TLR6) and leucocyte transendothelial migration pathway (including integrin subunit beta 1) were enriched during acute infection. Key DEGs, such as FcγR Ib and FcγR Ia, and interferon‐alpha inducible protein 27 showed correlation with alanine aminotransferase levels, and they were differentially expressed between acute and immune‐tolerant phases and between immune‐tolerant and immune‐clearance phases. The integrative analysis of DNA methylation profile showed that lowly methylated and highly expressed genes, including cytotoxic T lymphocyte‐associated protein 4 and mitogen‐activated protein kinase 3 were enriched in T cell receptor signalling pathway during acute infection. Highly methylated and lowly expressed genes, such as Ras association domain family member 1 and cyclin‐dependent kinase inhibitor 2A were identified in chronic infection. Furthermore, differentially expressed FcγR Ia, FcγR IIa and FcγR IIb, CD3−CD56+CD16+ natural killer cells and CD14highCD16+ monocytes were identified between immune‐tolerant and immune‐clearance phases by experimental validation. The above genes and pathways may be used to distinguish different immune statuses of HBV infection.

Maternal alloimmune IgG causes anti-glomerular basement membrane disease in perinatal transgenic mice that express human laminin α5

Mammalian immune systems are not mature until well after birth. However, transfer of maternal IgG to the fetus and newborn usually provides immunoprotection from infectious diseases. IgG transfer occurs before birth in humans across the placenta and continues after birth across the intestine in many mammalian species, including rodents. Transfer, which is mediated by the neonatal IgG Fc receptor, occurs by transcytosis across placental syncytiotrophoblasts and intestinal epithelium. Although maternal IgG is generally beneficial, harmful maternal allo- and autoantibodies can also be transferred to the fetus/infant, resulting in serious disease. To test this we generated transgenic mice that widely express human laminin α5 in their basement membranes. When huLAMA5 transgenic males were crossed with wild-type females, there was a maternal anti-human laminin α5 immune response. Maternal IgG alloantibody crossed the yolk sac and post-natal intestine invivo and bound in bright, linear patterns to kidney glomerular basement membranes of transgenic fetuses/neonates but not those of wild-type siblings. By postnatal day 18, most transgenic mice were proteinuric, had glomerular C3 deposits and inflammatory cell infiltrates, thickened and split glomerular basement membranes, and podocyte foot process effacement. Thus, our novel model of perinatal anti-glomerular basement membrane disease may prove useful for studying pediatric glomerulopathies, formation of the fetomaternal interface, and maternal alloimmunization.

A single infection with a malaria parasite protects mice from lethal autoimmune glomerulonephritis

Abstract The progression of autoimmune diseases such as Systemic Lupus Erythematous (SLE) can be altered by environmental factors, including parasitic infections. Epidemiological studies have shown a relative absence of SLE in West Africa (areas of endemic malaria), even though SLE is highly prevalent in populations of African descent living outside of Africa. We hypothesize that malaria infections might reduce the incidence of lupus. In the present work, we assess the effect of the non-lethal Plasmodium yoelii 17XNL in a well-characterized animal model of SLE (FcγRIIB−/−). Mice deficient in the inhibitory Fc receptor for IgG (FcγRIIB) develop spontaneous disease driven by the production of anti-nuclear autoantibodies, a broad immune activation, splenomegaly, anemia and lethal glomerulonephritis with a pathology similar to human SLE. We infected FcγRIIB−/− mice at 2 months of age with P. yoelii and followed the progression of autoimmune disease for 5 months. Infection with P. yoelii increased the titer of a broad range of autoantibodies, with concomitant immune-complex deposition and complement fixation in the kidney. While lupus-prone mice succumbed from renal disease detected by protein in urine, malaria-infected lupus-prone mice did not develop proteinuria or lethal disease. Infection with the parasite did not alter tolerance mechanisms leading to lupus but instead prevented end-point tissue pathology. P. yoelii infection seems to inhibit pathogenic leukocyte infiltration without affecting the autoimmune inflammation of glomeruli. These results indicate a possible route for lupus protection that specifically targets renal function, which remains a cause of the substantial morbidity and mortality in SLE patients.

The Ligands for Human IgG and Their Effector Functions.

Activation of the humoral immune system is initiated when antibodies recognize an antigen and trigger effector functions through the interaction with Fc engaging molecules. The most abundant immunoglobulin isotype in serum is Immunoglobulin G (IgG), which is involved in many humoral immune responses, strongly interacting with effector molecules. The IgG subclass, allotype, and glycosylation pattern, among other factors, determine the interaction strength of the IgG-Fc domain with these Fc engaging molecules, and thereby the potential strength of their effector potential. The molecules responsible for the effector phase include the classical IgG-Fc receptors (FcγR), the neonatal Fc-receptor (FcRn), the Tripartite motif-containing protein 21 (TRIM21), the first component of the classical complement cascade (C1), and possibly, the Fc-receptor-like receptors (FcRL4/5). Here we provide an overview of the interactions of IgG with effector molecules and discuss how natural variation on the antibody and effector molecule side shapes the biological activities of antibodies. The increasing knowledge on the Fc-mediated effector functions of antibodies drives the development of better therapeutic antibodies for cancer immunotherapy or treatment of autoimmune diseases.

Role of the Neonatal Fc Receptor for IgG in the Pathophysiology of Lupus

Role of the Neonatal Fc Receptor for IgG in the Pathophysiology of Lupus

Oral streptococci show diversity in resistance to complement immunity.

Mechanisms underlying systemic infections by oral species of Mitis (Streptococcus mitis, Streptococcus oralis) and Sanguinis (Streptococcus gordonii, Streptococcus sanguinis) commensal streptococci are poorly understood. This study investigates profiles of susceptibility to complement-mediated host immunity in representative strains of these four species, which were isolated from oral sites or from the bloodstream. Deposition of complement opsonins (C3b/iC3b), and surface binding to C-reactive protein (CRP) and to IgG antibodies were quantified by flow cytometry in 34 strains treated with human serum (HS), and compared to rates of opsonophagocytosis by human PMN mediated by complement (CR1/3) and/or IgG Fc (FcγRII/III) receptors. S. sanguinis strains showed reduced susceptibility to complement opsonization and low binding to CRP and to IgG compared to other species. Surface levels of C3b/iC3b in S. sanguinis strains were 4.5- and 7.8-fold lower than that observed in S. gordonii and Mitis strains, respectively. Diversity in C3b/iC3b deposition was evident among Mitis species, in which C3b/iC3b deposition was significantly associated with CR/FcγR-dependent opsonophagocytosis by PMN (P<0.05). Importantly, S. gordonii and Mitis group strains isolated from systemic infections showed resistance to complement opsonization when compared to oral isolates of the respective species (P<0.05). This study establishes species-specific profiles of susceptibility to complement immunity in Mitis and Sanguinis streptococci, and indicates that strains associated with systemic infections have increased capacity to evade complement immunity. These findings highlight the need for studies identifying molecular functions involved in complement evasion in oral streptococci.

Role of FcγRs in Antibody-Based Cancer Therapy.

Monoclonal antibodies can mediate antitumor activity by multiple mechanisms. They can bind directly to tumor receptors resulting in tumor cell death, or can bind to soluble growth factors, angiogenic factors, or their cognate receptors blocking signals required for tumor cell growth or survival. Monoclonal antibodies, upon binding to tumor cell, can also engage the host's immune system to mediate immune-mediated destruction of the tumor. The Fc portion of the antibody is essential in engaging the host immune system by fixing complement resulting in complement-mediated cytotoxicity (CDC) of the tumor, or by engaging Fc receptors for IgG (FcγR) expressed by leukocytes leading to antibody-dependent cellular cytotoxicity (ADCC) or antibody-dependent cellular phagocytosis (ADCP) of tumor cells. Antibodies whose Fc portion preferentially engage activating FcγRs have shown greater inhibition of tumor growth and metastasis. Monoclonal antibodies can also stimulate the immune system by binding to targets expressed on immune cells. These antibodies may stimulate antitumor immunity by antagonizing a negative regulatory signal, agonizing a costimulatory signal, or depleting immune cells that are inhibitory. The importance of Fc:FcγR interactions in antitumor therapy for each of these mechanisms have been demonstrated in both mouse models and clinical trials and will be the focus of this chapter.

The genomic organization and expression pattern of the low-affinity Fc gamma receptors (Fc\u03b3R) in the G\xf6ttingen minipig

Safety and efficacy of therapeutic antibodies are often dependent on their interaction with Fc receptors for IgG (FcγRs). The Göttingen minipig represents a valuable species for biomedical research but its use in preclinical studies with therapeutic antibodies is hampered by the lack of knowledge about the porcine FcγRs. Genome analysis and sequencing now enabled the localization of the previously described FcγRIIIa in the orthologous location to human FCGR3A. In addition, we identified nearby the gene coding for the hitherto undescribed putative porcine FcγRIIa. The 1′241 bp long FCGR2A cDNA translates to a 274aa transmembrane protein containing an extracellular region with high similarity to human and cattle FcγRIIa. Like in cattle, the intracellular part does not contain an immunoreceptor tyrosine-based activation motif (ITAM) as in human FcγRIIa. Flow cytometry of the whole blood and single-cell RNA sequencing of peripheral blood mononuclear cells (PBMCs) of Göttingen minipigs revealed the expression profile of all porcine FcγRs which is compared to human and mouse. The new FcγRIIa is mainly expressed on platelets making the minipig a good model to study IgG-mediated platelet activation and aggregation. In contrast to humans, minipig blood monocytes were found to express inhibitory FcγRIIb that could lead to the underestimation of FcγR-mediated effects of monocytes observed in minipig studies with therapeutic antibodies.

OMIP-051 - 28-color flow cytometry panel to characterize B cells and myeloid cells.

OMIP-051 - 28-color flow cytometry panel to characterize B cells and myeloid cells.

Antibody arrests γ-herpesvirus olfactory super-infection independently of neutralization.

Protecting against persistent viruses is an unsolved challenge. The clearest example for a gamma-herpesvirus is resistance to super-infection by Murid herpesvirus-4 (MuHV-4). Most experimental infections have delivered MuHV-4 into the lungs. A more likely natural entry site is the olfactory epithelium. Its protection remains unexplored. Here, prior exposure to olfactory MuHV-4 gave good protection against super-infection. The protection was upstream of B cell infection, which occurs in lymph nodes, and showed redundancy between antibody and T cells. Adding antibody to virions that blocked heparan binding strongly reduced olfactory host entry - unlike in the lungs, opsonized virions did not reach IgG Fc receptor+ myeloid cells. However, the nasal antibody response to primary infection was too low to reduce host entry. Instead, the antibody acted downstream, reducing viral replication in the olfactory epithelium. This depended on IgG Fc receptor engagement rather than virion neutralization. Thus antibody can protect against natural γ-herpesvirus infection before it reaches B cells and independently of neutralization.

Expression of a Recombinant High Affinity IgG Fc Receptor by Engineered NK Cells as a Docking Platform for Therapeutic mAbs to Target Cancer Cells.

Anti-tumor mAbs are the most widely used and characterized cancer immunotherapy. Despite having a significant impact on some malignancies, most cancer patients respond poorly or develop resistance to this therapy. A known mechanism of action of these therapeutic mAbs is antibody-dependent cell-mediated cytotoxicity (ADCC), a key effector function of human NK cells. CD16A on human NK cells has an exclusive role in binding to tumor-bound IgG antibodies. Though CD16A is a potent activating receptor, it is also a low affinity IgG Fc receptor (FcγR) that undergoes a rapid downregulation in expression by a proteolytic process involving ADAM17 upon NK cell activation. These regulatory processes are likely to limit the efficacy of tumor-targeting therapeutic mAbs in the tumor environment. We sought to enhance NK cell binding to anti-tumor mAbs by engineering these cells with a recombinant FcγR consisting of the extracellular region of CD64, the highest affinity FcγR expressed by leukocytes, and the transmembrane and cytoplasmic regions of CD16A. This novel recombinant FcγR (CD64/16A) was expressed in the human NK cell line NK92 and in induced pluripotent stem cells from which primary NK cells were derived. CD64/16A lacked the ADAM17 cleavage region in CD16A and it was not rapidly downregulated in expression following NK cell activation during ADCC. CD64/16A on NK cells facilitated conjugation to antibody-treated tumor cells, ADCC, and cytokine production, demonstrating functional activity by its two components. Unlike NK cells expressing CD16A, CD64/16A captured soluble therapeutic mAbs and the modified NK cells mediated tumor cell killing. Hence, CD64/16A could potentially be used as a docking platform on engineered NK cells for therapeutic mAbs and IgG Fc chimeric proteins, allowing for switchable targeting elements and a novel cancer cellular therapy.

Pentraxins and Fc Receptor-Mediated Immune Responses.

C-reactive protein (CRP) is a member of the pentraxin family of proteins. These proteins are highly conserved over the course of evolution being present as far back as 250 million years ago. Mammalian pentraxins are characterized by the presence of five identical non-covalently linked subunits. Each subunit has a structurally conserved site for calcium-dependent ligand binding. The biological activities of the pentraxins established over many years include the ability to mediate opsonization for phagocytosis and complement activation. Pentraxins have an important role in protection from infection from pathogenic bacteria, and regulation of the inflammatory response. It was recognized early on that some of these functions are mediated by activation of the classical complement pathway through C1q. However, experimental evidence suggested that cellular receptors for pentraxins also play a role in phagocytosis. More recent experimental evidence indicates a direct link between pentraxins and Fc receptors. The Fc receptors were first identified as the major receptors for immunoglobulins. The avidity of the interaction between IgG complexes and Fc receptors is greatly enhanced when multivalent ligands interact with the IgG binding sites and activation of signaling pathways requires Fc receptor crosslinking. Human pentraxins bind and activate human and mouse IgG receptors, FcγRI and FcγRII, and the human IgA receptor, FcαRI. The affinities of the interactions between Fc receptors and pentraxins in solution and on cell surfaces are similar to antibody binding to low affinity Fc receptors. Crystallographic and mutagenesis studies have defined the structural features of these interactions and determined the stoichiometry of binding as one-to-one. Pentraxin aggregation or binding to multivalent ligands increases the avidity of binding and results in activation of these receptors for phagocytosis and cytokine synthesis. This review will discuss the structural and functional characteristics of pentraxin Fc receptor interactions and their implications for host defense and inflammation.