The immunology of PF4 polyanion interactions.

PURPOSE OF REVIEW: Platelet factor 4 (PF4, CXCL4), the most abundant α-granule platelet-specific chemokine, forms tetramers with an equatorial ring of high positive charge that bind to a wide range of polyanions, after which it changes conformation to expose antigenic epitopes. Antibodies directed against PF4 not only help to clear infection but can also lead to the development of thrombotic disorders such as heparin-induced thrombocytopenia (HIT) and vaccine-induced thrombocytopenia and thrombosis (VITT). This review will outline the different mechanisms through which PF4 engagement with polyanions combats infection but also contributes to the pathogenesis of inflammatory and thrombotic disease states. RECENT FINDINGS: Recent work has shown that PF4 binding to microbial polyanions may improve outcomes in infection by enhancing leukocyte-bacterial binding, tethering pathogens to neutrophil extracellular traps (NETs), decreasing the thrombotic potential of NET DNA, and modulating viral infectivity. However, PF4 binding to nucleic acids may enhance their recognition by innate immune receptors, leading to autoinflammation. Lastly, while HIT is induced by platelet activating antibodies that bind to PF4/polyanion complexes, VITT, which occurs in a small subset of patients treated with COVID-19 adenovirus vector vaccines, is characterized by prothrombotic antibodies that bind to PF4 alone. SUMMARY: Investigating the complex interplay of PF4 and polyanions may provide insights relevant to the treatment of infectious disease while also improving our understanding of the pathogenesis of thrombotic disorders driven by anti-PF4/polyanion and anti-PF4 antibodies.

Phase 1 Trial of Antibody NI006 for Depletion of Cardiac Transthyretin Amyloid.

BACKGROUND: Transthyretin amyloid (ATTR) cardiomyopathy is a progressive and fatal disease caused by misfolded transthyretin. Despite advances in slowing disease progression, there is no available treatment that depletes ATTR from the heart for the amelioration of cardiac dysfunction. NI006 is a recombinant human anti-ATTR antibody that was developed for the removal of ATTR by phagocytic immune cells. METHODS: In this phase 1, double-blind trial, we randomly assigned (in a 2:1 ratio) 40 patients with wild-type or variant ATTR cardiomyopathy and chronic heart failure to receive intravenous infusions of either NI006 or placebo every 4 weeks for 4 months. Patients were sequentially enrolled in six cohorts that received ascending doses (ranging from 0.3 to 60 mg per kilogram of body weight). After four infusions, patients were enrolled in an open-label extension phase in which they received eight infusions of NI006 with stepwise increases in the dose. The safety and pharmacokinetic profiles of NI006 were assessed, and cardiac imaging studies were performed. RESULTS: The use of NI006 was associated with no apparent drug-related serious adverse events. The pharmacokinetic profile of NI006 was consistent with that of an IgG antibody, and no antidrug antibodies were detected. At doses of at least 10 mg per kilogram, cardiac tracer uptake on scintigraphy and extracellular volume on cardiac magnetic resonance imaging, both of which are imaging-based surrogate markers of cardiac amyloid load, appeared to be reduced over a period of 12 months. The median N-terminal pro-B-type natriuretic peptide and troponin T levels also seemed to be reduced. CONCLUSIONS: In this phase 1 trial of the recombinant human antibody NI006 for the treatment of patients with ATTR cardiomyopathy and heart failure, the use of NI006 was associated with no apparent drug-related serious adverse events. (Funded by Neurimmune; NI006-101 ClinicalTrials.gov number, NCT04360434.).

Cloned antibodies from patients with HIT provide new clues to HIT pathogenesis.

Heparin-induced thrombocytopenia (HIT) is a serious adverse drug reaction characterized by antibodies that recognize platelet factor 4/heparin complexes (PF4/H) and activate platelets to create a prothrombotic state. Although a high percentage of heparin-treated patients produce antibodies to PF4/H, only a subset also makes antibodies that are platelet activating (PA). A close correlation between PA antibodies and the likelihood of experiencing HIT has been demonstrated in clinical studies, but how PA (presumptively pathogenic) and nonactivating (NA) (presumptively benign) antibodies differ from each other at the molecular level is unknown. To address this issue, we cloned 7 PA and 47 NA PF4/H-binding antibodies from 6 patients with HIT and characterized their structural and functional properties. Findings showed that PA clones differed significantly from NA clones in possessing 1 of 2 heavy chain complementarity-determining region 3 (HCDR3) motifs, RX1-2R/KX1-2R/H (RKH) and YYYYY (Y5), in an unusually long complementarity-determining region 3 (≥20 residues). Mutagenic studies showed that modification of either motif in PA clones reduced or abolished their PA activity and that appropriate amino acid substitutions in HCDR3 of NA clones can cause them to become PA. Repertoire sequencing showed that the frequency of peripheral blood IgG+ B cells possessing RKH or Y5 was significantly higher in patients with HIT than in patients without HIT given heparin, indicating expansion of B cells possessing RKH or Y5 in HIT. These findings imply that antibodies possessing RKH or Y5 are relevant to HIT pathogenesis and suggest new approaches to diagnosis and treatment of this condition.

TNF-α inhibitor ameliorates immune-related arthritis and pneumonitis in humanized mice.

Objectives: This study aimed at establishing a mouse model of immune-related adverse in humanized BALB/c-hPD1/hCTLA4 mice to investigate their potential pathogenesis and explore therapeutic targets for immune-related arthritis and pneumonitis. Methods: Humanized BALB/c-hPD1/hCTLA4 mice were injected with vehicle or collagen-specific antibodies (CA) and immune checkpoint inhibitors (ICI, ipilimumab, anti-human CTLA-4; and nivolumab, anti-human PD-1), and some mice were treated with anti-TNF-α antibody, leading to the control, collagen antibody-induced arthritis (CAIA), CAIA+ICI and treatment groups. The severity of clinical arthritis and pneumonitis in mice was monitored longitudinally and the pathological changes in the joints and lungs were histologically analyzed and the contents of lung hydroxyproline were measured. The frequency of different subsets of T cells was analyzed by flow cytometry and multiplex immunofluorescency. Results: Compared with the control, the ICI group of mice developed the delayed onset of moderate degrees of arthritis while the CAIA+ICI group of mice exhibited the early onset of severe arthritis. Treatment with ICI caused severe pneumonitis, especially in the mice with CA. Flow cytometry analysis indicated a significantly higher frequency of splenic TNF-α+CD4+ and TNF-α+CD8+ T cells, but not other subsets of T cells tested, in the CAIA+ICI group of mice, relative to that in other groups of mice. Treatment with anti-TNF-α significantly mitigated the severity of arthritis and pneumonitis as well as deposition of collagen in lung of mice. The treatment also decreased the frequency of TNF-α+CD4+ and TNF-α+CD8+ T cells as well as effector memory T cells in the periphery lymph orangs and lungs of mice. Conclusions: We successfully established a humanized mouse model of ICI-related severe arthritis and pneumonitis with a higher frequency of TNF-α+ T cells, which were significantly mitigated by anti-TNF-α treatment. Conceptually, ICI treatment can induce multiple autoimmune-like diseases in autoimmune-prone individuals and TNF-α+ T cells may be therapeutic targets for intervention of immune-related arthritis and pneumonitis.

Natural history of PF4 antibodies in vaccine-induced immune thrombocytopenia and thrombosis.

The COVID-19 pandemic has resulted in the rapid development of a range of vaccines against SARS-CoV-2. Vaccine-induced immune thrombocytopenia and thrombosis (VITT) is a rare but life-threatening complication of primarily adenoviral-based vaccines associated with the presence of antibodies to a PF4/polyanion neoepitope and measured by using enzyme-linked immunosorbent assays. Presented are serial anti-PF4/polyanion antibody, platelet, and D-dimer measurements in a large cohort of patients and their relation to relapse. Overall, 51% of patients using the Stago assay had persistently positive anti-PF4/polyanion levels 100 days' postdiagnosis, whereas 94% of patients monitored by using the Immucor assay remain positive. The median duration of positivity of the PF4 assay is 87 days, with 72% of patients remaining positive after a median follow-up of 105 days. The use of plasma exchange seemed to reduce anti-PF4/polyanion levels and increase platelet counts in the acute setting more rapidly than other therapies. The rate of relapse in this study was 12.6%, with all relapsed cases exhibiting persistently positive PF4 antibodies and falling platelet counts. Only one patient had extension of their thrombosis. Overall, despite the persistence of PF4 antibodies in 72% of patients, the rate of relapse was low and did not seem to result in recrudescence of the aggressive clinical picture seen at index presentation. Monitoring of these patients in the UK cohort is ongoing and will aid in definition of the natural history of this novel condition.

Safety, pharmacokinetics, and antitumor activity of the anti-CEACAM5-DM4 antibody-drug conjugate tusamitamab ravtansine (SAR408701) in patients with advanced solid tumors: first-in-human dose-escalation study.

BACKGROUND: Tusamitamab ravtansine (SAR408701) is an antibody-drug conjugate composed of a humanized monoclonal antibody that binds carcinoembryonic antigen-related cell adhesion molecule-5 (CEACAM5) and a cytotoxic maytansinoid that selectively targets CEACAM5-expressing tumor cells. In this phase I dose-escalation study, we evaluated the safety, pharmacokinetics, and preliminary antitumor activity of tusamitamab ravtansine in patients with solid tumors. PATIENTS AND METHODS: Eligible patients were aged ≥18 years, had locally advanced/metastatic solid tumors that expressed or were likely to express CEACAM5, and had an Eastern Cooperative Oncology Group Performance Status of 0 or 1. Patients were treated with ascending doses of tusamitamab ravtansine intravenously every 2 weeks (Q2W). The first three dose levels (5, 10, and 20 mg/m2) were evaluated using an accelerated escalation protocol, after which an adaptive Bayesian procedure was used. The primary endpoint was the incidence of dose-limiting toxicities (DLTs) during the first two cycles, graded using National Cancer Institute Common Terminology Criteria for Adverse Events (NCI CTCAE) v4.03 criteria. RESULTS: Thirty-one patients received tusamitamab ravtansine (range 5-150 mg/m2). The DLT population comprised 28 patients; DLTs (reversible grade 3 microcystic keratopathy) occurred in three of eight patients treated with tusamitamab ravtansine 120 mg/m2 and in two of three patients treated with 150 mg/m2. The maximum tolerated dose was identified as 100 mg/m2. Twenty-two patients (71%) experienced ≥1 treatment-related treatment-emergent adverse event (TEAE), seven patients (22.6%) experienced ≥1 treatment-related grade ≥3 TEAE, and three patients (9.7%) discontinued treatment due to TEAEs. The most common TEAEs were asthenia, decreased appetite, keratopathy, and nausea. Three patients had confirmed partial responses. The mean plasma exposure of tusamitamab ravtansine increased in a dose-proportional manner from 10 to 150 mg/m2. CONCLUSIONS: Tusamitamab ravtansine had a favorable safety profile with reversible, dose-related keratopathy as the DLT. Based on the overall safety profile, pharmacokinetic data, and Bayesian model recommendations, the maximum tolerated dose of tusamitamab ravtansine was defined as 100 mg/m2 Q2W.

Bortezomib alleviates antibody-mediated rejection in kidney transplantation by facilitating Atg5 expression.

Antibody-mediated rejection (AMR) is one of the most dominant mechanisms responsible for the loss of kidney grafts. Previous researches have shown that donor-specific antibodies (DSAs) are the major mediators of AMR. In order to prolong the survival time of grafts, it is vital to reduce the incidence of AMR and inhibit the generation of DSAs. We established an animal model of AMR by performing kidney transplantation in pre-sensitized rats. Then, we investigated the effect of bortezomib (BTZ) on AMR. We found that BTZ could reduce the serum level of DSAs and alleviate post-transplantation inflammation in peritubular capillaries (PTCs) and glomeruli, which was demonstrated by the reduction of C4d and IgG deposition in PTCs, and the reduced number of B cell and plasma cell in peripheral blood and the transplanted kidney (p < 0.05). Our results also suggested that BTZ increased the number of regulatory T cell (Treg) and significantly reduced the proportion of T helper (Th17) cell (p < 0.05). Besides, BTZ induced the significant upregulation of anti-inflammatory cytokines but downregulated pro-inflammatory cytokines (p < 0.05). After dealing with Atg5 siRNA-lentivirus, the effect of BTZ alleviating AMR was reversed and Th17/Treg proportions were also significantly modulated. Collectively, these findings show that BTZ slows down the process of AMR and Atg5 may be the key mechanism. Furthermore, Atg5 silencing results may be demonstrated that Atg5 alleviated AMR by modulating the ratio of Th17/Treg.

REV-ERBα Agonist GSK4112 attenuates Fas-induced Acute Hepatic Damage in Mice.

Fas-induced apoptosis is a central mechanism of hepatocyte damage during acute and chronic hepatic disorders. Increasing evidence suggests that circadian clock plays critical roles in the regulation of cell fates. In the present study, the potential significance of REV-ERBα, a core ingredient of circadian clock, in Fas-induced acute liver injury has been investigated. The anti-Fas antibody Jo2 was injected intraperitoneally in mice to induce acute liver injury and the REV-ERBα agonist GSK4112 was administered. The results indicated that treatment of GSK4112 decreased the level of plasma ALT and AST, attenuated the liver histological changes, and promoted the survival rate in Jo2-insulted mice. Treatment with GSK4112 also downregulated the activities of caspase-3 and caspase-8, suppressed hepatocyte apoptosis. In addition, treatment with GSK4112 decreased the level of Fas and enhanced the phosphorylation of Akt. In conclusion, treatment with GSK4112 alleviated Fas-induced apoptotic liver damage in mice, suggesting that REV-ERBα agonist might have potential value in pharmacological intervention of Fas-associated liver injury.

Neutrophil-suppressive activity over T-cell proliferation and fungal clearance in a murine model of Fonsecaea pedrosoi infection.

Neutrophils are essential to control several fungal infections. These cells are commonly known for their pro-inflammatory activities. However, some studies have demonstrated the anti-inflammatory properties of neutrophils during certain infectious diseases, culminating in the inhibition of T cell proliferation. Chromoblastomycosis (CBM) is a deep and progressive mycosis that affects thousands of people worldwide. Although neutrophil infiltrates are observed in the lesion histopathology, the fungus can overtake the immune system response and destroy the host-infected tissue. The present study demonstrated that neutropenic animals had an increase in the IL-6 production in the spleen and liver, followed by a lower fungal burden in these organs up to 14 days of infection. Neutropenic animals also showed a lower F. pedrosoi-specific antibody production 14-days post infection and higher T-cell proliferation in the in vitro experiments after stimulation with F. pedrosoi-purified proteins. Taken together, our results suggest that the presence of regulatory neutrophils in the mouse model of F. pedrosoi infection could act favoring the spread of the fungus and the chronicity of the infection. These findings shed light on the CBM treatment, which might target neutrophil polarization as a new therapy approach to treat CBM lesions.

Pathogenic Mechanisms of Vaccine-Induced Immune Thrombotic Thrombocytopenia in People Receiving Anti-COVID-19 Adenoviral-Based Vaccines: A Proposal.

VITT is a rare, life-threatening syndrome characterized by thrombotic symptoms in combination with thrombocytopenia, which may occur in individuals receiving the first administration of adenoviral non replicating vectors (AVV) anti Covid19 vaccines. Vaccine-induced immune thrombotic thrombocytopenia (VITT) is characterized by high levels of serum IgG that bind PF4/polyanion complexes, thus triggering platelet activation. Therefore, identification of the fine pathophysiological mechanism by which vaccine components trigger platelet activation is mandatory. Herein, we propose a multistep mechanism involving both the AVV and the neo-synthetized Spike protein. The former can: i) spread rapidly into blood stream, ii), promote the early production of high levels of IL-6, iii) interact with erythrocytes, platelets, mast cells and endothelia, iv) favor the presence of extracellular DNA at the site of injection, v) activate platelets and mast cells to release PF4 and heparin. Moreover, AVV infection of mast cells may trigger aberrant inflammatory and immune responses in people affected by the mast cell activation syndrome (MCAS). The pre-existence of natural antibodies binding PF4/heparin complexes may amplify platelet activation and thrombotic events. Finally, neosynthesized Covid 19 Spike protein interacting with its ACE2 receptor on endothelia, platelets and leucocyte may trigger further thrombotic events unleashing the WITT syndrome.

Mannose Receptor Mediates the Activation of Chitooligosaccharides on Blunt Snout Bream (Megalobrama amblycephala) Macrophages.

Chitooligosaccharide (COS) is an important immune enhancer and has been proven to have a variety of biological activities. Our previous research has established an M1 polarization mode by COS in blunt snout bream (Megalobrama amblycephala) macrophages, but the mechanism of COS activation of blunt snout bream macrophages remains unclear. In this study, we further explored the internalization mechanism and signal transduction pathway of chitooligosaccharide hexamer (COS6) in blunt snout bream macrophages. The results showed that mannose receptor C-type lectin-like domain 4-8 of M. amblycephala (MaMR CTLD4-8) could recognize and bind to COS6 and mediate COS6 into macrophages by both clathrin-dependent and caveolin-dependent pathways. In the inflammatory response of macrophages activated by COS6, the gene expression of tumor necrosis factor (TNF)-α, interleukin (IL)-1ß, and nitric oxide synthase 2 (NOS2) was significantly inhibited after MaMR CTLD4-8-specific antibody blockade. However, even if it was blocked, the expression of these inflammation-related genes was still relatively upregulated, which suggested that there are other receptors involved in immune regulation. Further studies indicated that MaMR CTLD4-8 and Toll-like receptor 4 (TLR4) cooperated to regulate the pro-inflammatory response of macrophages caused by COS6. Taken together, these results revealed that mannose receptor (MR) CTLD4-8 is indispensable in the process of recognition, binding, internalization, and immunoregulation of COS in macrophages of blunt snout bream.

Brain Disposition of Antibody-Based Therapeutics: Dogma, Approaches and Perspectives.

Due to their high specificity, monoclonal antibodies have been widely investigated for their application in drug delivery to the central nervous system (CNS) for the treatment of neurological diseases such as stroke, Alzheimer's, and Parkinson's disease. Research in the past few decades has revealed that one of the biggest challenges in the development of antibodies for drug delivery to the CNS is the presence of blood-brain barrier (BBB), which acts to restrict drug delivery and contributes to the limited uptake (0.1-0.2% of injected dose) of circulating antibodies into the brain. This article reviews the various methods currently used for antibody delivery to the CNS at the preclinical stage of development and the underlying mechanisms of BBB penetration. It also describes efforts to improve or modulate the physicochemical and biochemical properties of antibodies (e.g., charge, Fc receptor binding affinity, and target affinity), to adapt their pharmacokinetics (PK), and to influence their distribution and disposition into the brain. Finally, a distinction is made between approaches that seek to modify BBB permeability and those that use a physiological approach or antibody engineering to increase uptake in the CNS. Although there are currently inherent difficulties in developing safe and efficacious antibodies that will cross the BBB, the future prospects of brain-targeted delivery of antibody-based agents are believed to be excellent.

Anti-platelet factor 4 antibodies causing VITT do not cross-react with SARS-CoV-2 spike protein.

Vaccine-induced immune thrombotic thrombocytopenia (VITT) is a severe adverse effect of ChAdOx1 nCoV-19 COVID-19 vaccine (Vaxzevria) and Janssen Ad26.COV2.S COVID-19 vaccine, and it is associated with unusual thrombosis. VITT is caused by anti-platelet factor 4 (PF4) antibodies activating platelets through their FcγRIIa receptors. Antibodies that activate platelets through FcγRIIa receptors have also been identified in patients with COVID-19. These findings raise concern that vaccination-induced antibodies against anti-SARS-CoV-2 spike protein cause thrombosis by cross-reacting with PF4. Immunogenic epitopes of PF4 and SARS-CoV-2 spike protein were compared using in silico prediction tools and 3D modeling. The SARS-CoV-2 spike protein and PF4 share at least 1 similar epitope. Reactivity of purified anti-PF4 antibodies from patients with VITT was tested against recombinant SARS-CoV-2 spike protein. However, none of the affinity-purified anti-PF4 antibodies from 14 patients with VITT cross-reacted with SARS-CoV-2 spike protein. Sera from 222 polymerase chain reaction-confirmed patients with COVID-19 from 5 European centers were tested by PF4-heparin enzyme-linked immunosorbent assays and PF4-dependent platelet activation assays. We found anti-PF4 antibodies in sera from 19 (8.6%) of 222 patients with COVID-19. However, only 4 showed weak to moderate platelet activation in the presence of PF4, and none of those patients developed thrombotic complications. Among 10 (4.5%) of 222 patients who had COVID-19 with thrombosis, none showed PF4-dependent platelet-activating antibodies. In conclusion, antibodies against PF4 induced by vaccination do not cross-react with the SARS-CoV-2 spike protein, indicating that the intended vaccine-induced immune response against SARS-CoV-2 spike protein is not the trigger of VITT. PF4-reactive antibodies found in patients with COVID-19 in this study were not associated with thrombotic complications.

Acute Hemolysis and Heme Suppress Anti-CD40 Antibody-Induced Necro-Inflammatory Liver Disease.

Clearance of red blood cells and hemoproteins is a key metabolic function of macrophages during hemolytic disorders and following tissue injury. Through this archetypical phagocytic function, heme is detoxified and iron is recycled to support erythropoiesis. Reciprocal interaction of heme metabolism and inflammatory macrophage functions may modify disease outcomes in a broad range of clinical conditions. We hypothesized that acute hemolysis and heme induce acute anti-inflammatory signals in liver macrophages. Using a macrophage-driven model of sterile liver inflammation, we showed that phenylhydrazine (PHZ)-mediated acute erythrophagocytosis blocked the anti-CD40 antibody-induced pathway of macrophage activation. This process attenuated the inflammatory cytokine release syndrome and necrotizing hepatitis induced by anti-CD40 antibody treatment of mice. We further established that administration of heme-albumin complexes specifically delivered heme to liver macrophages and replicated the anti-inflammatory effect of hemolysis. The anti-inflammatory heme-signal was induced in macrophages by an increased intracellular concentration of the porphyrin independently of iron. Overall, our work suggests that induction of heme-signaling strongly suppresses inflammatory macrophage function, providing protection against sterile liver inflammation.

Efficacy and Safety of Divaza for the Correction of Oxidative Disturbances in Patients with Cerebral Atherosclerosis: A Randomized Controlled Trial.

OBJECTIVE: The objective of this study was to determine if Divaza, a drug with nootropic and antioxidant effects, was safe and effective for the correction of oxidative disturbances and to stabilize cognitive impairment in patients with cerebral atherosclerosis. STUDY DESIGN: The study design consisted of a 12-week multicenter, randomized, double-blind, placebo-controlled, prospective trial in parallel groups. SETTING: The setting in which the study was conducted comprised 10 clinical centers across the Russian Federation. INTERVENTIONS: Patients were randomized into 2 groups and instructed to take either 2 tablets of the study drug or a placebo 3 times per day in conjunction with basic therapy. OUTCOMES: The primary outcome was a change in the average endogenous antioxidant potential after the completion of the study. The blood indicators of the oxidative stress (OS) were analyzed at the baseline and then after 12 weeks of therapy using iron-induced chemiluminescence analysis. The Montreal cognitive assessment test was used as a secondary outcome measure to evaluate cognitive impairment at the end of the study. RESULTS: 124 outpatients with a mean age of 60.7 ± 7.6 years were enrolled and randomly assigned to receive Divaza (n = 65) or a placebo (n = 59). An improvement of cognitive function was observed in all patients of the Divaza group at the end of the treatment; this was significantly better than the placebo group (100 [100] vs. 89.5 [89.1]%, respectively, p = 0.0272 [p = 0.0128]). The administration of Divaza restored the activity of the endogenous antioxidant system. The change in the average level of lipoprotein resistance to oxidation after 12 weeks of therapy, compared to the baseline, was significantly higher in the Divaza group (14.8 ± 14.7 [14.8 ± 14.7] seconds latent period vs. 6.4 ± 16.9 [6.9 ± 16.7] seconds in the placebo group (p = 0.007 [p = 0.0107]). CONCLUSIONS: Divaza is a safe and effective therapeutic option for attenuating OS and recovery of cognitive impairment in patients with cerebral atherosclerosis.

Immunological analysis of the murine anti-CD3-induced cytokine release syndrome model and therapeutic efficacy of anti-cytokine antibodies.

The aberrant release of inflammatory mediators often referred to as a cytokine storm or cytokine release syndrome (CRS), is a common and sometimes fatal complication in acute infectious diseases including Ebola, dengue, COVID-19, and influenza. Fatal CRS occurrences have also plagued the development of highly promising cancer therapies based on T-cell engagers and chimeric antigen receptor (CAR) T cells. CRS is intimately linked with dysregulated and excessive cytokine release, including IFN-γ, TNF-α, IL 1, IL-6, and IL-10, resulting in a systemic inflammatory response leading to multiple organ failure. Here, we show that mice intravenously administered the agonistic hamster anti-mouse CD3ε monoclonal antibody 145-2C11 develop clinical and laboratory manifestations seen in patients afflicted with CRS, including body weight loss, hepatosplenomegaly, thrombocytopenia, increased vascular permeability, lung inflammation, and hypercytokinemia. Blood cytokine levels and gene expression analysis from lung, liver, and spleen demonstrated a hierarchy of inflammatory cytokine production and infiltrating immune cells with differentiating organ-dependent kinetics. IL-2, IFN-γ, TNF-α, and IL-6 up-regulation preceded clinical signs of CRS. The co-treatment of mice with a neutralizing anti-cytokine antibody cocktail transiently improved early clinical and laboratory features of CRS. We discuss the predictive use of this model in the context of new anti-cytokine strategies to treat human CRS.

Targeting the Notch Signaling Pathway in Chronic Inflammatory Diseases.

The Notch signaling pathway regulates developmental cell-fate decisions and has recently also been linked to inflammatory diseases. Although therapies targeting Notch signaling in inflammation in theory are attractive, their design and implementation have proven difficult, at least partly due to the broad involvement of Notch signaling in regenerative and homeostatic processes. In this review, we summarize the supporting role of Notch signaling in various inflammation-driven diseases, and highlight efforts to intervene with this pathway by targeting Notch ligands and/or receptors with distinct therapeutic strategies, including antibody designs. We discuss this in light of lessons learned from Notch targeting in cancer treatment. Finally, we elaborate on the impact of individual Notch members in inflammation, which may lay the foundation for development of therapeutic strategies in chronic inflammatory diseases.