Complement-mediated Disorders Research Articles

How complement inhibitors are transforming the management of complement-mediated disorders.

How complement inhibitors are transforming the management of complement-mediated disorders.

Usefulness and analytical performances of complement multiplex assay for measuring complement biomarkers in plasma

IntroductionThe complement system is involved in numerous diseases, through diverse mechanisms and degree of activation. With the emergence of complement targeting therapeutic, simple and accessible tools to evaluate the extent of complement activation are strongly needed. MethodsWe evaluated two multiplex panels, measuring complement activation fragments (C4a, C3a, C5a, Bb, Ba, sC5b9) and intact components or regulators (C1q, C2, C3, C4, C5, FD, FP, FH, FI). The specificity of each measurement was assessed by using complement proteins depleted sera and plasma collected from patients with complement deficiencies. Normal values distribution was estimated using 124 plasma samples from healthy donors and complement activation profile was assessed in plasma collected from 31 patients with various complement-mediated disorders. ResultsWe observed good inter-assay variation. All tested protein deficiencies were accurately detected. We established assay-specific reference values for each analyte. Except for C3, C4 and C4a, the majority of the measurements were in good agreement with references methods or published data. ConclusionOur study substantiates the utility of the Complement Multiplex assay as a tool for measuring complement activation and deficiencies. Quantifying complement cleavage fragments in patients exhibiting classical or alternative pathway activation allowed evaluating the activation state of the whole cascade.

Characterization of the bispecific VHH antibody gefurulimab (ALXN1720) targeting complement component C5, and designed for low volume subcutaneous administration

(248/300)The bispecific antibody gefurulimab (also known as ALXN1720) was developed to provide patients with a subcutaneous treatment option for chronic disorders involving activation of the terminal complement pathway. Gefurulimab blocks the enzymatic cleavage of complement component 5 (C5) into the biologically active components C5a and C5b, which triggers activation of the terminal complement cascade. Heavy-chain variable region antigen-binding fragment (VHH) antibodies targeting C5 and human serum albumin (HSA) were isolated from llama immune-based libraries and humanized. Gefurulimab comprises an N-terminal albumin-binding VHH connected to a C-terminal C5-binding VHH via a flexible linker. The purified bispecific VHH antibody has the expected exact size by mass spectrometry and can be formulated at greater than 100mg/mL. Gefurulimab binds tightly to human C5 and HSA with dissociation rate constants at pH 7.4 of 54 pM and 0.9nM, respectively, and cross-reacts with C5 and serum albumin from cynomolgus monkeys. Gefurulimab can associate with C5 and albumin simultaneously, and potently inhibits the terminal complement activity from human serum initiated by any of the three complement pathways in Wieslab assays. Electron microscopy and X-ray crystallography revealed that the isolated C5-binding VHH recognizes the macroglobulin (MG) 4 and MG5 domains of the antigen and thereby is suggested to sterically prevent C5 binding to its activating convertase. Gefurulimab also inhibits complement activity supported by the rare C5 allelic variant featuring an R885H substitution in the MG7 domain. Taken together, these data suggest that gefurulimab may be a promising candidate for the potential treatment of complement-mediated disorders

Subclinical microvascular changes in ANCA-vasculitides: the role of optical coherence tomography angiography and nailfold capillaroscopy in the detection of disease-related damage

BackgroundBoth cardiovascular and complement-mediated disorders might lead to microvascular damages in anti-neutrophil cytoplasm autoantibodies (ANCA)-associated vasculitides (AAV). We aimed at investigating, for the first time, subclinical microvascular abnormalities with non-invasive techniques in AAV patients by analyzing both retinal and nailfold capillary changes. Retinal plexi were investigated using optical coherence tomography angiography (OCT-A), while nailfold capillary changes by video-capillaroscopy (NVC). Potential correlations between microvessels’ abnormalities and disease damage were also explored.MethodsAn observational study was conducted on consecutive patients who met the inclusion criteria of defined diagnosis of eosinophilic granulomatosis with polyangiitis (EGPA), granulomatosis with polyangiitis (GPA), and microscopic polyangiitis (MPA), age ≥ 18 ≤ 75 yrs, and no ophthalmological disorders. Disease activity was assessed by Birmingham Vasculitis Activity Score (BVAS), damage by Vasculitis Damage Index (VDI), and poorer prognosis by the Five Factor Score (FFS). Quantitative analysis of vessel density (VD) was performed by OCT-A in both superficial and deep capillary plexi. Figures and detailed analysis from NVC were performed for all subjects in the study.ResultsIncluded AAV patients (n = 23) were compared with 20 age/sex-matched healthy controls (HC). Retinal VD in superficial whole and parafoveal plexi resulted significantly decreased in AAV compared to HC (P = 0.02 and P = 0.01, respectively). Furthermore, deep whole and parafoveal vessel density was strongly reduced in AAV than HC (P ≤ 0.0001 for both). In AAV patients, significant inverse correlations occurred between VDI and OCTA-VD in both superficial (parafoveal, P = 0.03) and deep plexi (whole, P = 0.003, and parafoveal P = 0.02). Non-specific NVC pattern abnormalities occurred in 82% of AAV patients with a similar prevalence (75%) in HC. In AAV, common abnormalities were edema and tortuosity in a comparable distribution with HC. Correlations between NVC changes and OCT-A abnormalities have not been described.ConclusionSubclinical microvascular retinal changes occur in patients with AAV and correlate with the disease-related damage. In this context, the OCT-A can represent a useful tool in the early detection of vascular damage. AAV patients present microvascular abnormalities at NVC, whose clinical relevance requires further studies.

A Case of Neisseria sicca Bacteremia Due to Eculizumab Therapy

Eculizumab is a monoclonal antibody indicated for the treatment of complement-mediated disorders. Despite its utility, eculizumab increases risk for infection with Neisseria species, owing to membrane attack complex inhibition. Although this naturally evokes thought of infection with N meningitidis, other Neisseria species also benefit from membrane attack complex disruption. N sicca is a commensal organism that is typically considered nonpathogenic. Here, we describe a case of N sicca bacteremia in a patient receiving eculizumab for atypical hemolytic uremic syndrome and seek to elucidate limitations to current prophylactic recommendations for patients receiving anticomplement therapy.

5610823 TRIAL IN PROGRESS: THE RANDOMIZED, DOUBLE-BLIND, PLACEBO-CONTROLLED PHASE 1B CROSSWALK-A TRIAL EVALUATING THE SAFETY OF CROVALIMAB FOR THE MANAGEMENT OF ACUTE UNCOMPLICATED VASO-OCCLUSIVE EPISODES (VOES) IN PATIENTS (PTS) WITH SICKLE CELL DISEASE (SCD)

Background: SCD is a group of autosomal recessive red blood cell (RBC) disorders caused by a single point mutation in the β-globin gene, resulting in the production of hemoglobin S (HbS). HbS polymerizes within RBCs under certain conditions, leading to the distortion of the RBC membrane and generation of dense and sickle RBCs. These pathologic RBCs contribute to microvascular occlusions, which may present as acute, painful episodes called VOEs. Although most VOEs are managed at home, >70% of emergency department visits and >90% of hospitalizations for SCD are VOE-related.1,2 Additionally, acute chest syndrome is one of the most severe complications of VOEs and is a leading cause of mortality.3,4 As there are no targeted therapies for the management of VOEs, treatment is currently limited to pain management, blood exchange transfusion (with the risk of complications), and other supportive care, which represents a significant unmet medical need. Overall, accumulating nonclinical data suggest a multimodal role for complement dysregulation in the pathophysiology of SCD including vaso-occlusion, hemolysis, inflammation, thrombogenicity, endothelial activation, and end-organ damage.5 Complement pathway activation has been described in pts with SCD at baseline, in acute pain crises, and in pts with delayed hemolytic transfusion reaction. The complement pathway can be targeted with crovalimab, which is a novel, engineered, anti-complement C5 monoclonal antibody. In a Phase 1/2 study in patients with paroxysmal nocturnal hemoglobinuria, a complement-mediated disorder, crovalimab showed rapid and sustained complement inhibition with promising efficacy and safety.6 Aims: CROSSWALK-a (NCT04912869) is a randomized, double-blind, placebo-controlled, Phase 1b study evaluating the safety of crovalimab in managing acute uncomplicated VOEs in pts with SCD. Methods: Pts aged 12 to 55 years, weighing ≥40 kg, and with a confirmed diagnosis of SCD homozygous hemoglobin S (HbSS) or sickle cell β0 thalassemia (HbSβ0) will be enrolled (Figure). Pts must present with an acute uncomplicated VOE, requiring hospitalization and treatment with parenteral opioid analgesics. Vaccinations against Neisseria meningitidis, Hemophilus influenzae type B, and Streptococcus pneumoniae must be current. Eligible pts will be randomized 2:1 to receive either a single intravenous weight-based tiered dose of crovalimab or placebo. All pts will continue with pain management and other supportive care for their VOE, and may continue concurrent SCD-directed therapies. Pts will be followed during hospitalization until discharge and will also be followed post-discharge during an observational period. The study duration for an individual pt, including the hospitalization and observational periods, will be 12 weeks. The primary objective is to evaluate the incidence and severity of adverse events according to National Cancer Institute Common Terminology Criteria for Adverse Events, Version 5.0, incidence and severity of infusion-related reactions and hypersensitivity, and change from baseline in targeted vital signs and clinical laboratory test results. Efficacy, pharmacokinetic, pharmacodynamic, immunogenicity, and exploratory biomarker endpoints will also be evaluated. Protocol updates are in progress. Results: CROSSWALK-a is scheduled to be completed in September 2023. Summary: CROSSWALK-a is enrolling pts with SCD in five countries.

5612539 TRIAL IN PROGRESS: THE RANDOMIZED, DOUBLE-BLIND, PLACEBO-CONTROLLED PHASE 2A CROSSWALK-C TRIAL EVALUATING THE EFFICACY OF CROVALIMAB AS ADJUNCT TREATMENT IN THE PREVENTION OF VASO-OCCLUSIVE EPISODES (VOES) IN PATIENTS (PTS) WITH SICKLE CELL DISEASE (SCD)

Background: SCD is a group of autosomal recessive red blood cell (RBC) disorders caused by a single point mutation in the β-globin gene, with either homozygous inheritance or heterozygous co-inheritance with other pathogenic variants of the β-globin gene. Hemoglobin S, produced as a result of this point mutation, polymerizes within RBCs under certain conditions, distorting them and generating dense and sickle RBCs. These pathologic RBCs contribute to microvascular occlusions, which may present as acute painful episodes called VOEs. Pts with SCD may also have severe chronic anemia, chronic pain, immune dysfunction, and progressive multi-organ damage. Current therapies for SCD include hydroxyurea, as well as newer treatments such as L-glutamine, crizanlizumab, and voxelotor. Despite these treatments, considerable morbidity and mortality among pts with SCD represents a significant unmet medical need. Complement pathway activation has been reported in pts with SCD at baseline, in acute pain crises, and in delayed hemolytic transfusion reaction. Accumulating nonclinical data suggest the potential multimodal role for complement dysregulation in the pathophysiology of SCD, including vaso-occlusion, hemolysis, inflammation, thrombogenicity, endothelial activation, and end-organ damage.1 The complement pathway can be targeted with crovalimab, a novel anti-C5 monoclonal antibody that allows for small-volume subcutaneous (SC) self-injection. In a Phase 1/2 paroxysmal nocturnal hemoglobinuria (a complement-mediated disorder) study, crovalimab showed rapid and sustained complement inhibition with promising efficacy and safety.2 Aim: CROSSWALK-c (NCT05075824) is a randomized, double-blind, placebo-controlled, Phase 2a study evaluating the efficacy and safety of crovalimab as adjunct therapy in preventing VOEs in pts with SCD. Methods: Pts aged 12 to 55 years, weighing ≥40 kg, with a confirmed diagnosis of SCD, homozygous hemoglobin S (HbSS) or sickle cell β0 thalassemia (HbSβ0), and presenting with 2 to 10 VOEs are eligible, including pts on concurrent SCD-directed therapies. Vaccinations against Neisseria meningitidis, Haemophilus influenzae type B, and Streptococcus pneumoniae are required. Pts with a history of hematopoietic stem cell transplant will be excluded. Eligible pts will be randomized 1:1 to the crovalimab or placebo arms (Figure). An initial intravenous loading dose of crovalimab or placebo will be given on Week 1, Day 1, followed by SC dose on Week 1, Day 2, and then weekly SC doses on Weeks 2–4. Starting Week 5, a maintenance dose will be given every 4 weeks for 48 weeks. All study treatment will be given according to a weight-based tiered dosing schedule (pts weighing ≥ 40 kg to < 100 kg and pts ≥ 100 kg). The primary objective is to evaluate the efficacy of crovalimab vs placebo based on the annualized rate of medical facility VOEs. Key secondary efficacy objectives are the annualized rate of acute chest syndrome, the annualized rate of home VOE, and change in urinary albumin–creatinine ratio, tricuspid regurgitant jet velocity, and Patient-Reported Outcomes Measurement Information System (PROMIS)-Fatigue score in adults, from baseline to Week 49. Safety, pharmacokinetic, immunogenicity, and exploratory biomarker objectives will also be evaluated. Protocol updates are in progress. Results: Primary results are expected in July 2024. Summary: CROSSWALK-c is enrolling pts with SCD and chronic VOEs in six countries.

Identification of potent siRNA targeting complement C5 and its robust activity in pre-clinical models of myasthenia gravis and collagen-induced arthritis

Complement component 5 (C5), an important molecule in the complement cascade, blockade by antibodies shows clinical efficacy in treating complement-mediated disorders. However, insufficient blockading induced by single-nucleotide polymorphisms in the C5 protein or frequent development of "breakthrough" intravascular hemolysis in patients with paroxysmal nocturnal hemoglobinuria treated with eculizumab have been reported. Herein, we developed a lipid nanoparticle (LNP)-formulated siRNA targeting C5 that was efficiently delivered to the liver and silenced C5 expression. We identified a potent C5-siRNA with an invitro IC50 of 420 pM and invivo ED50 of 0.017mg/kg following a single administration. Single or repeated administrations of the LNP-formulated C5-siRNA allowed robust and durable suppression of liver C5 expression in mice. Complement C5 silencing ameliorated C5b-dependent anti-acetylcholine receptor antibody-induced myasthenia gravis and C5a-dependent collagen-induced arthritis symptoms. Similarly, in nonhuman primates, a single administration of C5-siRNA/LNP-induced dose-dependent plasma C5 suppression and concomitantly inhibited serum complement activity; complement activity recovered to the pre-treatment levels at 65days post administration, thus indicating that the complement activity can be controlled for a specific period. Our findings provide the foundation for further developing C5-siRNA delivered via LNPs as a potential therapeutic for complement-mediated diseases.

Soluble Complement Receptor 1 Therapeutics

Human Complement Receptor 1 (CR1/CD35) is a potent negative regulator of the complement system. Its mechanism of action is through interaction with the complement activation fragments, C3b and C4b to mediate decay acceleration of the C3 and C5 convertase complexes as well as cleavage of both ligands into inactive fragments via cofactor activity. The result is inhibition of the classical, lectin, and alternative complement pathways. This article will focus on recombinant soluble forms of CR1 that have been generated as potential therapeutics for complement-mediated disorders. Specifically, we will review and contrast the in vitro and in vivo properties of: sCR1 (BRL55730/TP10/CDX-1135), the soluble full-length extracellular domain of human CR1; sCR1-sLex (TP20), a glyco-engineered version of sCR1 additionally targeted to activated endothelium; APT070 (Mirococept), a CR1 fragment conjugated to a myristoylated peptide to enhance tissue targeting; and CSL040, a soluble truncated version of the CR1 extracellular domain which exhibits altered potency and pharmacokinetic properties as compared to the parental molecule. The data obtained from studies on the effects of these CR1-based molecules in animal models of disease and their therapeutic applications will also be discussed.

Atypical hemolytic uremic syndrome induced by SARS-CoV2 infection in infants with EXOSC3 mutation.

BackgroundAtypical hemolytic uremic syndrome (aHUS) is a rare disease characterized by systemic thrombotic microangiopathy mainly in the kidneys and mostly due to genetic disorders leading to uncontrolled activation of the complement system. Severe complications of SARS-CoV2 infection are linked to microvascular injury and complement activation is suspected to play a role in the pathogenesis of endothelial cell damage in severe COVID-19.MethodsWe present the first two cases of aHUS triggered by SARS-CoV-2 infection in two unrelated infants with the same mutation in the RNA exosome gene EXOSC3. This mutation is known to cause pontocerebellar hypoplasia type 1b, an autosomal-recessive neurodegenerative disease. So far, no kidney involvement in affected persons was reported.ResultsAs eculizumab treatment was unsuccessful and complement-mediated disorders were ruled out, we suppose that the atypical HUS in our two patients is not due to complement-mediated thrombotic microangiopathy but rather due to a dysfunction of the RNA exosome.ConclusionsThe RNA exosome is crucial for the precise processing and degradation of nuclear and cytoplasmatic RNA. We suspect that the SARS-CoV-2 infection led to changes in RNA that could not be offset by the defective RNA exosome in our two patients. The accumulation/wrong processing of the viral RNA must have led to the endothelial cell damage resulting in aHUS. This would be a new — “RNA-induced” — mechanism of aHUS.

P119: TRIAL IN PROGRESS: THE RANDOMIZED, DOUBLE-BLIND, PLACEBO-CONTROLLED PHASE 1B CROSSWALK-A TRIAL EVALUATING THE SAFETY OF CROVALIMAB FOR THE MANAGEMENT OF ACUTE UNCOMPLICATED VASO-OCCLUSIVE EPISODES (VOES) IN PATIENTS (PTS) WITH SICKLE CELL DISEASE (SCD)

Background: SCD is a group of autosomal recessive red blood cell (RBC) disorders caused by a single point mutation in the β-globin gene, resulting in the production of hemoglobin S (HbS). HbS polymerizes within RBCs under certain conditions, leading to the distortion of the RBC membrane and generation of dense and sickle RBCs. These pathologic RBCs contribute to microvascular occlusions, which may present as acute, painful episodes called VOEs. Although most VOEs are managed at home, >70% of emergency department visits and >90% of hospitalizations for SCD are VOE-related.1,2 Additionally, acute chest syndrome is one of the most severe complications of VOEs and is a leading cause of mortality.3,4 As there are no targeted therapies for the management of VOEs, treatment is currently limited to pain management, blood exchange transfusion (with the risk of complications), and other supportive care, which represents a significant unmet medical need. Overall, accumulating nonclinical data suggest a multimodal role for complement dysregulation in the pathophysiology of SCD including vaso-occlusion, hemolysis, inflammation, thrombogenicity, endothelial activation, and end-organ damage.5 Complement pathway activation has been described in pts with SCD at baseline, in acute pain crises, and in pts with delayed hemolytic transfusion reaction. The complement pathway can be targeted with crovalimab, which is a novel, engineered, anti-complement C5 monoclonal antibody. In a Phase 1/2 paroxysmal nocturnal hemoglobinuria (a complement-mediated disorder) study, crovalimab showed rapid and sustained complement inhibition with promising efficacy and safety.6 Aims: CROSSWALK-a (NCT04912869) is a randomized, double-blind, placebo-controlled, Phase 1b study evaluating the safety of crovalimab in managing acute uncomplicated VOEs in pts with SCD. Methods: Pts aged 12 to 55 years, weighing ≥40 kg, and with a confirmed diagnosis of SCD homozygous hemoglobin S (HbSS) or sickle cell β0 thalassemia (HbSβ0) will be enrolled (Figure). Pts must present with an acute uncomplicated VOE, requiring hospitalization and treatment with parenteral opioid analgesics. Vaccinations against Neisseria meningitidis, Hemophilus influenzae type B, and Streptococcus pneumoniae must be current. Eligible pts will be randomized 2:1 to receive either a single intravenous weight-based tiered dose of crovalimab or placebo. All pts will continue with pain management and other supportive care for their VOE, and may continue concurrent SCD-directed therapies. Pts will be followed during hospitalization until discharge and will also be followed post-discharge during an observational period. The maximum total study duration for an individual pt, including the hospitalization and observational periods, will be 12 weeks. The primary objective is to evaluate the incidence and severity of adverse events according to National Cancer Institute Common Terminology Criteria for Adverse Events, Version 5.0, incidence and severity of infusion-related reactions and hypersensitivity, and change from baseline in targeted vital signs and clinical laboratory test results. Efficacy, pharmacokinetics, pharmacodynamics, immunogenicity, and exploratory biomarker endpoints will also be evaluated. Results: CROSSWALK-a is scheduled to be completed in September 2023. Summary: CROSSWALK-a is enrolling pts with SCD in 5 countries.

Trial in Progress: The Randomized, Double-Blind, Placebo-Controlled Phase Ib CROSSWALK-a Trial Evaluating the Safety of Crovalimab for the Management of Acute Uncomplicated Vaso-Occlusive Episodes (VOEs) in Patients with Sickle Cell Disease (SCD)

BackgroundSCD is a group of autosomal recessive red blood cell (RBC) disorders caused by a single point mutation in the β-globin gene, resulting in the production of hemoglobin S. Hemoglobin S polymerizes within RBCs under certain conditions, leading to the distortion of the RBC membrane and generation of dense and sickle RBCs. These pathologic RBCs contribute to microvascular occlusions in patients with SCD, which present as acute painful episodes called VOEs. Despite the majority of VOEs being managed at home, they remain the most common reason for emergency department (ED) visits and hospitalization among patients with SCD, with > 70% of ED visits and > 90% of hospital admissions for SCD being related to VOEs (Ballas et al. Am J Hematol 2005; Lanzkron et al. Am J Hematol 2010). One of the most severe complications of VOEs is acute chest syndrome, which is a leading cause of mortality among patients with VOEs (Vichinsky et al. N Engl J Med 2000; Bartolucci et al. eBioMedicine 2016). In the absence of targeted therapies for the management of VOEs in patients with SCD, treatment is currently limited to pain management, blood exchange transfusion (with the risk of complications), and other supportive care, representing a significant unmet medical need. Activation of the complement pathway has been described in patients with SCD at baseline, in acute pain crises, and in patients with delayed hemolytic transfusion reaction. Accumulating nonclinical data have suggested a multimodal role for complement dysregulation in the pathophysiology of SCD including vaso-occlusion, hemolysis, inflammation, thrombogenicity, endothelial activation, and end-organ damage (Roumenina et al. Am J Hematol 2020). Crovalimab is a novel, engineered, anti-complement C5 monoclonal antibody. In a Phase I/II study (Röth et al. Blood 2020) in patients with paroxysmal nocturnal hemoglobinuria, a complement-mediated disorder, crovalimab demonstrated rapid and sustained complement inhibition with promising efficacy and safety.Study Design and MethodsCROSSWALK-a (NCT04912869) is a randomized, double-blind, placebo-controlled, Phase Ib study evaluating the safety of crovalimab for the management of acute uncomplicated VOEs in patients with SCD. Patients aged ≥ 12 years to ≤ 55 years, weighing ≥ 40 kg, and with a confirmed diagnosis of SCD homozygous hemoglobin S (HbSS) or sickle cell β 0 thalassemia (HbSβ 0) are eligible for the study (Figure). Patients must present with an acute uncomplicated VOE, requiring hospitalization and treatment with parenteral opioid analgesics. Vaccinations against Neisseria meningitidis, Hemophilus influenzae type B, and Streptococcus pneumoniae must be up to date. Eligible patients will be randomized 2:1 to receive either a single intravenous weight-based tiered dose of crovalimab or placebo. Patients in both study arms will continue to undergo pain management and receive other supportive care for their VOE and may also continue to receive ongoing concurrent SCD-directed therapies. Patients will be followed during the hospitalization until discharge and will continue to be followed post-discharge during an observational period. The maximum total study duration for an individual patient will be 12 weeks, which includes the hospitalization and observational periods. The primary objective is to evaluate the incidence and severity of adverse events according to National Cancer Institute Common Terminology Criteria for Adverse Events, Version 5.0, incidence and severity of infusion-related reactions and hypersensitivity, and change from baseline in targeted vital signs and clinical laboratory test results. Efficacy, pharmacokinetics, pharmacodynamics, immunogenicity, and exploratory biomarker endpoints will also be evaluated. [Display omitted] DisclosuresBartolucci: F. Hoffmann-La Roche Ltd: Consultancy; Bluebird: Consultancy, Research Funding; Emmaus: Consultancy; Hemanext: Consultancy; Jazz Pharma: Other: Lecture fees; AGIOS: Consultancy; GBT: Consultancy; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: Lecture fees, Steering committee, Research Funding; INNOVHEM: Other: Co-founder; Fabre Foundation: Research Funding; Addmedica: Consultancy, Other: Lecture fees, Research Funding. Ataga: Forma Therapeutics: Membership on an entity's Board of Directors or advisory committees; Novo Nordisk: Membership on an entity's Board of Directors or advisory committees; F. Hoffmann-La Roche Ltd: Consultancy; Novartis: Membership on an entity's Board of Directors or advisory committees; Agios Pharmaceuticals: Consultancy; Global Blood Therapeutics: Membership on an entity's Board of Directors or advisory committees. Callaghan: Alnylum: Current equity holder in publicly-traded company; Roche/Genentech: Consultancy, Speakers Bureau; Takeda: Consultancy, Speakers Bureau; Sanofi: Consultancy; Pfizer: Consultancy; Global Blood Therapeutics: Consultancy, Speakers Bureau; uniQure: Consultancy; Spark: Consultancy; Biomarin: Consultancy; Kedrion: Consultancy; Hema Biologics: Consultancy; Forma: Consultancy; Chesei: Consultancy; Agios Pharmaceuticals: Current Employment. De Franceschi: F. Hoffmann-La Roche Ltd: Consultancy. Minniti: F. Hoffmann-La Roche: Consultancy; Bluebird Bio: Other: Endpoint adjudicator; Forma: Consultancy; GBT: Consultancy; Novo Nordisk: Consultancy; CSL Behring: Other: Endpoint adjudicator; Novartis: Consultancy; Chiesi: Consultancy. Alexandrou: F. Hoffmann-La Roche Ltd: Consultancy, Current equity holder in publicly-traded company; Pfizer: Current equity holder in publicly-traded company. Imbs: F. Hoffmann-La Roche Ltd: Consultancy; Certara Inc.: Current Employment. Fox: Parexel International: Current Employment; Genentech, Inc.: Current Employment. Patel: Genentech, Inc.: Current Employment. Sostelly: F. Hoffmann-La Roche Ltd: Current Employment.

Tissue-specific complement inhibition by antibody-Complement Regulatory Protein (CRP) fusions utilizing Therapeutic Autoimmune reguLatOry proteiN (TALON™) platform

Abstract The complement system plays a major role in humoral innate immunity. Upon activation, complement proteins trigger a cascade of enzymatic events leading to the production of inflammatory mediators and membrane attack complexes. Tight regulation of complement proteins is essential for immune homeostasis due to their potency and abundance in fluid phase. Complement Regulatory Proteins (CRPs) that are membrane-bound (e.g., CD59), or in fluid phase (e.g., Factor H), prevent aberrant activation of complement pathways against self and mitigate autoimmunity. The importance of CRPs is underscored by the finding that genetic variations in the CRPs can lead to autoimmune diseases such as paroxysmal nocturnal hemoglobinuria (PNH) characterized by the lysis of host red blood cells from dysregulated complement system. Systemic complement inhibition via anti-C5 antibody has clinically been shown to benefit some patients with complement-mediated disorders, but the therapy requires a high, frequent dosing scheme and supplemental vaccinations to compensate for systemic immunosuppression. In addition, other diseases such as C3 glomerulopathy affect specific tissues and involve pathways beyond C5, limiting treatment options. To this end, local immunomodulation through targeted delivery of CRPs to specific tissues is an attractive strategy to restore homeostasis in complement-mediated autoimmune diseases while circumventing the need for high dose systemic inhibition of complement factors. Here, we present a proof-of-concept study for the design, generation, and evaluation of tissue-tethering antibodies fused with CRPs. Also, we demonstrate the pharmacokinetics and pharmacodynamics of tissue-targeting CRP fusions in mouse models.

A novel soluble complement receptor 1 fragment with enhanced therapeutic potential

Human complement receptor 1 (HuCR1) is a pivotal regulator of complement activity, acting on all three complement pathways as a membrane-bound receptor of C3b/C4b, C3/C5 convertase decay accelerator, and cofactor for factor I-mediated cleavage of C3b and C4b. In this study, we sought to identify a minimal soluble fragment of HuCR1, which retains the complement regulatory activity of the wildtype protein. To this end, we generated recombinant, soluble, and truncated versions of HuCR1 and compared their ability to inhibit complement activation in vitro using multiple assays. A soluble form of HuCR1, truncated at amino acid 1392 and designated CSL040, was found to be a more potent inhibitor than all other truncation variants tested. CSL040 retained its affinity to both C3b and C4b as well as its cleavage and decay acceleration activity and was found to be stable under a range of buffer conditions. Pharmacokinetic studies in mice demonstrated that the level of sialylation is a major determinant of CSL040 clearance in vivo. CSL040 also showed an improved pharmacokinetic profile compared with the full extracellular domain of HuCR1. The in vivo effects of CSL040 on acute complement-mediated kidney damage were tested in an attenuated passive antiglomerular basement membrane antibody-induced glomerulonephritis model. In this model, CSL040 at 20 and 60 mg/kg significantly attenuated kidney damage at 24 h, with significant reductions in cellular infiltrates and urine albumin, consistent with protection from kidney damage. CSL040 thus represents a potential therapeutic candidate for the treatment of complement-mediated disorders.

Ravulizumab in the treatment of paroxysmal nocturnal hemoglobinuria

ABSTRACT Introduction Eculizumab, the anti-C5 monoclonal antibody therapeutic, has revolutionized the treatment of Paroxysmal Nocturnal Hemoglobinuria (PNH) and has shown efficacy in other complement-mediated disorders including atypical Hemolytic Uremic Syndrome (aHUS) and Myasthenia Gravis (MG). Despite the effectiveness of eculizumab, challenges remain in the treatment of these diseases, including breakthrough hemolysis, frequent intravenous infusions, and cost; in an effort to mitigate these challenges, the new monoclonal therapeutic ravulizumab has been developed. Areas covered In this paper, we review the characteristics and clinical performance of ravulizumab and assess its potential utility in the treatment of PNH, and its position in this rapidly-developing therapeutic landscape. A review of published data up to Phase III of ravulizumab had been performed. Studies were identified via Google Scholar, PubMed, the United States National Library of Medicine Clinical Trials, citation chasing, and topic knowledge of the authors. Expert opinion Ravulizumab represents an important advance in the clinical care of complement-mediated disorders. Clinical trials demonstrate non-inferior efficacy and indistinguishable safety and tolerability to eculizumab, with added patient-preferred benefits including longer intervals between infusions. Though improved, physicians should be aware of the limitations of ravulizumab, including the need for intravenous infusions and possible breakthrough hemolysis.

SKY59, A Novel Recycling Antibody for Complement-mediated Diseases.

The complement system usually helps protect against microbial infection, but it could also be involved in the onset of various diseases. Inhibition of complement component 5 (C5) with eculizumab has resulted in a significant reduction of hemolysis, reduction of thromboembolic events, and increased survival in patients with Paroxysmal Nocturnal Hemoglobinuria (PNH). However, eculizumab requires frequent intravenous infusions due to the abundance of C5 in plasma and some patients may still experience breakthrough hemolysis. This review introduces the recent body of knowledge on recycling technology and discusses the likely therapeutic benefits of SKY59, a novel recycling antibody, for PNH and complement-mediated disorders. By using recycling technology, we created a novel anti-C5 antibody, SKY59, capable of binding to C5 pH-dependently. In cynomolgus monkeys, SKY59 robustly inhibited C5 and complement activity for significantly longer than a conventional antibody. SKY59 also showed an inhibitory effect on C5 variant p.Arg885His, whereas eculizumab does not suppress complement activity in patients with this type of mutation. SKY59 is a promising anti-C5 biologic agent that has significant advantages over current therapies such as long duration of action and efficacy against C5 variants.

Microthrombosis associated with GI bleeding in COVID-19

A 35-year-old man with sickle-cell trait and diabetes who had undergone renal transplantation presented with weeks of episodic severe cramping, abdominal pain, diarrhea, maroon stool, and anorexia despite using empiric antibiotics for gastroenteritis. At presentation, his laboratory results noted a mild leukocytosis and a hemoglobin of 6.4 g/dL. Real-time polymerase chain reaction of a nasopharyngeal sample gave a positive result for SARS-CoV-2 despite the lack of respiratory symptoms or fever. A CT scan demonstrated diffusely thickening of the stomach and small-bowel walls (A). Upper endoscopy revealed normal gastric mucosa but edematous, friable duodenal mucosa with contact bleeding (B). Examination of biopsy specimens from the duodenal mucosa demonstrated hemorrhage in the lamina propria with fibrin microthrombi (C, H&E, orig. mag. ×40). A biopsy specimen from a small gastroesophageal junction polyp incidentally showed dysplasia with a focus of adenocarcinoma (D). Magnetic resonance angiography failed to show any explanatory large-vessel disease. The patient’s melena resolved 4 weeks after symptom onset and 3 blood transfusions. Repeated endoscopy at 5 weeks showed normal mucosa; the polyp was successfully removed en bloc by submucosal dissection. No other thromboses were noted during his course; d-dimer peaked at 1633 mg/mL. GI involvement of SARS-CoV-2 is common; typical symptoms include diarrhea and nausea, although bleeding has been rarely described. To our knowledge, this is the first endoscopic description of biopsy-confirmed COVID-19–associated microthrombosis, reinforcing the importance of endoscopy with adequate biopsy. All authors disclosed no financial relationships. Commentary As our knowledge about COVID-19 evolves, we are starting to view the illness as a thrombotic microvascular injury of systemic nature in which the lung, skin, kidney, GI tract, and other organs are affected. Microthrombosis in COVID-19 is a complement-mediated disorder. It is unclear whether COVID-19–related coagulopathy is a byproduct of severe inflammatory response or a virus-specific–mediated coagulopathy. As seen in this case, COVID-19–related microthrombotic disease of the small intestine presents endoscopically as edema, granularity, and patchy erythema, as seen in other ischemic disorders. Biopsy specimens showing hemorrhage in the lamina propria with fibrin microthrombi are diagnostic of the condition. With the current rise of COVID-19 cases in hospitals worldwide, endoscopists should be aware of COVID-19–related microthrombosis syndrome as a cause of GI bleeding in this subset of patients. How prevalent this condition is among our hospitalized patients with COVID-19 and whether other comorbid conditions pose a synergetic effect on developing microthromboses are questions that remain to be answered. Mohamed O. Othman, MD, Associate Editor for Focal Points

Complement-Mediated Disorders in Pregnancy.

Complement-mediated disorders in pregnancy span a large spectrum and have been implicated in all three complement pathways: classical, lectin, and alternative. Our understanding of these disorders in recent years has advanced due to a better understanding of complement regulatory proteins, such as complement factor H, complement factor I, membrane cofactor protein, and thrombomodulin that particularly affect the alternative complement pathway. Enthusiasm in genotyping for mutations that encode these proteins has allowed us to study the presence of genetic variants which may predispose women to develop conditions such as pregnancy-associated hemolytic uremic syndrome (P-aHUS), thrombotic thrombocytopenic purpura, preeclampsia/hemolysis, elevated liver enzymes, low platelets (HELLP), systemic lupus erythematosus/antiphospholipid syndrome, and peripartum cardiomyopathy. The advent of the anti-C5-antibody eculizumab to quench the complement cascade has already proven in small case series to improve maternal kidney outcomes in complement-mediated obstetric catastrophes such as P-aHUS and HELLP. In this review, we will detail the pathogenesis behind these complement-mediated pregnancy disorders, the role of complementvariants in disease phenotype, and the most up-to-date experience with eculizumab in this population.

Investigational RNAi Therapeutic Targeting C5 Is Efficacious in Pre-clinical Models of Myasthenia Gravis

Complement-mediated damage to the neuromuscular junction (NMJ) is a key mechanism of pathology in myasthenia gravis (MG), and therapeutics inhibiting complement have shown evidence of efficacy in the treatment of MG. In this study, we describe the development of a subcutaneously administered N-acetylgalactosamine (GalNAc)-conjugated small interfering RNA (siRNA) targeting the C5 component of complement that silences C5 expression in the liver (ALN-CC5). Treatment of wild-type rodents with ALN-CC5 resulted in robust and durable suppression of liver C5 expression. Dose-dependent serum C5 suppression was observed in non-human primates, with a lowering of serum C5 of up to 97.5% and the concomitant inhibition of serum complement activity. C5 silencing was efficacious in ameliorating disease symptoms in two standard rat models of MG, demonstrating the key role of circulating C5 in pathology at the NMJ. Improvement in disease activity scores and NMJ pathology was observed at intermediate levels of complement activity inhibition, suggesting that complete ablation of complement activity may not be required for efficacy in MG. The pre-clinical studies of ALN-CC5 and efficacy of C5 silencing in rat models of MG support further clinical development of ALN-CC5 as a potential therapeutic for the treatment of MG and other complement-mediated disorders.

Novel insights into the treatment of complement-mediated hemolytic anemias.

Complement-mediated hemolytic anemias can either be caused by deficiencies in regulatory complement components or by autoimmune pathogenesis that triggers inappropriate complement activation. In paroxysmal nocturnal hemoglobinuria (PNH) hemolysis is entirely complement-driven. Hemolysis is also thought to be complement-dependent in cold agglutinin disease (CAD) and in paroxysmal cold hemoglobinuria (PCH), whereas warm antibody autoimmune hemolytic anemia (wAIHA) is a partially complement-mediated disorder, depending on the subtype of wAIHA and the extent of complement activation. The pathophysiology, clinical presentation, and current therapies for these diseases are reviewed in this article. Novel, complement-directed therapies are being rapidly developed. Therapeutic terminal complement inhibition using eculizumab has revolutionized the therapy and prognosis in PNH but has proved less efficacious in CAD. Upstream complement modulation is currently being investigated and appears to be a highly promising therapy, and two such agents have entered phase II and III trials. Of these, the anti-C1s monoclonal antibody sutimlimab has shown favorable activity in CAD, while the anti-C3 cyclic peptide pegcetacoplan appears to be promising in PNH as well as CAD, and may also have a therapeutic potential in wAIHA.