Complement Component C3 Research Articles

The Case | A benign reason behind a malignant finding

A 21-year-old man presented to the office for deranged kidney function. He was diagnosed to have C3 glomerulonephritis at the age of 18 years when he presented to his first physician with hypertension. Laboratory parameters showed high serum creatinine (2.7 mg/dl), low serum albumin (2.9 g/dl), low total protein (5.2 g/dl), dysmorphic red blood cells on urinalysis, high 24-hour urine protein (5.7 g/d), low complement C3 (32 mg/dl), and low 50% hemolytic complement (10 U/ml). Kidney biopsy showed a membranoproliferative pattern of injury with 3+ reactivity for complement C3 and significant chronic changes. There was moderate tubulointerstitial fibrosis and 30% of all sampled glomeruli were globally sclerosed. Clinically, there were no signs of atypical hemolytic uremic syndrome and pathologically no evidence of thrombotic microangiopathy. He was started on steroids and mycophenolate mofetil, which were discontinued after 3 months in view of intolerable side effects. Thereafter, ravulizumab, a new complement component C5 inhibitor with a longer half-life, was started and he received 4 infusions (3 doses of 3300 mg and 1 dose of 2700 mg at 8 weekly intervals) until his recent visit to the office. Laboratory parameters at that time showed persistently high serum creatinine (2.3 mg/dl), high 24-hour urine protein (6 g/d), and dysmorphic red blood cells on urinalysis; however, 50% hemolytic complement (40 U/ml) and complement C3 (117 mg/dl) normalized. A repeat kidney biopsy was done in view of no clinical response to therapy, and the direct immunofluorescence findings of pretreatment and post-treatment kidney biopsies are shown in Figure 1.

Design, Synthesis of Dioxopiperidinamide Derivatives by Amide Coupling Reaction and Study of Their Biological Activity

AbstractA series of fifteen dioxopiperidinamide derivatives synthesized and their spectra chemical data were reported. An alternative amide coupling reaction has been followed by using 3‐aminopiperidine‐2,6‐dione through amide bond functionalization between N‐Ethyl‐N′‐(3‐dimethylaminopropyl)carbodiimide hydrochloride (EDC.HCl) and Hydroxybenzotriazole (HOBt). All the designed compounds were docked in to the active sites of 2 A73: Human Complement Component C3. All the docked compounds is favorable to make both hydrogen and hydrophobic interaction with amino acids located at the binding site. The most of potential compounds (E)‐N‐(2,6‐dioxopiperidin‐3‐yl)‐3‐(m‐tolyl)acrylamide (4 g), (E)‐3‐(4‐bromophenyl)‐N‐(2,6‐dioxopiperidin‐3‐yl)acrylamide (4 h) and N‐(2,6‐dioxopiperidin‐3‐yl)‐3‐methyl‐4‐oxo‐4,6,7,8,9,10‐hexahydrothieno [2′,3′:4,5]pyrimido[1,2‐a]azepine‐2‐carboxamide (7) studied for their anticancer activity on the THP‐1 cancer cells line. It was also further established their Anti‐inflammatory activity and Antioxidant activity with the comparable efficiency against those of clinically used drugs.

Protein Expression and Bioinformatics Study of Granulosa Cells of Polycystic Ovary Syndrome Expressed Under the Influence of DHEA.

The reproductive system is heavily dependent on ovarian follicles, which are made up of germ cells (oocytes) and granulosa cells (GCs), including cumulus granulosa cells (CGCs) and mural granulosa cells (MGCs). Understanding their normal and steroid-induced functions is the key to understanding the pathophysiology of endocrinal diseases in women. This study investigated the differentially expressed proteins by CGCs and MGCs of patients with polycystic ovarian syndrome (PCOS) and without subsequent exposure to dehydroepiandrosterone sulfate (DHEAS) and functional differentiation. The present study was observational and experimental study carried out in hospital involving 80 female patients undergoing IVF for infertility. In this study, we isolated CGCs and MGCs from the follicular fluid of both PCOS and non-PCOS patients undergoing in vitro fertilization (IVF). The cells were cultured and treated with DHEAS for 48 hours, and these cells were extracted, digested, and analyzed by tandem mass spectrometry followed by processing of the results using open-source bioinformatics tools. The present investigation discovered 276 and 341 proteins in CGCs and MGCs, respectively. DHEAS reduced the number of proteins expressed by CGCs and MGCs to 34 and 57 from 91 and 94, respectively. Venn results of CGCs revealed 49, 53, 36, and 21 proteins in normal CGCs, PCOS-CGCs, post-DHEAS, and PCOS-CGCs, respectively. Venn analysis of MGCs showed 51 proteins specific to PCOS and 29 shared by normal and PCOS samples after DHEAS therapy. MGCs express the most binding and catalytic proteins, whereas CGCs express transporter-related proteins. A protein pathway study demonstrated considerable differences between normal and PCOS samples, while DHEAS-treated samples of both cell lines showed distinct pathways. String findings identified important network route components such as albumin, actin, apolipoprotein, complement component C3, and heat shock protein. This is the first study to show how DHEAS-induced stress affects the expression of proteins by MGCs and CGCs isolated from normal and PCOS patients. Further studies are recommended to identify PCOS biomarkers from CGCs and MGCs expressed under the influence of DHEAS.

“PNH is getting very popular!”: New clinical trial data and management insights from ASH 2023

Paroxysmal nocturnal hemoglobinuria (PNH) is a rare acquired disorder of hematopoietic stem cells that leaves red blood cells (RBCs) susceptible to complement-mediated lysis, with clinical manifestations that can include anemia, hemoglobinuria, and thrombosis. Inhibition of the terminal complement component C5 with eculizumab or ravulizumab can control intravascular hemolysis (IVH) and improve clinical parameters, patient survival, and quality of life (QoL). However, terminal complement inhibition alone may leave RBCs vulnerable to extravascular hemolysis (EVH) and persistent anemia in a minority of patients. This report focuses on clinical trial and real-world data presented at the recent annual meeting of the American Society of Hematology (ASH) for currently available and novel complement inhibitors in PNH.

Dendritic spine loss in epileptogenic Type II focal cortical dysplasia: Role of enhanced classical complement pathway activation.

Dendritic spines are the postsynaptic sites for most excitatory glutamatergic synapses. We previously demonstrated a severe spine loss and synaptic reorganization in human neocortices presenting Type II focal cortical dysplasia (FCD), a developmental malformation and frequent cause of drug-resistant focal epilepsy. We extend the findings, investigating the potential role of complement components C1q and C3 in synaptic pruning imbalance. Data from Type II FCD were compared with those obtained in focal epilepsies with different etiologies. Neocortical tissues were collected from 20 subjects, mainly adults with a mean age at surgery of 31 years, admitted to epilepsy surgery with a neuropathological diagnosis of: cryptogenic, temporal lobe epilepsy with hippocampal sclerosis, and Type IIa/b FCD. Dendritic spine density quantitation, evaluated in a previous paper using Golgi impregnation, was available in a subgroup. Immunohistochemistry, in situ hybridization, electron microscopy, and organotypic cultures were utilized to study complement/microglial activation patterns. FCD Type II samples presenting dendritic spine loss were characterized by an activation of the classical complement pathway and microglial reactivity. In the same samples, a close relationship between microglial cells and dendritic segments/synapses was found. These features were consistently observed in Type IIb FCD and in 1 of 3 Type IIa cases. In other patient groups and in perilesional areas outside the dysplasia, not presenting spine loss, these features were not observed. In vitro treatment with complement proteins of organotypic slices of cortical tissue with no sign of FCD induced a reduction in dendritic spine density. These data suggest that dysregulation of the complement system plays a role in microglia-mediated spine loss. This mechanism, known to be involved in the removal of redundant synapses during development, is likely reactivated in Type II FCD, particularly in Type IIb; local treatment with anticomplement drugs could in principle modify the course of disease in these patients.

Altered expression of microglial markers of phagocytosis in schizophrenia

BackgroundCognitive disturbances in schizophrenia have been linked to a lower density of dendritic spines on pyramidal neurons in the prefrontal cortex (PFC). Complement component C4, which has previously been found at higher levels in schizophrenia, marks synapses for phagocytosis by microglia. Thus, elevated consumption of dendritic spines by microglia mediated through excessive complement activity may play a role in lower spine density in schizophrenia. However, it is unclear if microglia themselves have the molecular capacity for enhanced phagocytosis of spines in schizophrenia. MethodsTranscript levels for complement components and microglia-specific phagocytic markers were quantified using quantitative PCR in the PFC of 62 matched pairs of schizophrenia and unaffected comparison subjects and in antipsychotic-exposed monkeys. ResultsRelative to comparison subjects, schizophrenia subjects had higher mRNA levels for C4 (+154 %); C1q (+69 %), which initiates the classical complement pathway that includes C4; and for microglia-specific markers that enable phagocytic activity including TAM receptor tyrosine kinases Axl (+27 %) and MerTK (+27 %) and lysosome-associated glycoprotein CD68 (+27 %) (all p ≤ .042). Transcript levels for microglial phagocytic markers were correlated with C4 mRNA levels in schizophrenia subjects (all r ≥ 0.31, p ≤ .015). We also found further evidence consistent with microglial activation in schizophrenia, including higher mRNA levels for THIK1 (TWIK-related halothane-inhibited potassium channel: +30 %) and lower mRNA levels for the purinergic receptor P2Y12 (−27 %) (all p ≤ .016). Transcript levels were unchanged in antipsychotic-exposed monkeys. ConclusionsThese results are consistent with the presence of increased complement activity and an elevated molecular capacity of microglia for phagocytosis in the same schizophrenia subjects.

Complement C3b contributes to Escherichia coli-induced platelet aggregation in human whole blood.

Platelets have essential functions as first responders in the immune response to pathogens. Activation and aggregation of platelets in bacterial infections can lead to life-threatening conditions such as arterial thromboembolism or sepsis-associated coagulopathy. In this study, we investigated the role of complement in Escherichia coli (E. coli)-induced platelet aggregation in human whole blood, using Multiplate® aggregometry, flow cytometry, and confocal microscopy. We found that compstatin, which inhibits the cleavage of complement component C3 to its components C3a and C3b, reduced the E. coli-induced platelet aggregation by 42%-76% (p = 0.0417). This C3-dependent aggregation was not C3a-mediated as neither inhibition of C3a using a blocking antibody or a C3a receptor antagonist, nor the addition of purified C3a had any effects. In contrast, a C3b-blocking antibody significantly reduced the E. coli-induced platelet aggregation by 67% (p = 0.0133). We could not detect opsonized C3b on platelets, indicating that the effect of C3 was not dependent on C3b-fragment deposition on platelets. Indeed, inhibition of glycoprotein IIb/IIIa (GPIIb/IIIa) and complement receptor 1 (CR1) showed that these receptors were involved in platelet aggregation. Furthermore, aggregation was more pronounced in hirudin whole blood than in hirudin platelet-rich plasma, indicating that E. coli-induced platelet aggregation involved other blood cells. In conclusion, the E. coli-induced platelet aggregation in human whole blood is partly C3b-dependent, and GPIIb/IIIa and CR1 are also involved in this process.

Dietary Effect of Borage (Borago officinalis) Powder on Growth Performance, Immune Response, and Blood Biochemical Parameters in Common Carp (Cyprinus carpio) Juvenile

In this study, the effect of borage (Borago officinalis) powder on juveniles of common carp (Cyprinus carpio) were investigated. The diets contained control (0), 0.5%, 1.5%, and 2.5% of borage powder. The fish with the initial average weight of 8.40 ± 1.37 were allotted to 12 circular tanks of 300 L capacity at a density of ten fish per each tank and fed to apparent satiety three times a day. The growth performance of the fish showed no significant difference compared to the control group (P > 0.05). By adding 1.5% borage powder to the control diet the mucosal immunity of fish in terms of lysozyme activity (14.26%), total protein (69.64%), immunoglobulin (30.78%), and humoral immunity in term of lysozyme activity (9.7%), complement components C3 (8.27%), C4 (21.64%), and total protein (9.33%) were increased significantly (P < 0.05). A total number of red blood cell (12.39%), mean corpuscular volume (MCV, 8.99%), mean corpuscular hemoglobin (MCH, 10.22%), and hemoglobin (2.69%) showed a significant increase in the fish fed supplemented diet with 1.5% borage powder compared to the control (P < 0.05). The results showed supplemented diet with 1.5% dietary borage powder induced high immunity and survival rate of juvenile common carp.

Plasma complement component C2: a potential biomarker for predicting abdominal aortic aneurysm related complications.

Blood-based adjunctive measures that can reliably predict abdominal aortic aneurysm (AAA)-related complications hold promise for mitigating the AAA disease burden. In this pilot study, we sought to evaluate the prognostic performance of complement factors in predicting AAA-related clinical outcomes. We recruited consecutive AAA patients (n = 75) and non-AAA patients (n = 75) presenting to St. Michael's Hospital. Plasma levels of complement proteins were assessed at baseline, as well as prospectively measured regularly over a period of 2years. The primary outcome was the incidence of rapidly progressing AAA (i.e. aortic expansion), defined as change in AAA diameter by either 0.5cm in 6months, or 1cm in 12months. Secondary outcomes included incidence of major adverse aortic events (MAAE) and major adverse cardiovascular events (MACE). All study outcomes (AAA diameter, MACE and MAAE) were obtained during follow-up. Multivariable adjusted Cox regression analyses were performed to assess the prognostic value of plasma C2 levels in patients with AAA regarding rapid aortic expansion and MAAE and MACE. Event-free survival rates of both groups were also compared. Compared to non-AAA patients, patients with AAA demonstrated significantly higher plasma concentrations of C1q, C4, Factor B, Factor H and Factor D, and significantly lower plasma concentrations of C2, C3, and C4b (p = 0.001). After a median of 24months from initial baseline measurements, C2 was determined as the strongest predictor of rapid aortic expansion (HR 0.10, p = 0.040), MAAE (HR 0.09, p = 0.001) and MACE (HR 0.14, p = 0.011). Based on the data from the survival analysis, higher levels of C2 at admission in patients with AAA predicted greater risk for rapid aortic expansion and MAAE (not MACE). Plasma C2 has the potential to be a biomarker for predicting rapid aortic expansion, MAAE, and the eventual need for an aortic intervention in AAA patients.

Proteomic profiling of circulating plasma exosomes reveals novel biomarkers of Alzheimer’s disease

BackgroundNeuronal- and astrocyte-derived exosomes have been identified as an optimal source for screening biomarkers for Alzheimer’s disease (AD). However, few studies focus on the bulk exosome population isolated from plasma of AD. This study investigated whether proteins in bulk exosomes can aid in the diagnosis of AD.MethodsThe plasma exosomes were collected by ultracentrifuge. Protein samples were extracted from exosomes. Cerebrospinal fluid levels of amyloid β (Aβ)42 and phosphorylated tau (P-tau)181 were measured for diagnostic purposes. A pilot study (controls, 20; AD, 20) followed by a second dataset (controls, 56; AD, 58) was used to establish a diagnostic model of AD. Mass spectrometry-based proteomics was performed to profile the plasma exosomal proteome. Parallel reaction monitoring was used to further confirm the differentially expressed proteins.ResultsIn total, 328 proteins in plasma exosomes were quantified. Among them, 31 proteins were altered in AD patients, and 12 were validated. The receiver operating characteristic curve analysis revealed a combination of six proteins (upregulated: Ig-like domain-containing protein (A0A0G2JRQ6), complement C1q subcomponent subunit C (C1QC), complement component C9 (CO9), platelet glycoprotein Ib beta chain (GP1BB), Ras suppressor protein 1 (RSU1); downregulated: disintegrin and metalloproteinase domain 10 (ADA10)) has the capacity to differentiate AD patients from healthy controls with high accuracy. Linear correlation analysis showed that the combination was significantly correlated with cognitive performance.ConclusionsThe combination of plasma exosomal proteins A0A0G2JRQ6, C1QC, CO9, GP1BB, RSU1, and ADA10 acts as a novel candidate biomarker to differentiate AD patients from healthy individuals.

Changes in Maternal Serum Levels of C3 and C4 Complement Components in Different Delivery Methods and Postpartum Hemorrhage.

Activation of the complement system may play a role in the pathophysiology of human labor. Yet no unanimous conclusion has been drawn. To compare the differences in maternal complement components C3 and C4 serum levels in cesarean section and the vaginal delivery at term and in the postpartum hemorrhage. One hundred and sixty six women delivered at term were enrolled in this study. Maternal blood samples were obtained from 47 cases of elective cesarean section and 119 cases of the vaginal delivery. Serum complement levels were measured subsequently by immuno-scatter turbidimetry. The maternal complement levels declined significantly during delivery by both the cesarean section and the vaginal delivery (p＜0.01) in comparison with the baseline. A much larger drop of C3 serum level was found in the postpartum hemorrhage and in the vaginal delivery, and the incidence of the postpartum hemorrhage has a positive correlation with the complement decline rate. The complement system may be involved in the delivery process and represents a predictive value in postpartum hemorrhage.

Clinical efficacy and safety of sirolimus in childhood-onset systemic lupus erythematosus in real world.

To investigate the effectiveness and safety of sirolimus in childhood-onset systemic lupus erythematosus in a real world. This is a retrospective real world clinical study. All childhood-onset systemic lupus erythematosus patients treated with sirolimus in Children's Hospital of Hebei Province China were analyzed. They were treated with sirolimus and followed up regularly. The patients had systemic Lupus Erythematosus Disease Activity Index 2000 (SLEDAI-2K) score, levels of antidouble-stranded DNA antibody, complement components C3 and C4, 24-hour proteinuria and corticosteroid reduction were recorded at baseline and at 6, 12, and 18 months. Adverse events were also collected. Thirty-two patients were enrolled in the study. SLEDAI-2K were improved on all time-points (P < .05). Complement levels increased and the levels of antidouble-stranded DNA antibody decreased during treatment. The mean dose of prednisone tapered and achieved significant reduction after 12 months therapy (15.4 ± 5.8 mg/d to 4.8 ± 2.1 mg/d; P < .05). Sirolimus was well tolerated and only 5 patients (15.6%) experienced adverse events, all of which were classified as infections (2 bacterial infection and 3 viral infections). No deaths, severe infusion reactions, or hypersensitivity reactions were found. Sirolimus use was associated with a decrease in disease activity and ability to tolerate tapering of oral glucocorticoid dose with a favorable risk-benefit profile.

Preliminary Dose-Escalation Results from a Phase 1/2 Study of GEN3014 (HexaBody®-CD38) in Patients (pts) with Relapsed or Refractory Multiple Myeloma (RRMM)

Background: CD38-targeted antibodies (Abs) are foundational treatments (Tx) for pts with newly diagnosed or RRMM. GEN3014 (HexaBody®-CD38) is a novel human IgG1 anti-CD38 monoclonal Ab (mAb) with an E430G hexamerization-enhancing mutation that facilitates highly efficient complement-dependent cytotoxicity (CDC). GEN3014 showed preclinical tumor cell killing through highly potent CDC (more efficient than daratumumab) and Ab-dependent cellular cytotoxicity and phagocytosis. We present preliminary data from dose escalation in the first-in-human, phase 1/2 trial of GEN3014 in pts with RRMM (NCT04824794). Methods: Adults with RRMM were enrolled if they had ≥3 prior lines of Tx including a proteasome inhibitor (PI) and an immunomodulatory imide drug (IMiD), were double refractory to a PI and IMiD, or had ≥2 prior lines of Tx if 1 combined a PI and IMiD. GEN3014 was tested at 6 dose levels ranging from 0.2 to 24 mg/kg and administered intravenously in 28-d cycles (first dose split equally between D1 and D2 of cycle 1, then full doses QW through cycle 2; Q2W, cycles 3-6; Q4W, cycles ≥7). Primary objectives were to determine the recommended phase 2 dose (RP2D) and maximum tolerated dose. Other endpoints included pharmacokinetic (PK) and pharmacodynamic (PD) characterization. PD biomarker analysis included immunophenotyping and quantification of complement factors and cytokines in peripheral blood. Results: As of July 10, 2022, 24 pts with RRMM were treated (median age, 65 y; range, 45-84). Pts were heavily pretreated with a median of 7 prior lines of Tx (range, 3-13). Most pts (67%) had prior exposure to a daratumumab- or isatuximab-containing regimen, with 8 pts naive to anti-CD38 mAb. GEN3014 was well tolerated; the most common Tx-emergent AEs were infusion-related reactions (IRRs; 75.0%), neutropenia (62.5%), anemia (33.3%), diarrhea (33.3%), pyrexia (25.0%), and thrombocytopenia (25.0%). IRRs were mostly low grade (G; 62.5% G1-2, 12.5% G3, no G4), occurred mainly during the first infusion, and were manageable. There were no Tx-related deaths or dose-limiting toxicities observed at doses up to 16 mg/kg, the RP2D. Disease progression led to discontinuation in 80% of pts. Among 19 response-evaluable pts (5 naive to anti-CD38 mAb, 14 refractory to anti-CD38 mAb), preliminary antitumor activity was seen. In pts naive to anti-CD38 mAb, 2 pts achieved very good partial response (1 each at 4 and 24 mg/kg); 1 achieved minimal response (MR; 16 mg/kg). In pts refractory to anti-CD38 mAb, 2 pts achieved MR (1 each at 8 and 16 mg/kg). Five pts remained on Tx (4 naive and 1 refractory to anti-CD38 mAb). Updated data will be presented. Peak GEN3014 concentrations at the end of infusion increased roughly proportionally with dose. The interval area under the concentration-time curve showed a more than proportional increase with doses ≤8 mg/kg but a proportional or less than proportional increase at doses >8 mg/kg. These results suggest target-mediated drug disposition at lower doses and a high degree of target saturation at >8 mg/kg. Trough concentrations increased with weekly administration, most notably in pts responding to Tx, indicating target cell depletion over time. GEN3014 Tx was associated with a rapid, sustained decrease in peripheral blood natural killer cells at all doses in all pts (mean 92.4% ± 9.2% peak reduction from baseline [BL], n=21). T cells transiently decreased after administration of first doses ≥4 mg/kg, followed by expansion (≥100% increase from BL), particularly in pts naive to anti-CD38 mAb. GEN3014 induced transient reduction in complement component C2 (mean 58% ± 18% peak reduction from BL, n=18) and total complement lytic activity (CH50; mean 48% ± 24% peak reduction from BL for ≥8 mg/kg, n=11) at all evaluable doses, suggesting CDC. Complement parameters rapidly returned to BL, indicating that Tx does not exhaust complement. Plasma cytokine (IL-2, IL-6, IL-8, IL-10, IFNγ, and TNFα) levels generally remained low after Tx. Conclusions: Dose-escalation results show GEN3014 had a tolerable safety profile and clinical activity in pts with RRMM, including pts with prior anti-CD38 mAb. The PK profile is characterized by target-mediated drug disposition at lower doses and signs of saturation at doses >8 mg/kg. Biomarker analyses confirm biological activity in pts with RRMM at all evaluated doses. The expansion part of the trial evaluating the 16 mg/kg RP2D is ongoing.

Patient-Reported Outcomes from a Phase 2, Randomized Trial Evaluating the Safety and Efficacy of Pozelimab and Cemdisiran in Patients with Paroxysmal Nocturnal Hemoglobinuria

Background: Paroxysmal nocturnal hemoglobinuria (PNH) is an ultra-rare acquired genetic disease characterized by uncontrolled complement activation, which results in intravascular hemolysis and an increased tendency to develop thrombosis. PNH causes symptoms such as severe fatigue, which can negatively impact a patient's physical functioning and health-related quality of life (QoL). Cemdisiran is an investigational N-acetylgalactosamine-conjugated small interfering RNA (siRNA) that suppresses liver production of complement component C5, while pozelimab is an investigational fully human monoclonal antibody inhibitor of C5. The combination of pozelimab and cemdisiran is being evaluated in an ongoing phase 2, randomized, open-label, two-arm study that is designed to evaluate the safety and efficacy of combination therapy in patients with PNH who were transitioning from pozelimab monotherapy (NCT04811716). Interim patient-reported outcomes data are presented here. Methods: Twenty-two patients were randomized (1:1) to two treatment regimens; both arms received subcutaneous (SC) cemdisiran 200 mg every 4 weeks (Q4W) plus pozelimab 400 mg SC at a frequency of either Q4W (arm 1) or every 2 weeks (arm 2). Patients completed the Functional Assessment of Chronic Illness Therapy-Fatigue (FACIT-Fatigue) scale (score range 0-52), and the European Organization for Research and Treatment of Cancer: Quality-of-Life Questionnaire global health status (GHS)/QoL and physical function assessments (score range 0-100). Higher scores indicate a better level of functioning or GHS/QoL, or less fatigue. Results: Prior to receiving pozelimab monotherapy, mean (standard deviation [SD]) pre-treatment values were 32.3 (15.2) for FACIT-Fatigue scores, 60.6 (22.4) for GHS/QoL scores, and 70.9 (22.5) for physical functioning scores (Table 1). For the current trial, baseline values were representative of the pozelimab monotherapy that the patients were receiving prior to transitioning to the combination therapy. The mean (SD) FACIT-Fatigue score at baseline was 45.4 (5.5) for arm 1 and 45.0 (3.4) for arm 2 (Table 1). Over Weeks 2-16, the mean FACIT-Fatigue scores were 40.1-45.6 for arm 1 and 39.2-41.4 for arm 2. The mean (SD) GHS/QoL scores were well controlled and similar for both treatment arms at baseline (77.8 [14.4] for arm 1 and 77.4 [20.8] for arm 2). Over Weeks 2-16, the mean GHS/QoL scores were 68.1-77.8 for arm 1 and 54.2-78.6 for arm 2. The mean physical functioning score at baseline was 93.3 for both treatment groups (SD: 8.8 for arm 1 and 9.4 for arm 2). Over Weeks 2-16, the mean physical functioning scores were 89.6-95.6 for arm 1 and 81.0-83.3 for arm 2 (Table 1). Conclusions: Patients with PNH who transitioned from pozelimab monotherapy had improved baseline scores compared with pre-treatment for their GHS/QoL, physical functioning, and fatigue scores. Improvements in these scores were maintained by the combination treatment through to Week 16, particularly with the pozelimab Q4W and cemdisiran dose regimen. This preliminary evidence, with limited sample size, suggests that pozelimab and cemdisiran combination therapy, particularly the Q4W regimen, maintains improvements in patient fatigue, GHS/QoL, and physical functioning. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Impact of Pegcetacoplan on Paroxysmal Nocturnal Hemoglobinuria Symptoms in Patients Previously Treated with C5-Inhibitors

Background Paroxysmal nocturnal hemoglobinuria (PNH) is a rare, acquired hematologic condition associated with significant morbidity, mortality, and impact on health-related quality of life (HRQoL). In the past, the treatment paradigm for PNH consisted of blood transfusions and treatment with complement component C5 inhibitors (C5-is), eculizumab or ravulizumab. Pegcetacoplan was approved in the US and EU in May 2021 and December 2021, respectively, and is the first approved C3 inhibitor (C3-i) for US adults with PNH and EU adults with PNH who remain anemic despite stable C5 inhibitor treatment for ≥3 months. The objective of this study was to characterize the impact of pegcetacoplan on PNH symptom incidence, frequency, and severity in comparison to C5-is in the Voice of the Patient. Methods This mixed-methods study enrolled consenting, adult patients (>21 years old) with PNH in the US for telephone depth interviews. Eligible participants were required to be currently treated with and have at least 3 months of treatment experience with pegcetacoplan. Symptoms were described on a 7-point Likert scale (where 1 is "infrequent" or "very mild" and 7 is "always" or "very severe"). Results Results from 15 patients (8 female and 7 male) are reported here. Enrolled patients had a median age of 40 years (range 24 - 77) and all had received treatment with a C5-i prior to switching to pegcetacoplan. On average, patients had 5.6 months of pegcetacoplan experience following an average of 29.3 months of C5-i experience. The majority of patients (n=14) reported experiencing PNH symptoms while on C5-is, most notably fatigue (n=14), cognitive symptoms (n=12), shortness of breath (n=12), muscle weakness (n=8), and gastrointestinal problems (n=8). In addition to continuing to experience these PNH symptoms, patients described experiencing moderate to high symptom frequency and severity, with fatigue estimated as a 6.0 and 5.5, respectively, on a 7-point Likert scale, cognitive symptoms as a 5.4 and 4.8, shortness of breath as 5.1 and 4.8, muscle weakness as 5.4 and 4.6, and GI symptoms as a 5.3 and 4.9. After switching from C5-is to pegcetacoplan, fewer patients reported experiencing these residual PNH symptoms, including fatigue (n=12 on pegcetacoplan vs n=14 on C5-is), cognitive symptoms (n=11 vs n=12), shortness of breath (n=7 vs n=12), muscle weakness (n=5 vs n=8), and gastrointestinal problems (n=5 vs n=8) (Figure 1). While PNH symptoms persisted on pegcetacoplan, patients reported reduction in symptom frequency and severity, including fatigue (-21% and -33% relative reduction, respectively), cognitive symptoms (-16% and -18% relative reduction), shortness of breath (-30% and -31% relative reduction), muscle weakness (-18% and -14% relative reduction), and gastrointestinal problems (-16% and -14% relative reduction) compared to their experience on C5-is. Select patients, however, reported onset of new symptoms after switching to pegcetacoplan, including shortness of breath (n=1/15) and muscle weakness (n=1/15), though these were reported as infrequent (1.0 and 2.0, respectively) and very mild (1.0 and 1.0, respectively). Patient-reported symptom experience on C5-is and pegcetacoplan is summarized in Figure 2. Conclusions This study revealed that patients continue to experience residual PNH symptoms while on C5-is and describes improvement of patients' symptom management as it relates to incidence, frequency, and severity after switching to pegcetacoplan. Characterizing the real-world experience of PNH patients on therapy is essential to optimizing treatment options and tailoring therapies to the needs of patients. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

A Phase 2, Open-Label Study Evaluating the Safety and Efficacy of Combination Pozelimab and Cemdisiran Therapy in Patients with Paroxysmal Nocturnal Hemoglobinuria Who Switch from Eculizumab

Background: Paroxysmal nocturnal hemoglobinuria (PNH) is an ultra-rare, acquired disorder caused by a mutation in the phosphatidylinositol glycan class A (PIGA) gene, which leads to impaired expression of complement-regulating proteins on the surface of hematopoietic cells. Clinical presentation of PNH includes hemolytic anemia, hemoglobinuria, and thrombosis. Complement component C5 inhibitors such as eculizumab are part of the current standard of care for patients with PNH; however, this is an intravenous treatment which relies on nurse administration. In addition, some patients experience an incomplete response to therapy, and may still experience breakthrough hemolytic events. Pozelimab and cemdisiran are investigational agents with a subcutaneous maintenance regimen that may be self-administered. Both agents act together to inhibit terminal complement through complementary mechanisms of action. Pozelimab is a fully human monoclonal antibody inhibitor of C5, while cemdisiran is an N-acetylgalactosamine-conjugated small interfering RNA (siRNA) that suppresses liver production of C5. The efficacy and safety of the combination of pozelimab and cemdisiran is being evaluated in an ongoing, phase 2, open-label single-arm study in patients with PNH who switch from eculizumab therapy (NCT04888507). The interim safety and preliminary efficacy results are presented here. Methods: This ongoing phase 2 trial consists of a screening period (up to 42 days), a 32-week open-label treatment period (OLTP), an optional 52-week open-label extension period, and a 52-week post-treatment safety follow-up period. Patients transitioned from eculizumab therapy to the combination of pozelimab and cemdisiran over a 4-week period, and thereafter received the combination (pozelimab 400 mg and cemdisiran 200 mg) subcutaneously every 4 weeks. Results: At the time of this interim data analysis (data cut-off March 18, 2022) six patients were enrolled and five patients were ongoing. While one patient discontinued after 29 days of treatment, the remaining five patients had a treatment duration of at least 169 days, with one completing the OLTP (Day 225). Up to the time of data cut-off, no patient had a lactate dehydrogenase (LDH) level greater than 1.5 x the upper limit of normal (ULN; Figure 1) or experienced an event of breakthrough hemolysis, including the two patients who were previously treated with higher doses of eculizumab (1200 mg and 1500 mg every 2 weeks, respectively). Furthermore, all but two LDH values remained normal (≤1.0 x ULN) at all timepoints evaluated. All patients achieved hemoglobin stabilization. CH50, a measure of terminal complement activity, remained fully suppressed at 0 kIU/L throughout the study up to the time of the data cut-off. There were no serious or severe treatment emergent adverse events (TEAEs). Importantly, there were no meningococcal infections or adverse events due to potential large drug-target-drug immune complexes, or TEAEs leading to death, in this study. One patient discontinued study treatment due to two mild, non-serious TEAEs of headache at Days 2 and 16. Conclusions: These interim results from a limited data set suggest that in patients with PNH transitioning from eculizumab treatment, including patients treated with higher than standard of care doses, the combination of pozelimab and cemdisiran was generally well tolerated, providing sustained control of intravascular hemolysis without any breakthrough hemolysis events. These findings support the ongoing development of pozelimab and cemdisiran combination therapy. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Machine learning for morbid glomerular hypertrophy

A practical research method integrating data-driven machine learning with conventional model-driven statistics is sought after in medicine. Although glomerular hypertrophy (or a large renal corpuscle) on renal biopsy has pathophysiological implications, it is often misdiagnosed as adaptive/compensatory hypertrophy. Using a generative machine learning method, we aimed to explore the factors associated with a maximal glomerular diameter of ≥ 242.3 μm. Using the frequency-of-usage variable ranking in generative models, we defined the machine learning scores with symbolic regression via genetic programming (SR via GP). We compared important variables selected by SR with those selected by a point-biserial correlation coefficient using multivariable logistic and linear regressions to validate discriminatory ability, goodness-of-fit, and collinearity. Body mass index, complement component C3, serum total protein, arteriolosclerosis, C-reactive protein, and the Oxford E1 score were ranked among the top 10 variables with high machine learning scores using SR via GP, while the estimated glomerular filtration rate was ranked 46 among the 60 variables. In multivariable analyses, the R2 value was higher (0.61 vs. 0.45), and the corrected Akaike Information Criterion value was lower (402.7 vs. 417.2) with variables selected with SR than those selected with point-biserial r. There were two variables with variance inflation factors higher than 5 in those using point-biserial r and none in SR. Data-driven machine learning models may be useful in identifying significant and insignificant correlated factors. Our method may be generalized to other medical research due to the procedural simplicity of using top-ranked variables selected by machine learning.

Complement inhibition prevents glial nodal membrane injury in a GM1 antibody-mediated mouse model.

The involvement of the complement pathway in Guillain-Barré syndrome pathogenesis has been demonstrated in both patient biosamples and animal models. One proposed mechanism is that anti-ganglioside antibodies mediate neural membrane injury through the activation of complement and the formation of membrane attack complex pores, thereby allowing the uncontrolled influx of ions, including calcium, intracellularly. Calcium influx activates the calcium-dependent protease calpain, leading to the cleavage of neural cytoskeletal and transmembrane proteins and contributing to subsequent functional failure. Complement inhibition has been demonstrated to provide effective protection from injury in anti-ganglioside antibody-mediated mouse models of axonal variants of Guillain-Barré syndrome; however, the role of complement in the pathogenesis of demyelinating variants has yet to be established. Thus, it is currently unknown whether complement inhibition would be an effective therapeutic for Guillain-Barré syndrome patients with injuries to the Schwann cell membrane. To address this, we recently developed a mouse model whereby the Schwann cell membrane was selectively targeted with an anti-GM1 antibody resulting in significant disruption to the axo-glial junction and cytoplasmic paranodal loops, presenting as conduction block. Herein, we utilize this Schwann cell nodal membrane injury model to determine the relevance of inhibiting complement activation. We addressed the early complement component C2 as the therapeutic target within the complement cascade by using the anti-C2 humanized monoclonal antibody, ARGX-117. This anti-C2 antibody blocks the formation of C3 convertase, specifically inhibiting the classical and lectin complement pathways and preventing the production of downstream harmful anaphylatoxins (C3a and C5a) and membrane attack complexes. Here, we demonstrate that C2 inhibition significantly attenuates injury to paranodal proteins at the node of Ranvier and improves respiratory function in ex vivo and in vivo Schwann cell nodal membrane injury models. In parallel studies, C2 inhibition also protects axonal integrity in our well-established model of acute motor axonal neuropathy mediated by both mouse and human anti-GM1 antibodies. These data demonstrate that complement inhibition prevents injury in a Schwann cell nodal membrane injury model, which is representative of neuropathies associated with anti-GM1 antibodies, including Guillain-Barré syndrome and multifocal motor neuropathy. This outcome suggests that both the motor axonal and demyelinating variants of Guillain-Barré syndrome should be included in future complement inhibition clinical trials.

Complement component C3 and C5b-9 deposition on hypoxia reperfused endothelial cells by non-HLA antibodies against RhoGDI2: A player involved in graft failure?

Antibodies against Rho GDP-dissociation inhibitor 2 (RhoGDI2) are associated with inferior graft survival in transplant patients receiving a kidney from deceased donors. Although this suggests that these antibodies contribute to graft injury because of ischemia, it remains unknown whether they are also pathogenically involved in the process of graft loss. To study this, we firstly analyzed the IgG subclass profile of anti-RhoGDI2 antibodies in kidney transplant recipients, and whether antibody titers change over time or because of acute rejection. Next, we investigated the expression of RhoGDI2 on primary kidney and lung endothelial cells (ECs) upon hypoxia reperfusion. In addition, the complement-fixing properties of anti-RhoGDI2 antibodies were studied using imaging flow cytometry. Anti-RhoGDI2 antibodies in patients are mainly IgG1, and titers remained stable and seemed not be changed because of rejection. Antibodies against RhoGDI2, which surface expression seemed to increase upon hypoxia reperfusion, co-localized with C3 on ECs. Binding of human IgG1 monoclonal anti-RhoGDI2 antibodies as well as patient derived antibodies, resulted in complement activation, suggesting that these antibodies are complement fixing. This study suggested a potential pathogenic role of anti-RhoGDI2 antibodies in kidney graft loss. During ischemia reperfusion, the ability of these antibodies to fix complement could be one of the mechanisms resulting in tissue injury.

Anti-IL-6 therapies in central nervous system inflammatory demyelinating diseases.

Current treatments for central nervous system (CNS) inflammatory demyelinating diseases (IDDs) include corticosteroids, plasma exchange, intravenous immunoglobulin, and immunosuppressant drugs. However, some patients do not respond well to traditional therapies. In recent years, novel drugs, such as monoclonal antibodies, targeting the complement component C5, CD19 on B cells, and the interleukin-6 (IL-6) receptor, have been used for the treatment of patients with refractory CNS IDDs. Among these, tocilizumab and satralizumab, humanized monoclonal antibodies against the IL-6 receptor, have shown beneficial effects in the treatment of this group of diseases. In this review, we summarize current research progress and prospects relating to anti-IL-6 therapies in CNS IDDs.