Total Hemolytic Complement Activity Research Articles

Performance of classification and diagnostic criteria for IgG4-related disease and comparison of patients with and without IgG4-related disease

IgG4-related disease (IgG4-RD) was recently described in Japan. It is characterised by extensive organ involvement with tissue fibrosis. We assessed the performance of the 2019 American College of Rheumatology and European League Against Rheumatism (ACR/EULAR) classification criteria and the 2020 revised comprehensive diagnostic (RCD) criteria as well as differences between patients with and without IgG4-RD. In this retrospective, single-centre study of 50 patients admitted with suspected IgG4-RD, we evaluated the sensitivity and specificity of both criteria. We also compared clinical characteristics and laboratory data of patients with IgG4-RD (n = 42) and patients without IgG4-RD (n = 8). The ACR/EULAR classification criteria had 88.1% sensitivity and 87.5% specificity for IgG4-RD diagnosis. The RCD criteria had 100% sensitivity and 50% specificity. Patients with IgG4-RD had significantly more affected organs (p = 0.002). Patients with a single affected organ and IgG4-RD had significantly higher serum IgG4/IgG ratios (p = 0.027), lower serum C-reactive protein levels (p = 0.020), and lower total haemolytic complement activity (p = 0.044) than those without IgG4-RD. The ACR/EULAR classification criteria have high specificity and the RCD criteria have high sensitivity for diagnosing IgG4-RD. The number of affected organs is important for diagnosing IgG4-RD.

Efficacy and Safety of Biosimilar Candidate ABP 959 As Compared with Eculizumab Reference Product in Paroxysmal Nocturnal Hemoglobinuria

Introduction: ABP 959 is a biosimilar candidate to eculizumab reference product (RP). Eculizumab is a humanized recombinant immunoglobulin antibody that binds to the human complement component 5 protein to inhibit progression of complement cascades and is approved for the treatment of paroxysmal nocturnal hemoglobinuria (PNH), atypical hemolytic uremic syndrome, generalized myasthenia gravis, and neuromyelitis optica spectrum disorder. Methods: This multicenter, randomized, double-blind, active-controlled, 2-period crossover study compared the efficacy and safety of ABP 959 and eculizumab RP in adult patients with PNH who were stable on eculizumab 900 mg IV every 14 ± 2 days. The primary objective of this study was to evaluate the efficacy of ABP 959 compared to eculizumab RP based on control of intravascular hemolysis, with a secondary objective to compare their safety, pharmacokinetics (PK), and immunogenicity. Results from Period 1 of the crossover study are presented here. Patients were randomized 1:1 to receive either ABP 959 at 900 mg IV for 52 weeks in Period 1 followed by eculizumab RP 900 mg IV for 26 weeks in Period 2 (ABP 959/RP) or eculizumab RP 900 mg IV for 52 weeks followed by ABP 959 at 900 mg IV for 26 weeks (RP/ABP 959) (Figure). The primary efficacy endpoint for the parallel comparisons was hemolysis, measured by lactate dehydrogenase (LDH) at week 27; clinical similarity between treatments was assessed by comparing the 1-sided 97.5% upper confidence interval (CI) limit for the geometric least squares (LS) mean ratio between ABP 959 and eculizumab RP, with a non-inferiority (NI) margin of 2.873. To assess the robustness of the primary analysis result, a sensitivity analysis was conducted using a subset of randomized patients who did not experience any important protocol deviation affecting their primary efficacy evaluation. Secondary endpoints included total complement (50% total hemolytic complement activity [CH50]), LDH-time profile, number of units of packed red blood cells (pRBCs) transfused, safety, immunogenicity, and PK area under the serum concentration-time curve (PK AUC) of ABP 959 and eculizumab RP. Results: Forty-two patients (20 in the ABP 959/RP group; 22 in the RP/ABP 959 group) were randomized and treated in Period 1 across 25 centers. Forty-one (97.6%) patients (20 [100%] in ABP 959/RP and 21 [95.5%] in RP/ABP 959) completed Period 1 dosing. Baseline demographic and disease characteristics were comparable between the 2 treatment groups - mean (SD) age: 50.2 (16.6) years; time since original PNH diagnosis: 128.3 (116.1) months; duration of pre-study treatment: 68.4 (39.8) months. Results for week 27 LDH geometric LS means are presented below (Table). The ratio of the geometric LS means of LDH (ABP 959/RP) was 1.0628, with a 1-sided 97.5% upper CI of 1.1576 contained within the prespecified NI margin of 2.873, establishing similarity in clinical efficacy between ABP 959 and eculizumab RP in the parallel comparison; 95% CI was 0.9758 to 1.1576. Results from the sensitivity analysis were identical with results from the primary efficacy analysis. Total complement (CH50), total hemoglobin, serum-free hemoglobin, haptoglobin, bilirubin, and type III erythrocytes values were generally stable over time and results were comparable between the 2 treatment groups. The overall LDH profile and the mean number of units of pRBCs transfused during the study period were also comparable. Total PK AUC from week 13 to week 15 were similar between the 2 treatment groups. A comparable safety profile was observed in Period 1 between treatment groups, with only a single patient (4.5%) in the eculizumab RP group discontinuing the study due to any adverse event (grade 2 non-serious asthenia and grade 2 non-serious fatigue). The nominal difference in serious adverse event incidence was determined to be not clinically meaningful. Additionally, immunogenicity results through Period 1 were similar between ABP 959 and eculizumab RP groups. No patients in either treatment group tested positive for binding antidrug antibodies (ADAs), neutralizing ADAs, or treatment-boosted ADAs. Conclusions: Similarity in clinical efficacy was established between ABP 959 and eculizumab RP in the primary endpoint of hemolysis, measured by LDH at week 27. Additionally, comparable safety, PK, and immunogenicity results support the conclusion of clinical similarity between ABP 959 and eculizumab RP. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Dynamics of cervical mucus immune factors in patients with candida and mycoplasma infection before and after topical laser therapy

The article presents an analysis of the content and dynamics of indicators of humoral factors in the cervical mucus of women with inflammatory diseases of the lower reproductive tract caused by combined candida and mycoplasma infection. It has been shown that the use of topical laser therapy (wavelength 635±10 nm) with variable pulse generation leads to normalization of the sIgA, IgA production, restoration of the activity of C1, C2, C3, C4, and C5 complement components, total haemolytic complement activity, and the balance of cytokines in the cervical mucus.

The Pharmacokinetics, Pharmacodynamics, and Safety of Intravenous Pegcetacoplan Treatment in Healthy Subjects

Background: The complement cascade is part of innate immunity and is involved in multiple inflammatory processes and implicated in several diseases. Pegcetacoplan (PEG) is a pegylated, cyclic peptide that binds to complement protein C3 and is a broad inhibitor of the complement cascade. Subcutaneous (SC) dosing of PEG has demonstrated efficacy in the treatment of chronic conditions, such as paroxysmal nocturnal hemoglobinuria (PNH) and was recently approved by the FDA for the treatment of PNH in adults. Intravenous (IV) PEG administration may allow for more rapid and robust reduction of uncontrolled complement activation, especially in an acute setting, such as an acute hemolytic episode in PNH.Aims: To determine the safety, pharmacokinetics (PK), and pharmacodynamics (PD) of IV PEG in acetate-buffered saline treatment in a Phase 1 single ascending dose study (ACTRN12616000700437) in healthy subjects.Methods: On Day 1, four cohorts with PEG doses (200mg, 600mg, 1500mg, 2300mg) received a single bolus of PEG IV (or matching placebo) administered over 30min. Blood samples for PK analyses of PEG concentration and PD analyses of alternative complement pathway hemolytic activity (AH50), total complement hemolytic activity (CH50), C3 and C3a levels were collected at 15, 30, and 60min, 4, 8, 12, and 24hrs, and Days 3, 4, 5, 6, 7, 8, 15, 22, 29, and 43. Subjects were monitored during a safety period from Day 2 to 8 by physical examination, ECG, hematology, serum chemistry, monitoring for injection site reaction and treatment emergent adverse events (TEAEs). Follow-up safety assessments were performed on Days 15, 22, 29, and 43.Results: Twenty subjects were enrolled and allocated 4:1 to PEG or placebo per cohort (PEG-200mg, n=4; PEG-600mg, n=4; PEG-1500mg, n=4; PEG-2300mg, n=4; pooled placebo, n=4). Following a single IV dose, peak concentration (C max) of PEG was observed at 1hr post-dose (infusion start) for most cohorts (mean serum concentration: PEG-200mg, 61μg/mL; PEG-600mg, 193μg/mL; PEG-2300mg, 708μg/mL) except PEG-1500mg (occurred at 4hrs, 542μg/mL). PEG concentration at the end of infusion was similar to the observed C max. PEG concentration declined in a mono-exponential manner, with a terminal elimination half-life ranging from 200 to 285 hrs (Figure). Total body clearance of PEG after IV administration was similar across cohorts.Early, immediate decreases in mean AH50 values were detected within 1hr in all PEG cohorts, with 1500 and 2300mg doses decreasing AH50 to undetectable levels (Figure). Decreases in mean AH50 values were maintained for at least 12, 72, 144 and 168hrs after single doses of 200, 600, 1500 and 2300mg PEG, respectively.All PEG groups had an initial rapid decrease within 1hr in mean C3a levels, with all dose groups having trough mean C3a levels within 24hrs of dosing. Dose related decreases in mean C3a were not observed, all doses recorded a max mean decrease of 47% to 57%. No changes seen with placebo for C3a. C3 and CH50 results will be forthcoming.Of the twenty subjects included in the study, 11 (55.0%) experienced a treatment-related adverse event (TEAE). The most common TEAEs in the PEG group were headache, (n=6, 37.5%); upper respiratory infections attributed to seasonal viral infection (n=2, 12.5%); diarrhea (n=2, 12.5%). No serious adverse events, deaths, or severe TEAEs occurred. One subject (5.0%) in the PEG-2300mg cohort experienced a moderate TEAE (infusion-related reaction, dizziness, clamminess, nausea) that led to study discontinuation.Conclusions: These results suggest that administration of IV PEG in a sodium acetate solution has a favorable safety profile and effectively increases PEG serum concentrations while decreasing complement activity within the first hour post-dose in healthy subjects. Although the safety and efficacy of SC PEG treatment has been demonstrated in patients with PNH, IV PEG administration could serve as a useful therapeutic option for patients with a need for rapid control of complement activity. While this formulation is different than the commercially available PEG (EMPAVELI), which is administered SC and is suspended in sorbitol, the results suggest that IV PEG is well tolerated and provides the grounds for future investigations of IV PEG administration. [Display omitted] DisclosuresGrossi: Apellis Pharmaceuticals: Current Employment, Current equity holder in publicly-traded company. Yeh: Apellis Pharmaceuticals, Inc.: Current Employment, Current equity holder in publicly-traded company. Xu: Apellis Pharmaceuticals: Current Employment. Deschatelets: Apellis Pharmaceuticals: Current Employment, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties. OffLabel Disclosure:Pegcetacoplan, a subcutaneously administered C3-inhibitor that was recently approved by the US FDA for the treatment of PNH, controls IVH and prevents EVH. While subcutaneous pegcetacoplan is safe and effective, the aim of this study was to determine the safety, pharmacokinetics, and pharmacodynamics of IV pegcetacoplan in acetate-buffered saline treatment, different from current FDA approved formulation.

Total Haemolytic Complement Activity at Diagnosis as an Indicator of the Baseline Activity of Antineutrophil Cytoplasmic Antibody-associated Vasculitis.

The total haemolytic complement activity (CH50) assay evaluates the functioning of the complement system Accumulating evidence indicates that the activation of the complement system plays a critical role in the pathogenesis of antineutrophil cytoplasmic antibody-associated vasculitis (AAV) Therefore, this study aimed to investigate whether CH50 levels at diagnosis could reflect the baseline activity of AAV. This retrospective study included 101 immunosuppressive drug-naïve patients with AAV At diagnosis, all patients underwent clinical assessments for disease activity, including measurement of the Birmingham Vasculitis Activity Score (BVAS) and Five Factor Score (FFS), and laboratory evaluations, such as tests for CH50, C3, and C4 levels The association between CH50 levels and disease activity was determined. The median BVAS and FFS at diagnosis were 120 and 10, respectively, whereas the median CH50 level was 604 U/mL There was a negative correlation between the CH50 level and BVAS (r=-0241; p=0015) A CH50 cut-off value of 621 U/mL was used to classify the patients into two groups: patients with CH50 levels <621 U/mL (low-CH50 group) and those with CH50 levels ≥621 U/mL (high-CH50 group) The low-CH50 group had a higher proportion of patients with high disease activity, based on the BVAS, than the high-CH50 group (525% vs 238%, p=0004) Additionally, the low-CH50 group exhibited a lower relapse-free survival rate than the high-CH50 group; however, this difference was not statistically significant (p=0082). Low CH50 levels at diagnosis may reflect high baseline activity of AAV.

Serum phosphatidylserine-specific phospholipase A1 as a novel biomarker for monitoring systemic lupus erythematosus disease activity.

To assess the utility of serum levels of phosphatidylserine-specific phospholipase A1 (PS-PLA1 ), a lipase involved in the production of lysophosphatidylserine with multi-immunomodulatory effects, in systemic lupus erythematosus (SLE). Serum PS-PLA1 was measured in 161 patients with SLE (including 54 untreated patients), 80 disease controls (35 active rheumatoid arthritis [RA], 23 Sjögren's syndrome [SS], and 22 systemic sclerosis [SSc]), and 237 healthy controls. Serum PS-PLA1 was significantly higher in SLE patients than in healthy controls, RA and SS patients. Although PS-PLA1 was significantly elevated in SSc and SS patients compared with healthy controls, PS-PLA1 was significantly higher in untreated SLE patients than in treated SLE patients and disease control patients. Receiver operating characteristic analysis revealed that a cut-off value of 18.2ng/mL distinguished untreated SLE from disease control, with sensitivity and specificity of 71.4% and 57.5%, respectively. PS-PLA1 was significantly correlated with SLE Disease Activity Index (SLEDAI) and immunoglobulin G (IgG), and inversely correlated with white blood cell counts, lymphocyte counts, total complement hemolytic activity (CH50), complements C3, and C4 in SLE patients overall. Stepwise multiple regression identified SLEDAI, CH50, and IgG as significant parameters. In SLEDAI-based disease activity groups, PS-PLA1 was significantly higher in SLE patients with high disease activity than in those with low disease activity. PS-PLA1 decreased significantly in parallel with SLEDAI in 35 SLE patients whose paired serum samples were available pre- and post-treatment. Serum PS-PLA1 is associated with disease activity of SLE, indicating its possible use as a biomarker for monitoring SLE disease activity.

PF347 PHARMACOKINETIC AND PHARMACODYNAMIC SIMILARITY OF ABP 959 WITH ECULIZUMAB: RESULTS FROM A RANDOMIZED, DOUBLE-BLIND, SINGLE-DOSE, PARALLEL-GROUP STUDY IN HEALTHY SUBJECTS

Background: ABP 959 is being developed as a biosimilar candidate to eculizumab. Eculizumab is a recombinant humanized IgG2/4 chimeric monoclonal antibody that binds to the human C5 complement protein (C5), thereby inhibiting the complement cascade. It is indicated for the treatment of patients with generalized myasthenia gravis (gMG), paroxysmal nocturnal hemoglobinuria (PNH) to reduce hemolysis, and atypical hemolytic uremic syndrome (aHUS) to inhibit complement-mediated thrombotic microangiopathy. ABP 959 has been evaluated to be analytically similar to eculizumab. Aims: To demonstrate pharmacokinetic (PK) and pharmacodynamic (PD) similarity of ABP 959 relative to the reference product (RP) eculizumab. Methods: This was a randomized, double-blind, single-dose, 3-arm, parallel-group study in healthy adult male subjects, between 18 to 45 years of age, with a body mass index of 18 to 30.0 kg/m2, who were randomized to receive a 300-mg IV infusion of ABP 959, or FDA-licensed eculizumab (eculizumab US), or EU-authorized eculizumab (eculizumab EU). Serum samples for PK and PD evaluations were collected over 56 days. Primary objectives were to demonstrate PK and PD equivalence of ABP 959 versus eculizumab US and ABP 959 versus eculizumab EU, as assessed by area under the total serum concentration—time curve (AUC) from time 0 extrapolated to infinity (AUCinf) and an area between the effect curve (ABEC) of 50% total hemolytic complement activity (CH50). Secondary PK endpoints included AUC from time 0 to the time of the last observed quantifiable concentration (AUClast) and maximum observed concentration (Cmax). Pre-specified equivalence criterion for the primary PK and PD parameters was 90% confidence interval (CI) for geometric mean (GM) ratios within 0.80 to 1.25. Other secondary endpoints included the safety, tolerability, and immunogenicity of the investigational products (IPs). Results: A total of 219 subjects were enrolled (ABP 959, n = 71; eculizumab US, n = 74; eculizumab EU, n = 74). The GM of PK and PD parameters following a single IV infusion of IP, were similar between ABP 959 vs. eculizumab US and ABP 959 vs. eculizumab EU. The GM of AUCinf (h.μg/mL) was: 18996.3 for ABP 959; 19777.5 for eculizumab US and 18921.1 for eculizumab EU. The GM of ABEC of CH50 (%∗h) was 17724.5 for ABP 959; 16549.4 for eculizumab US and 16361.1 for eculizumab EU. For both PK and PD comparisons of ABP 959 with eculizumab US and ABP 959 with eculizumab EU, the 90% CIs of the GM ratios were fully contained within the bioequivalence criteria of 0.80 to 1.25 for all PK (AUCinf, Cmax, AUClast) and PD (ABEC of CH50) evaluations, confirming the PK and PD bioequivalence between ABP 959 and eculizumab US and ABP 959 and eculizumab EU. Additionally, eculizumab US and eculizumab EU were similar in PK and PD assessments. There were no deaths or treatment-emergent adverse events (TEAEs) leading to study discontinuation. Five subjects experienced 8 SAEs; only the SAE of headache was considered probably related to study drug. The incidence of overall TEAEs or any single category of TEAE was similar across treatment groups. A total of 19 (8.8%) subjects had positive binding ADAs over the course of the study; the incidence of binding ADAs was similar across treatments. No subjects developed neutralizing antibodies. Summary/Conclusion: This phase 1 study demonstrated PK and PD bioequivalence of ABP 959 to eculizumab RP. Safety and immunogenicity profiles were similar between ABP 959 and eculizumab RP.

Clinical outcomes in patients with systemic lupus erythematosus treated with belimumab in clinical practice settings: a retrospective analysis of results from the OBSErve study in Switzerland.

To describe patterns of systemic lupus erythematosus (SLE) care and the clinical effectiveness of belimumab plus standard of care therapy in a real-world clinical setting in Switzerland. This multicentre, observational, retrospective cohort study included adults with SLE who initiated belimumab as part of their usual care at least six months before data analysis. The primary outcome was the overall clinical response, assessed by a physician on a Physician&rsquo;s Global Assessment-like scale, to six months&rsquo; treatment with belimumab. Secondary outcomes included improvement in disease activity, SLE manifestations and changes in corticosteroid use. 53 patients (81% female) from three hospitals were included. At index (belimumab initiation), 23 patients (43%) had mild, 23 (43%) had moderate, and 7 (13%) had severe SLE. Overall improvement in disease activity in patients receiving belimumab was: &ge;80% in 6 patients (11%), &ge;50% in 12 (23%), &ge;20% in 31 (58%), &lt;20% in 13 (25%), and no improvement in 9 (17%). Mean Safety of Estrogens in Lupus Erythematosus National Assessment-SLE Disease Activity Index score decreased from 8.0 at index to 3.6 at six months post index in the 27 patients assessed. In addition, a &ge;50% improvement in arthritis, fatigue, rash, low complement (C3, C4 or total haemolytic complement activity), and anti-double-stranded deoxyribonucleic acid antibody levels was experienced six months post index by 10 (38%), 3 (16%), 6 (38%), 2 (12%) and 4 (16%) patients who presented the manifestations at index respectively. At index, 41 patients (77%) received oral corticosteroids at a mean dose of 11.6 mg/day, which decreased to 5.9 mg/day at six months post index. Of the 31 patients receiving a high dose of corticosteroids (&ge;7.5 mg/day) at index, 18 required &lt;7.5 mg/day and a further two discontinued corticosteroids at six months post index. This study provides real-world insight into belimumab use in clinical practice in Switzerland. In line with findings from other countries, Swiss patients with SLE who received belimumab demonstrated clinical and serological improvements in SLE and a reduction in corticosteroid use after six months of treatment.

Significant Fluctuations in Total Complement Hemolytic Activity as a Nutritional Parameter After Transition from Hemodialysis to Online Hemodiafiltration

Significant Fluctuations in Total Complement Hemolytic Activity as a Nutritional Parameter After Transition from Hemodialysis to Online Hemodiafiltration

High fish oil diet promotes liver inflammation and activates the complement system.

Diets rich in n-3 polyunsaturated fatty acid (n-3 PUFA) fish oil (FO) have beneficial effects in obesity‑associated metabolic disease. However, contradictory roles in inflammatory disease intervention have been reported. Our previous work revealed that a high‑FO diet promoted myeloid cell differentiation by modifying the bone marrow microenvironment; however, its effects on liver inflammation and complement system activation remain unknown. By performing ELISA, reverse transcription‑quantitative polymerase chain reaction, flow cytometry and histology on mice fed with high‑FO and low‑fat diets, the present study demonstrated that a 4‑week high‑FO diet promoted liver inflammation in mice without affecting body or liver weight. The livers of high‑FO diet mice exhibited increased infiltration of T cells and CD11b+ Gr‑1+ myeloid cells. Additionally, a higher level of IL‑1β and MCP‑1 mRNA expression was detected, suggesting that the high‑FO diet promoted liver inflammation. Further experiments indicated that the high‑FO diet increased the total hemolytic complement activity (CH50), promoted the production of the membrane attack complex and increased the levels of various complement proteins invivo, including complement components C3, C4b, C1qb and factor B. Furthermore, higher concentrations of triglyceride were detected in the peripheral blood of high‑FO diet mice, indicating the potential protective roles of n‑3 PUFAs in FO against lipotoxicity in hyperlipidemia. Collectively, the present study demonstrated that high FO intake induced inflammation and activated the complement system in the liver. However, further study is required to determine the exact mechanisms.

The complement system in hypertension and renal damage in the Dahl SS rat.

Evidence indicates the immune system is important in development of hypertension and kidney disease. In the Dahl Salt‐Sensitive (SS) rat model, lymphocytes play a role in development of hypertension and kidney damage after increased sodium intake. Recent transcriptomic analyses demonstrate upregulation of the innate immune complement system in the kidney of Dahl SS rat fed a high‐salt diet, leading us to hypothesize that inhibition of complement activation would attenuate development of hypertension and kidney damage. Male Dahl SS rats on a low salt (0.4% NaCl) diet were instrumented with telemeters for continuous monitoring of arterial blood pressure. Animals received saline vehicle (Control) or sCR1, a soluble form of endogenous Complement Receptor 1 (CR1; CD35) that inhibits complement activation. At Day 0, rats were switched to high salt (4.0% NaCl) diet and assigned to sCR1 (15 mg/kg per day) or Control groups with daily ip injections either days 1–7 or days 14–18. Urine was collected overnight for determination of albumin excretion. Treatment with sCR1, either immediately after high‐salt diet was initiated, or at days 14–18, did not alter development of hypertension or albuminuria. The sCR1 dose effectively inhibited total hemolytic complement activity as well as C3a generation. High salt caused an increase in message for complement regulator Cd59, with minimal change in Crry that controls the C3 convertase. Thus, innate immune complement activation in the circulation is not critical for development of hypertension and kidney damage due to increased sodium intake, and therapeutic manipulation of the complement system is not indicated in salt‐sensitive hypertension.

Micronutrient Supplementation does not Change Complement System Response to Heavy Training

We aimed to examine the micronutrient supplementation effect on complement system activity after heavy training. 24 male firefighters were randomly divided into supplemented and placebo groups, and tested for immunology-related parameters using venous blood samples in the fasting state pre- and post-5 weeks of nutritional supplementation. C3 and C4 complement components were determined in a nephelometer from immune complexes formed through specific human antisera and total haemolytic complement activity (CH100) was determined by enzyme immunoassay. Differences between pre- and post-supplementation were observed only for CH100 on placebo group (p=0.004; mean diff −26.92; 95%CI −43.58 to −10.25) and no interaction, treatment or time effects were observed for C3 and C4. Although interaction accounted for 8.8% of the total variance in CH100 (with time effect pre- vs post-accounting for 19.5% of the total variance), the treatment effect (supplemented vs placebo) was not significant. The absence of effects on the complement system response to supplementation during heavy training could be justified by the fact that: (i) nutritional supplements do not improve humoral innate immunity in well-fed subjects; (ii) selected supplements unlikely improve the innate immune system in situations of adequate nutritional status; and/or (iii) selected doses of supplementation were not sufficient to elicit immune changes.

Variable Eculizumab Clearance Requires PharmacodynamicMonitoring to Optimize TherapyforThrombotic Microangiopathy after HematopoieticStem Cell Transplantation.

Thrombotic microangiopathy (TMA) after hematopoietic stem cell transplantation (HSCT) associated with terminal complement activation, as measured by elevated plasma terminal complement (sC5b-9) concentrations, has a very high mortality. The complement inhibitor eculizumab may be a therapeutic option for HSCT-associated TMA. We examined the pharmacokinetics and pharmacodynamics (PK/PD) of eculizumab in children and young adult HSCT recipients with TMA and activated complement to determine drug dosing requirements for future efficacy trials. We analyzed prospectively collected laboratory samples and clinical data from 18 HSCT recipients with high-risk TMA presenting with complement activation who were treated with eculizumab. We measured eculizumab serum concentrations, total hemolytic complement activity, and plasma sC5b-9 concentrations. Population PK/PD analyses correlated eculizumab concentrations with complement blockade and clinical response and determined interindividual differences in PK parameters. We also compared transplant survival in patients treated with eculizumab (n=18) with patients with the same high-risk TMA features who did not receive any targeted therapy during a separate prospective observational study (n=11). In the PK analysis, we found significant interpatient variability in eculizumab clearance, ranging from 16 to 237mL/hr/70kg in the induction phase. The degree of complement activation measured by sC5b-9 concentrations at the start of therapy, in addition to actual body weight, was a significant determinant of eculizumab clearance and disease response. Sixty-one percent of treated patients had complete resolution of TMA and were able to safely discontinue eculizumab without disease recurrence. Overall survival was significantly higher in treated subjects compared with untreated patients (56% versus 9%, P=.003). Complement blocking therapy is associated with improved survival in HSCT patients with high-risk TMA who historically have dismal outcomes, but eculizumab pharmacokinetics in HSCT recipients differ significantly from reports in other diseases like atypical hemolytic uremic syndrome and paroxysmal nocturnal hemoglobinuria. Our eculizumab dosing algorithm, including pr-treatment plasma sC5b-9 concentrations, patient's actual body weight, and the first eculizumab dose (mg), accurately determined eculizumab concentration-time profiles for HSCT recipients with high-risk TMA. This algorithm may guide eculizumab treatment and ensure that future efficacy studies use the most clinically appropriate and cost-efficient dosing schedules.

Eculizumab Therapy in Children with Severe Hematopoietic Stem Cell Transplantation–Associated Thrombotic Microangiopathy

We recently observed that dysregulation of the complement system may be involved in the pathogenesis of hematopoietic stem cell transplantation–associated thrombotic microangiopathy (HSCT-TMA). These findings suggest that the complement inhibitor eculizumab could be a therapeutic option for this severe HSCT complication with high mortality. However, the efficacy of eculizumab in children with HSCT-TMA and its dosing requirements are not known. We treated 6 children with severe HSCT-TMA using eculizumab and adjusted the dose to achieve a therapeutic level >99 μg/mL. HSCT-TMA resolved over time in 4 of 6 children after achieving therapeutic eculizumab levels and complete complement blockade, as measured by low total hemolytic complement activity (CH50). To achieve therapeutic drug levels and a clinical response, children with HSCT-TMA required higher doses or more frequent eculizumab infusions than currently recommended for children with atypical hemolytic uremic syndrome. Two critically ill patients failed to reach therapeutic eculizumab levels, even after dose escalation, and subsequently died. Our data indicate that eculizumab may be a therapeutic option for HSCT-TMA, but HSCT patients appear to require higher medication dosing than recommended for other conditions. We also observed that a CH50 level ≤ 4 complement activity enzyme units correlated with therapeutic eculizumab levels and clinical response, and therefore CH50 may be useful to guide eculizumab dosing in HSCT patients as drug level monitoring is not readily available.

Relationship among antineutrophil cytoplasmic antibody, blood urea nitrogen and complement in patients with eosinophilic granulomatosis polyangiitis (Churg-Strauss syndrome)

Eosinophilic granulomatosis with polyangiitis (EGPA), also known as Churg-Strauss syndrome, is an antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis characterized by a history of asthma, hypereosinophilia. The prevalence of ANCA in EGPA is less common than in other ANCA-associated vasculitis. Increasing evidence of complement activation in the pathogenesis of ANCA-associated vasculitis has been provided by studies in animal models. We examined EGPA patients with cutaneous manifestations as an initial sign and investigated the correlations among clinical, serological and histopathological findings. We focused on differences among ANCA, blood urea nitrogen and complement levels such as complement 3 (C3), C4 and total complement hemolytic activity (CH50). We retrospectively investigated the records of 22 patients (11 male and 11 female) with EGPA admitted to our hospital from 1997-2012. Ten of the 22 patients (46%) were positive for serum myeloperoxidase (MPO)-ANCA. In contrast, all the patients were negative for serum proteinase 3 ANCA. There was a significantly positive correlation between serum CH50 and C4 levels in patients with EGPA. Serum blood urea nitrogen (BUN) levels differed significantly between MPO-ANCA-positive and -negative patients. Serum CH50 levels were higher in MPO-ANCA-positive patients compared to negative patients. Serum BUN levels were higher in elevated CH50 patients compared to normal and low CH50-negative patients. We propose that positive findings for MPO-ANCA with CH50 high activity may be a risk factor for developing renal insufficiency. Assuming there are correlations between the presence of ANCA and complements, earlier diagnosis based on initial efficacious treatment for EGPA.

Laboratory Features Throughout the Disease Course of Influenza A (H1N1) Virus Infection

Influenza has emerged every year but a complete profile of laboratory indices throughout the disease course remains unknown. Clinical data was collected from 28 confirmed cases of the pandemic influenza H1N1 2009. The levels of serum iron (Fe), carbon dioxide combining power (CO2-CP), total complement hemolytic activity (CH50), C-reactive protein (CRP), and white blood cell (WBC) and differential count were analyzed. Major laboratory abnormalities recokled for patients upon admission were lymphopenia (96.4%), eosinopenia (50.0%), hypoferremia (92.9%), decreased levels of serum CO2-CP (60.7%), increased levels of serum CRP (84.6%) and serum CH50 (71.4%). The serum iron and CO2-CP concentration and the counts for lymphocytes, eosinophils, and basophils were significantly increased four days after sickness was noticed compared with the first three days of illness (p < 0.05). The total WBC and neutrophil counts were significantly decreased four days after onset of illness compared with the counts over the first three days (p < 0.05). The monocyte count and CRP concentration was significantly decreased 7 days after onset of illness compared with first 3 days after illness onset (p < 0.05). The serum CH50 concentrations were higher than the normal range during disease course and significantly elevated 7 days after onset of illness compared with the first 6 days after illness onset (p < 0.05). The serum levels of iron, CO2-CP, CH50, CRP, and WBC and differential count Were significantly varied during the whole pandemic influenza (H1N1) 2009. The development of WBC count in patients with influenza may be an effective predictor for severity of illness.

Beneficial Effect of the Polysaccharides from Bupleurum smithii var. parvifolium on “Two-Hit” Acute Lung Injury in Rats

Radix Bupleuri is a traditional Chinese herb frequently used in the prescriptions for the treatment of inflammatory diseases. This study was to determine whether the crude polysaccharides isolated from the roots of Bupleurum smithii var. parvifolium (BPs) had beneficial effects on a "two-hit" acute lung injury model in rats. A two-hit lung injury model characterized by hemorrhagic shock followed by reperfusion and intratracheal lipopolysaccharide (1mgkg(-1)) administration was established in Wistar rats. Drugs and saline were administered 30min after the start of resuscitation. The two-hit acute lung injury model was successfully established. After oral administration, all BPs groups ameliorated pathological injury with lessened complement C3c deposit in lung. BPs 10 and 20mgkg(-1) diminished the ratio of wet-to-dry weight. In bronchoalveolar lavage (BAL) fluids, the protein concentration, the leukocytes counts, and the lung myeloperoxidase were significantly reduced. BPs also mediated the decreases in interleukin-6 and tumor necrosis factor-α in BAL fluids and in sera. Furthermore, BPs 20mgkg(-1) decreased the total complement hemolytic activity. BPs had beneficial effects on two-hit acute lung injury. The mechanisms of BPs on inflammatory disease might relate to its inhibitory effect on increased production of proinflammatory mediators and on overactivation of complement.

Levels of total hemolytic complement activity in paired maternal newborn sheep sera.

Summary The levels of total hemolytic complement (CH50) in sera obtained from newborn lambs and their corresponding dams were compared. The mean concentration of CH50 units/ml. in sera from 23 lambs was 71.1 with a standard deviation of 18.9, whereas, for the ewes it was 189.5 ± 29.6. The differences between the mean neonatal and maternal CH50 concentrations were highly significant (P « 0.001). In each case CH50 titers were higher in maternal than newborn sera, with the ratio of ewe: lamb serum CH50 values ranging from 1.48 to 5.31 with a mean of 2.87. Anticomplementary activity was not detected in lamb sera. Zusammenfassung Hämolytische Komplementaktivität in Seren aus Mutterschaf-Lamm-Paaren Seren wurden von neugeborenen Lämmern und deren Müttern gewonnen und die Aktivität des hämolytischen Komplements (CH50) darin bestimmt. Die mittlere Konzentration in CH50 Einheiten/ml betrug in den Seren von 23 Lämmern 71,1 mit einer Standardabweichung von 18,9; die Werte der Mutterschafe waren 189,5 ± 29,6. Der Unterschied in den mittleren Konzentrationen von Lämmern und Mutterschafen war hoch signifikant (P < 0,001). Immer waren die CH50 Titer der Mütter höher als bei den Neugeborenen, wobei das Verhältnis Mutterschaf: Lammserum von 1,48 bis 5,31 variierte mit einem Mittelwert von 2,87. Das Lämmerserum hatte keine antikomplementäre Aktivität. Résumé Activité hémolytique du complément dans des sérums de paires brebis-agneau Des sérums d'agneaux nouveau-nés et de leurs mères ont été pris et on a déterminé l'activité du complément hémolytique (CH50). La concentration moyenne en CH50 unités/ml fut de 71,1 dans les sérums de 23 agneaux avec une déviation standard de 18,9; les valeurs des brebis furent 189,5 ± 29,6. La différence des concentrations moyennes entre agneaux et brebis fut hautement significative (P < 0,001). Les titres CH50 des mères furent toujours plus élevés que ceux des nouveau-nés alors que le rapport sérum brebis-agneau varia de 1,48 à 5,31 avec une valeur moyenne de 2,87. Le sérum des agneaux n'a eu aucune activité anticomplémentaire. Resumen Actividad hemolítica del complemento en sueros sanguíneos de parejas oveja madre-cordero Se obtuvieron sueros sanguíneos de corderos recién nacidos y de sus madres, valorádose en ellos la actividad del complemento hemolítico (CH50). La concentración media en unidades CH50/ml importó en los sueros de 23 corderos 71,1 con una desviación estandar de 18,9; los valores de las ovejas madres eran 189,5 ± 29,6. La diferencia en las concentraciones de corderos y ovejas madres era altamente significante (P < 0,001). Los títulos CH50 de las madres eran siempre mayores que los de los recien nacidos, variando la relaciön suero sanguíneo de oveja madre: cordero de 1,48 a 5,31 con un valor medio de 2,87. El suero de cordero no tenía ninguna actividad anticomplementaria.

Un exceptionnel déficit héréditaire en fraction C3 du complément dépisté par électrophorèse des protéines sériques

We report the case of a 5-years old child referred to the pediatric clinic due to a prolonged history of recurrent otitis. Initial immunologic investigation was normal but a severe C3 complement deficiency was detected by the absence of beta 2-globulin protein fraction using serum protein capillary electrophoresis. C3 was not detected in serum and total complement haemolytic activity was decreased. His mother and father had half of the C3 normal plasma level and a heterozygous mutation of the C3 gene. The diagnosis of hereditary deficiency of the third complement component (C3) with compound heterozygous mutation of the gene was made. This defect in complement protein C3, described to date in only 20 families in the world, is associated with repeated infections. The child is treated with oracillin with relatively good control of symptoms.

Human C3 Deficiency Associated with Impairments in Dendritic Cell Differentiation, Memory B Cells, and Regulatory T Cells

Primary C3 deficiency, a rare autosomal inherited disease (OMIM 120700), was identified in a 2-year-old male suffering from recurrent pyogenic infections from early infancy with undetectable total complement hemolytic activity (CH50) and C3 values. The nonconsanguineous parents and the two patients' two siblings had 50% normal serum C3 concentration. The molecular abnormality associated a paternal allele coding C3 with the missense mutation p.Ser(550)Pro and an apparently null maternal allele, with production of a defective protein that could no longer be secreted. Vaccination of the child did not induce a long-term Ab response. Accordingly, switched memory IgD(-)CD27(+) B cells were barely detected, amounting to only 2.3% of peripheral blood CD19(+) cells. Cells were significantly defective in stimulating alloreactive responses. The in vitro development of immature dendritic cells and their maturation capacity were greatly impaired, with decreased CD1a expression and IL-12p70 secretion ability. These cells were unable to induce autologous B cell proliferation and Ig secretion in the presence of CD40L and C3. Finally, the regulatory T cell development ability of CD4(+) T cells after CD3 and CD46 activation in the presence of IL-2 was significantly impaired. Thus, the association of important functional defects of dendritic cells, acquisition of B cell memory, and regulatory T cells with human C3 deficiency strongly supports a major role for C3 in bridging innate and adaptive immunity in humans.