Shiga toxin targets the podocyte causing hemolytic uremic syndrome through endothelial complement activation.

BACKGROUND: Shiga toxin (Stx)-producing Escherichia coli hemolytic uremic syndrome (STEC-HUS) is the leading cause of acute kidney injury in children, with an associated mortality of up to 5%. The mechanisms underlying STEC-HUS and why the glomerular microvasculature is so susceptible to injury following systemic Stx infection are unclear. METHODS: Transgenic mice were engineered to express the Stx receptor (Gb3) exclusively in their kidney podocytes (Pod-Gb3) and challenged with systemic Stx. Human glomerular cell models and kidney biopsies from patients with STEC-HUS were also studied. FINDINGS: Stx-challenged Pod-Gb3 mice developed STEC-HUS. This was mediated by a reduction in podocyte vascular endothelial growth factor A (VEGF-A), which led to loss of glomerular endothelial cell (GEnC) glycocalyx, a reduction in GEnC inhibitory complement factor H binding, and local activation of the complement pathway. Early therapeutic inhibition of the terminal complement pathway with a C5 inhibitor rescued this podocyte-driven, Stx-induced HUS phenotype. CONCLUSIONS: This study potentially explains why systemic Stx exposure targets the glomerulus and supports the early use of terminal complement pathway inhibition in this devastating disease. FUNDING: This work was supported by the UK Medical Research Council (MRC) (grant nos. G0901987 and MR/K010492/1) and Kidney Research UK (grant nos. TF_007_20151127, RP42/2012, and SP/FSGS1/2013). The Mary Lyon Center is part of the MRC Harwell Institute and is funded by the MRC (A410).

Mechanism and intervention of murine transfusion-related acute lung injury caused by anti-CD36 antibodies.

Anti-CD36 Abs have been suggested to induce transfusion-related acute lung injury (TRALI) upon blood transfusion, particularly in Asian populations. However, little is known about the pathological mechanism of anti-CD36 Ab-mediated TRALI, and potential therapies have not yet been identified. Here, we developed a murine model of anti-CD36 Ab-mediated TRALI to address these questions. Administration of mouse mAb against CD36 (mAb GZ1) or human anti-CD36 IgG, but not GZ1 F(ab')2 fragments, induced severe TRALI in Cd36+/+ male mice. Predepletion of recipient monocytes or complement, but not neutrophils or platelets, prevented the development of murine TRALI. Moreover, plasma C5a levels after TRALI induction by anti-CD36 Abs increased more than 3-fold, implying a critical role of complement C5 activation in the mechanism of Fc-dependent anti-CD36-mediated TRALI. Administration of GZ1 F(ab')2, antioxidant (N-acetyl cysteine, NAC), or C5 blocker (mAb BB5.1) before TRALI induction completely protected mice from anti-CD36-mediated TRALI. Although no significant amelioration in TRALI was observed when mice were injected with GZ1 F(ab')2 after TRALI induction, significant improvement was achieved when mice were treated postinduction with NAC or anti-C5. Importantly, anti-C5 treatment completely rescued mice from TRALI, suggesting the potential role of existing anti-C5 drugs in the treatment of patients with TRALI caused by anti-CD36.

Discovery of functionally distinct anti-C7 monoclonal antibodies and stratification of anti-nicotinic AChR positive Myasthenia Gravis patients.

Myasthenia Gravis (MG) is mediated by autoantibodies against acetylcholine receptors that cause loss of the receptors in the neuromuscular junction. Eculizumab, a C5-inhibitor, is the only approved treatment for MG that mechanistically addresses complement-mediated loss of nicotinic acetylcholine receptors. It is an expensive drug and was approved despite missing the primary efficacy endpoint in the Phase 3 REGAIN study. There are two observations to highlight. Firstly, further C5 inhibitors are in clinical development, but other terminal pathway proteins, such as C7, have been relatively understudied as therapeutic targets, despite the potential for lower and less frequent dosing. Secondly, given the known heterogenous mechanisms of action of autoantibodies in MG, effective patient stratification in the REGAIN trial may have provided more favorable efficacy readouts. We investigated C7 as a target and assessed the in vitro function, binding epitopes and mechanism of action of three mAbs against C7. We found the mAbs were human, cynomolgus monkey and/or rat cross-reactive and each had a distinct, novel mechanism of C7 inhibition. TPP1820 was effective in preventing experimental MG in rats in both prophylactic and therapeutic dosing regimens. To enable identification of MG patients that are likely to respond to C7 inhibition, we developed a patient stratification assay and showed in a small cohort of MG patients (n=19) that 63% had significant complement activation and C7-dependent loss of AChRs in this in vitro set up. This study provides validation of C7 as a target for treatment of MG and provides a means of identifying patients likely to respond to anti-C7 therapy based on complement-activating properties of patient autoantibodies.

C5b-9 Membrane Attack Complex Formation and Extracellular Vesicle Shedding in Barrett's Esophagus and Esophageal Adenocarcinoma.

The early complement components have emerged as mediators of pro-oncogenic inflammation, classically inferred to cause terminal complement activation, but there are limited data on the activity of terminal complement in cancer. We previously reported elevated serum and tissue C9, the terminal complement component, in esophageal adenocarcinoma (EAC) compared to the precursor condition Barrett's Esophagus (BE) and healthy controls. Here, we investigate the level and cellular fates of the terminal complement complex C5b-9, also known as the membrane attack complex. Punctate C5b-9 staining and diffuse C9 staining was detected in BE and EAC by multiplex immunohistofluorescence without corresponding increase of C9 mRNA transcript. Increased C9 and C5b-9 staining were observed in the sequence normal squamous epithelium, BE, low- and high-grade dysplasia, EAC. C5b-9 positive esophageal cells were morphologically intact, indicative of sublytic or complement-evasion mechanisms. To investigate this at a cellular level, we exposed non-dysplastic BE (BAR-T and CP-A), high-grade dysplastic BE (CP-B and CP-D) and EAC (FLO-1 and OE-33) cell lines to the same sublytic dose of immunopurified human C9 (3 µg/ml) in the presence of C9-depleted human serum. Cellular C5b-9 was visualized by immunofluorescence confocal microscopy. Shed C5b-9 in the form of extracellular vesicles (EV) was measured in collected conditioned medium using recently described microfluidic immunoassay with capture by a mixture of three tetraspanin antibodies (CD9/CD63/CD81) and detection by surface-enhanced Raman scattering (SERS) after EV labelling with C5b-9 or C9 antibody conjugated SERS nanotags. Following C9 exposure, all examined cell lines formed C5b-9, internalized C5b-9, and shed C5b-9+ and C9+ EVs, albeit at varying levels despite receiving the same C9 dose. In conclusion, these results confirm increased esophageal C5b-9 formation during EAC development and demonstrate capability and heterogeneity in C5b-9 formation and shedding in BE and EAC cell lines following sublytic C9 exposure. Future work may explore the molecular mechanisms and pathogenic implications of the shed C5b-9+ EV.

Whole blood-based measurement of SARS-CoV-2-specific T cells reveals asymptomatic infection and vaccine immunogenicity in healthy subjects and patients with solid-organ cancers.

Accurate assessment of SARS-CoV-2 immunity is critical in evaluating vaccine efficacy and devising public health policies. Whilst the exact nature of effective immunity remains incompletely defined, SARS-CoV-2-specific T-cell responses are a critical feature that will likely form a key correlate of protection against COVID-19. Here, we developed and optimized a high-throughput whole blood-based assay to determine the T-cell response associated with prior SARS-CoV-2 infection and/or vaccination amongst 231 healthy donors and 68 cancer patients. Following overnight in vitro stimulation with SARS-CoV-2-specific peptides, blood plasma samples were analysed for TH 1-type cytokines. Highly significant differential IFN-γ+ /IL-2+ SARS-CoV-2-specific T-cell responses were seen amongst previously infected COVID-19-positive healthy donors in comparison with unknown / naïve individuals (p < 0·0001). IFN-γ production was more effective at identifying asymptomatic donors, demonstrating higher sensitivity (96·0% vs. 83·3%) but lower specificity (84·4% vs. 92·5%) than measurement of IL-2. A single COVID-19 vaccine dose induced IFN-γ and/or IL-2 SARS-CoV-2-specific T-cell responses in 116 of 128 (90·6%) healthy donors, reducing significantly to 27 of 56 (48·2%) when measured in cancer patients (p < 0·0001). A second dose was sufficient to boost T-cell responses in the majority (90·6%) of cancer patients, albeit IFN-γ+ responses were still significantly lower overall than those induced in healthy donors (p = 0·034). Three-month post-vaccination T-cell responses also declined at a faster rate in cancer patients. Overall, this cost-effective standardizable test ensures accurate and comparable assessments of SARS-CoV-2-specific T-cell responses amenable to widespread population immunity testing, and identifies individuals at greater need of booster vaccinations.

Genetic Insights into the Impact of Complement in Alzheimer's Disease.

The presence of complement activation products at sites of pathology in post-mortem Alzheimer's disease (AD) brains is well known. Recent evidence from genome-wide association studies (GWAS), combined with the demonstration that complement activation is pivotal in synapse loss in AD, strongly implicates complement in disease aetiology. Genetic variations in complement genes are widespread. While most variants individually have only minor effects on complement homeostasis, the combined effects of variants in multiple complement genes, referred to as the "complotype", can have major effects. In some diseases, the complotype highlights specific parts of the complement pathway involved in disease, thereby pointing towards a mechanism; however, this is not the case with AD. Here we review the complement GWAS hits; CR1 encoding complement receptor 1 (CR1), CLU encoding clusterin, and a suggestive association of C1S encoding the enzyme C1s, and discuss difficulties in attributing the AD association in these genes to complement function. A better understanding of complement genetics in AD might facilitate predictive genetic screening tests and enable the development of simple diagnostic tools and guide the future use of anti-complement drugs, of which several are currently in development for central nervous system disorders.

C3 Glomerulopathy and Related Disorders in Children: Etiology-Phenotype Correlation and Outcomes.

BACKGROUND AND OBJECTIVES: Membranoproliferative GN and C3 glomerulopathy are rare and overlapping disorders associated with dysregulation of the alternative complement pathway. Specific etiologic data for pediatric membranoproliferative GN/C3 glomerulopathy are lacking, and outcome data are based on retrospective studies without etiologic data. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: A total of 80 prevalent pediatric patients with membranoproliferative GN/C3 glomerulopathy underwent detailed phenotyping and long-term follow-up within the National Registry of Rare Kidney Diseases (RaDaR). Risk factors for kidney survival were determined using a Cox proportional hazards model. Kidney and transplant graft survival was determined using the Kaplan-Meier method. RESULTS: Central histology review determined 39 patients with C3 glomerulopathy, 31 with immune-complex membranoproliferative GN, and ten with immune-complex GN. Patients were aged 2-15 (median, 9; interquartile range, 7-11) years. Median complement C3 and C4 levels were 0.31 g/L and 0.14 g/L, respectively; acquired (anticomplement autoantibodies) or genetic alternative pathway abnormalities were detected in 46% and 9% of patients, respectively, across all groups, including those with immune-complex GN. Median follow-up was 5.18 (interquartile range, 2.13-8.08) years. Eleven patients (14%) progressed to kidney failure, with nine transplants performed in eight patients, two of which failed due to recurrent disease. Presence of >50% crescents on the initial biopsy specimen was the sole variable associated with kidney failure in multivariable analysis (hazard ratio, 6.2; 95% confidence interval, 1.05 to 36.6; P<0.05). Three distinct C3 glomerulopathy prognostic groups were identified according to presenting eGFR and >50% crescents on the initial biopsy specimen. CONCLUSIONS: Crescentic disease was a key risk factor associated with kidney failure in a national cohort of pediatric patients with membranoproliferative GN/C3 glomerulopathy and immune-complex GN. Presenting eGFR and crescentic disease help define prognostic groups in pediatric C3 glomerulopathy. Acquired abnormalities of the alternative pathway were commonly identified but not a risk factor for kidney failure.

Development of EndoScreen Chip, a Microfluidic Pre-Endoscopy Triage Test for Esophageal Adenocarcinoma.

The current endoscopy and biopsy diagnosis of esophageal adenocarcinoma (EAC) and its premalignant condition Barrett's esophagus (BE) is not cost-effective. To enable EAC screening and patient triaging for endoscopy, we developed a microfluidic lectin immunoassay, the EndoScreen Chip, which allows sensitive multiplex serum biomarker measurements. Here, we report the proof-of-concept deployment for the EAC biomarker Jacalin lectin binding complement C9 (JAC-C9), which we previously discovered and validated by mass spectrometry. A monoclonal C9 antibody (m26 3C9) was generated and validated in microplate ELISA, and then deployed for JAC-C9 measurement on EndoScreen Chip. Cohort evaluation (n = 46) confirmed the expected elevation of serum JAC-C9 in EAC, along with elevated total serum C9 level. Next, we asked if the small panel of serum biomarkers improves detection of EAC in this cohort when used in conjunction with patient risk factors (age, body mass index and heartburn history). Using logistic regression modeling, we found that serum C9 and JAC-C9 significantly improved EAC prediction from AUROC of 0.838 to 0.931, with JAC-C9 strongly predictive of EAC (vs. BE OR = 4.6, 95% CI: 1.6-15.6, p = 0.014; vs. Healthy OR = 4.1, 95% CI: 1.2-13.7, p = 0.024). This proof-of-concept study confirms the microfluidic EndoScreen Chip technology and supports the potential utility of blood biomarkers in improving triaging for diagnostic endoscopy. Future work will expand the number of markers on EndoScreen Chip from our list of validated EAC biomarkers.

Role of complement and potential of complement inhibitors in myasthenia gravis and neuromyelitis optica spectrum disorders: a brief review.

The complement system is a powerful member of the innate immune system. It is highly adept at protecting against pathogens, but exists in a delicate balance between its protective functions and overactivity, which can result in autoimmune disease. A cascade of complement proteins that requires sequential activation, and numerous complement regulators, exists to regulate a proportionate response to pathogens. In spite of these mechanisms there is significant evidence for involvement of the complement system in driving the pathogenesis of variety of diseases including neuromyelitis optica spectrum disorders (NMOSD) and myasthenia gravis (MG). As an amplification cascade, there are an abundance of molecular targets that could be utilized for therapeutic intervention. Clinical trials assessing complement pathway inhibition in both these conditions have recently been completed and include the first randomized placebo-controlled trial in NMOSD showing positive results. This review aims to review and update the reader on the complement system and the evolution of complement-based therapeutics in these two disorders.

Treatment of COVID-19 with remdesivir in the absence of humoral immunity: a case report.

The response to the coronavirus disease 2019 (COVID-19) pandemic has been hampered by lack of an effective severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antiviral therapy. Here we report the use of remdesivir in a patient with COVID-19 and the prototypic genetic antibody deficiency X-linked agammaglobulinaemia (XLA). Despite evidence of complement activation and a robust T cell response, the patient developed persistent SARS-CoV-2 pneumonitis, without progressing to multi-organ involvement. This unusual clinical course is consistent with a contribution of antibodies to both viral clearance and progression to severe disease. In the absence of these confounders, we take an experimental medicine approach to examine the in vivo utility of remdesivir. Over two independent courses of treatment, we observe a temporally correlated clinical and virological response, leading to clinical resolution and viral clearance, with no evidence of acquired drug resistance. We therefore provide evidence for the antiviral efficacy of remdesivir in vivo, and its potential benefit in selected patients.

A novel mouse model expressing human forms for complement receptors CR1 and CR2.

BACKGROUND: The complement cascade is increasingly implicated in development of a variety of diseases with strong immune contributions such as Alzheimer's disease and Systemic Lupus Erythematosus. Mouse models have been used to determine function of central components of the complement cascade such as C1q and C3. However, species differences in their gene structures mean that mice do not adequately replicate human complement regulators, including CR1 and CR2. Genetic variation in CR1 and CR2 have been implicated in modifying disease states but the mechanisms are not known. RESULTS: To decipher the roles of human CR1 and CR2 in health and disease, we engineered C57BL/6J (B6) mice to replace endogenous murine Cr2 with human complement receptors, CR1 and CR2 (B6.CR2CR1). CR1 has an array of allotypes in human populations and using traditional recombination methods (Flp-frt and Cre-loxP) two of the most common alleles (referred to here as CR1long and CR1short) can be replicated within this mouse model, along with a CR1 knockout allele (CR1KO). Transcriptional profiling of spleens and brains identified genes and pathways differentially expressed between mice homozygous for either CR1long, CR1short or CR1KO. Gene set enrichment analysis predicts hematopoietic cell number and cell infiltration are modulated by CR1long, but not CR1short or CR1KO. CONCLUSION: The B6.CR2CR1 mouse model provides a novel tool for determining the relationship between human-relevant CR1 alleles and disease.

Monoclonal Antibodies Capable of Inhibiting Complement Downstream of C5 in Multiple Species.

Better understanding of roles of complement in pathology has fuelled an explosion of interest in complement-targeted therapeutics. The C5-blocking monoclonal antibody (mAb) eculizumab, the first of the new wave of complement blocking drugs, was FDA approved for treatment of Paroxysmal Nocturnal Hemoglobinuria in 2007; its expansion into other diseases has been slow and remains restricted to rare and ultra-rare diseases such as atypical hemolytic uremic syndrome. The success of eculizumab has provoked other Pharma to follow this well-trodden track and made C5 blockade the busiest area of complement drug development. C5 blockade inhibits generation of C5a and C5b, the former an anaphylatoxin, the latter the nidus for formation of the pro-inflammatory membrane attack complex. In order to use anti-complement drugs in common complement-driven diseases, more affordable and equally effective therapeutics are needed. To address this, we explored complement inhibition downstream of C5. Novel blocking mAbs targeting C7 and/or the C5b-7 complex were generated, identified using high throughput functional assays and specificity confirmed by immunochemical assays and surface plasmon resonance (SPR). Selected mAbs were tested in rodents to characterize pharmacokinetics, and therapeutic capacity. Administration of a mouse C7-selective mAb to wildtype mice, or a human C7 specific mAb to C7-deficient mice reconstituted with human C7, completely inhibited serum lytic activity for >48 h. The C5b-7 complex selective mAb 2H2, most active in rat serum, efficiently inhibited serum lytic activity in vivo for over a week from a single low dose (10 mg/kg); this mAb effectively blocked disease and protected muscle endplates from destruction in a rat myasthenia model. Targeting C7 and C7-containing terminal pathway intermediates is an innovative therapeutic approach, allowing lower drug dose and lower product cost, that will facilitate the expansion of complement therapeutics to common diseases.

Development and characterization of novel anti-C5 monoclonal antibodies capable of inhibiting complement in multiple species.

Over the last decade there has been an explosion in complement therapies; one-third of the drugs in the clinic or in development target C5 protein. Eculizumab, a monoclonal antibody (mAb) that binds C5 and blocks its cleavage by the convertase, is the current reference standard treatment for atypical haemolytic uraemic syndrome (aHUS) and paroxysmal nocturnal haemoglobinuria (PNH) and in clinical trials for many other diseases. Here we describe a panel of novel anti-C5 mAb, including mAb that, like Eculizumab, are efficient inhibitors of complement but, unlike Eculizumab, inhibit across species, including human, rat, rabbit and guinea pig. Several inhibitory anti-C5 mAb were identified and characterized for C5 binding and lytic inhibitory capacity in comparison to current therapeutic anti-C5 mAb; three clones, 4G2, 7D4 and 10B6, were selected and further characterized for ligand specificity and affinity and cross-species inhibitory activity. The mAb 10B6 was human-specific whereas mAb 4G2 and 7D4 efficiently inhibited lysis by human, rabbit and rat serum, and weakly inhibited guinea pig complement; 7D4 also weakly inhibited mouse complement in vitro The rat C5-cross-reactive mAb 4G2, when administered intraperitoneally in a rat model of myasthenia gravis, effectively blocked the disease and protected muscle endplates from destruction. To our knowledge this is the first report of an anti-C5 function blocking mAb that permits preclinical studies in rats.

Complement system biomarkers in first episode psychosis.

Several lines of evidence implicate immunological/inflammatory factors in development of schizophrenia. Complement is a key driver of inflammation, and complement dysregulation causes pathology in many diseases. Here we explored whether complement dysregulation occurred in first episode psychosis (FEP) and whether this provides a source of biomarkers. Eleven complement analytes (C1q, C3, C4, C5, factor B [FB], terminal complement complex [TCC], factor H [FH], FH-related proteins [FHR125], Properdin, C1 inhibitor [C1inh], soluble complement receptor 1 [CR1]) plus C-reactive protein (CRP) were measured in serum from 136 first episode psychosis (FEP) cases and 42 mentally healthy controls using established in-house or commercial ELISA. The relationship between caseness and variables (analytes measured, sex, age, ethnicity, tobacco/cannabis smoking) was tested by multivariate logistic regression. When measured individually, only TCC was significantly different between FEP and controls (p=0.01). Stepwise selection demonstrated interdependence between some variables and revealed other variables that significantly and independently contributed to distinguishing cases and controls. The final model included demographics (sex, ethnicity, age, tobacco smoking) and a subset of analytes (C3, C4, C5, TCC, C1inh, FHR125, CR1). A receiver operating curve analysis combining these variables yielded an area under the curve of 0.79 for differentiating FEP from controls. This model was confirmed by multiple replications using randomly selected sample subsets. The data suggest that complement dysregulation occurs in FEP, supporting an underlying immune/inflammatory component to the disorder. Classification of FEP cases according to biological variables rather than symptoms would help stratify cases to identify those that might most benefit from therapeutic modification of the inflammatory response.

Hexamerization-enhanced CD20 antibody mediates complement-dependent cytotoxicity in serum genetically deficient in C9.

We examined complement-dependent cytotoxicity (CDC) by hexamer formation-enhanced CD20 mAb Hx-7D8 of patient-derived chronic lymphocytic leukemia (CLL) cells that are relatively resistant to CDC. CDC was analyzed in normal human serum (NHS) and serum from an individual genetically deficient for C9. Hx-7D8 was able to kill up to 80% of CLL cells in complete absence of C9. We conclude that the narrow C5b-8 pores formed without C9 are sufficient for CDC due to efficient antibody-mediated hexamer formation. In the absence of C9, we observed transient intracellular increases of Ca2+ during CDC (as assessed with FLUO-4) that were extended in time. This suggests that small C5b-8 pores allow Ca2+ to enter the cell, while dissipation of the fluorescent signal accompanying cell disintegration is delayed. The Ca2+ signal is retained concomitantly with TOPRO-3 (viability dye) staining, thereby confirming that Ca2+ influx represents the most proximate mediator of cell death by CDC.

The Correlation between Inflammatory Biomarkers and Polygenic Risk Score in Alzheimer's Disease.

Plasma biomarkers to aid the early diagnosis of Alzheimer's disease (AD) or to monitor disease progression have long been sought and continue to be widely studied. Biomarkers that correlate with AD polygenic risk score, a measure of the polygenic architecture of the disease and highly predictive of AD status, would be excellent candidates. Therefore, we undertook a preliminary study to assess the association of plasma inflammatory biomarkers with an overall AD polygenic risk score as well as with an inflammation-specific AD polygenic risk score in a sample set of 93 AD cases. We measured five complement biomarkers [complement receptor 1 (CR1), clusterin, complement component 9 (C9), C1 inhibitor (C1inh), terminal complement complex (TCC)] and the benchmark inflammatory marker C-reactive protein (CRP). Plasma clusterin level showed an association with overall AD polygenic risk score, while clusterin, C1inh, and CRP levels each displayed some association with the inflammatory-specific AD polygenic risk score. The results suggest that elevated plasma levels of inflammatory biomarkers, including complement proteins, associate with polygenic risk scores in AD, further strengthening the link between genetic and biomarker disease predictors and indicating a potential role for these markers in disease prediction and patient stratification in AD.

Complement Membrane Attack and Tumorigenesis: A SYSTEMS BIOLOGY APPROACH.

Tumor development driven by inflammation is now an established phenomenon, but the role that complement plays remains uncertain. Recent evidence has suggested that various components of the complement (C) cascade may influence tumor development in disparate ways; however, little attention has been paid to that of the membrane attack complex (MAC). This is despite abundant evidence documenting the effects of this complex on cell behavior, including cell activation, protection from/induction of apoptosis, release of inflammatory cytokines, growth factors, and ECM components and regulators, and the triggering of the NLRP3 inflammasome. Here we present a novel approach to this issue by using global gene expression studies in conjunction with a systems biology analysis. Using network analysis of MAC-responsive expression changes, we demonstrate a cluster of co-regulated genes known to have impact in the extracellular space and on the supporting stroma and with well characterized tumor-promoting roles. Network analysis highlighted the central role for EGF receptor activation in mediating the observed responses to MAC exposure. Overall, the study sheds light on the mechanisms by which sublytic MAC causes tumor cell responses and exposes a gene expression signature that implicates MAC as a driver of tumor progression. These findings have implications for understanding of the roles of complement and the MAC in tumor development and progression, which in turn will inform future therapeutic strategies in cancer.

Deletion of the membrane complement inhibitor CD59a drives age and gender-dependent alterations to bone phenotype in mice.

Degenerative joint diseases such as osteoarthritis are characterised by aberrant region-specific bone formation and abnormal bone mineral content. A recent study suggested a role for the complement membrane attack complex in experimental models of osteoarthritis. Since CD59a is the principal regulator of the membrane attack complex in mice, we evaluated the impact of CD59a gene deletion upon maintenance of bone architecture. In vivo bone morphology analysis revealed that male CD59a-deficient mice have increased femur length and cortical bone volume, albeit with reduced bone mineral density. However, this phenomenon was not observed in female mice. Histomorphometric analysis of the trabecular bone showed increased rates of bone homeostasis, with both increased bone resorption and mineral apposition rate in CD59a-deficient male mice. When bone cells were studied in isolation, in vitro osteoclastogenesis was significantly increased in male CD59a-deficient mice, although osteoblast formation was not altered. Our data reveal, for the first time, that CD59a is a regulator of bone growth and homeostasis. CD59a ablation in male mice results in longer and wider bones, but with less density, which is likely a major contributing factor for their susceptibility to osteoarthritis. These findings increase our understanding of the role of complement regulation in degenerative arthritis.

Complement activation in leprosy: a retrospective study shows elevated circulating terminal complement complex in reactional leprosy.

Mycobacterium leprae infection gives rise to the immunologically and histopathologically classified spectrum of leprosy. At present, several tools for the stratification of patients are based on acquired immunity markers. However, the role of innate immunity, particularly the complement system, is largely unexplored. The present retrospective study was undertaken to explore whether the systemic levels of complement activation components and regulators can stratify leprosy patients, particularly in reference to the reactional state of the disease. Serum samples from two cohorts were analysed. The cohort from Bangladesh included multi-bacillary (MB) patients with (n = 12) or without (n = 46) reaction (R) at intake and endemic controls (n = 20). The cohort from Ethiopia included pauci-bacillary (PB) (n = 7) and MB (n = 23) patients without reaction and MB (n = 15) patients with reaction. The results showed that the activation products terminal complement complex (TCC) (P ≤ 0·01), C4d (P ≤ 0·05) and iC3b (P ≤ 0·05) were specifically elevated in Bangladeshi patients with reaction at intake compared to endemic controls. In addition, levels of the regulator clusterin (P ≤ 0·001 without R; P < 0·05 with R) were also elevated in MB patients, irrespective of a reaction. Similar analysis of the Ethiopian cohort confirmed that, irrespective of a reaction, serum TCC levels were increased significantly in patients with reactions compared to patients without reactions (P ≤ 0·05). Our findings suggests that serum TCC levels may prove to be a valuable tool in diagnosing patients at risk of developing reactions.

Complement, a target for therapy in inflammatory and degenerative diseases.

The complement system is a key innate immune defence against infection and an important driver of inflammation; however, these very properties can also cause harm. Inappropriate or uncontrolled activation of complement can cause local and/or systemic inflammation, tissue damage and disease. Complement provides numerous options for drug development as it is a proteolytic cascade that involves nine specific proteases, unique multimolecular activation and lytic complexes, an arsenal of natural inhibitors, and numerous receptors that bind to activation fragments. Drug design is facilitated by the increasingly detailed structural understanding of the molecules involved in the complement system. Only two anti-complement drugs are currently on the market, but many more are being developed for diseases that include infectious, inflammatory, degenerative, traumatic and neoplastic disorders. In this Review, we describe the history, current landscape and future directions for anti-complement therapies.