Targeting of oxidized Macrophage Migration Inhibitory Factor (oxMIF) with antibody ON104 attenuates the severity of glomerulonephritis.

The oxidized form of Macrophage Migration Inhibitory Factor (oxMIF) has been identified as the disease-related isoform of MIF, exerting pathological functions in inflamed tissue. In this study, we aimed to explore the in vivo effects of the neutralizing anti-oxMIF antibody ON104 in a rat model of crescentic glomerulonephritis (CGN), to better understand its disease modifying activities. WKY rats received a single intravenous injection of a rabbit nephrotoxic serum (NTS), targeting rat glomerular basement membrane to induce CGN. On day 4 and day 6, ON104 was given intraperitoneally (i.p.) and on day 8 urine, blood and kidney tissue were collected. ON104 substantially attenuated the severity of CGN demonstrated by reduced proteinuria, hematuria, as well as lower levels of kidney injury molecule (KIM)-1. ON104 treatment preserved the glomerular morphology and suppressed crescent formation, a hallmark of the disease. On the cellular level, oxMIF neutralization by ON104 strongly reduced the number of macrophages and neutrophils within the inflamed kidneys. In vitro, we identified human neutrophils, but not monocytes, as main producers of oxMIF among total peripheral cells. The present study demonstrates that oxMIF is a pertinent therapeutic target in a model of CGN which mechanistically resembles human immune mediated CGN. In this model, neutralization of oxMIF by ON104 leads to an improvement in both urinary abnormalities and histological pathological characteristics of the disease. ON104, thus has the potential to become a novel disease-modifying drug for the treatment of glomerulonephritis and other inflammatory kidney diseases.

Description and Cross-Sectional Analyses of 25,880 Adults and Children in the UK National Registry of Rare Kidney Diseases Cohort.

Introduction: The National Registry of Rare Kidney Diseases (RaDaR) collects data from people living with rare kidney diseases across the UK, and is the world's largest, rare kidney disease registry. We present the clinical demographics and renal function of 25,880 prevalent patients and sought evidence of bias in recruitment to RaDaR. Methods: RaDaR is linked with the UK Renal Registry (UKRR, with which all UK patients receiving kidney replacement therapy [KRT] are registered). We assessed ethnicity and socioeconomic status in the following: (i) prevalent RaDaR patients receiving KRT compared with patients with eligible rare disease diagnoses receiving KRT in the UKRR, (ii) patients recruited to RaDaR compared with all eligible unrecruited patients at 2 renal centers, and (iii) the age-stratified ethnicity distribution of RaDaR patients with autosomal dominant polycystic kidney disease (ADPKD) was compared to that of the English census. Results: We found evidence of disparities in ethnicity and social deprivation in recruitment to RaDaR; however, these were not consistent across comparisons. Compared with either adults recruited to RaDaR or the English population, children recruited to RaDaR were more likely to be of Asian ethnicity (17.3% vs. 7.5%, P-value < 0.0001) and live in more socially deprived areas (30.3% vs. 17.3% in the most deprived Index of Multiple Deprivation (IMD) quintile, P-value < 0.0001). Conclusion: We observed no evidence of systematic biases in recruitment of patients into RaDaR; however, the data provide empirical evidence of negative economic and social consequences (across all ethnicities) experienced by families with children affected by rare kidney diseases.

Effects of rare kidney diseases on kidney failure: a longitudinal analysis of the UK National Registry of Rare Kidney Diseases (RaDaR) cohort.

BACKGROUND: Individuals with rare kidney diseases account for 5-10% of people with chronic kidney disease, but constitute more than 25% of patients receiving kidney replacement therapy. The National Registry of Rare Kidney Diseases (RaDaR) gathers longitudinal data from patients with these conditions, which we used to study disease progression and outcomes of death and kidney failure. METHODS: People aged 0-96 years living with 28 types of rare kidney diseases were recruited from 108 UK renal care facilities. The primary outcomes were cumulative incidence of mortality and kidney failure in individuals with rare kidney diseases, which were calculated and compared with that of unselected patients with chronic kidney disease. Cumulative incidence and Kaplan-Meier survival estimates were calculated for the following outcomes: median age at kidney failure; median age at death; time from start of dialysis to death; and time from diagnosis to estimated glomerular filtration rate (eGFR) thresholds, allowing calculation of time from last eGFR of 75 mL/min per 1·73 m2 or more to first eGFR of less than 30 mL/min per 1·73 m2 (the therapeutic trial window). FINDINGS: Between Jan 18, 2010, and July 25, 2022, 27 285 participants were recruited to RaDaR. Median follow-up time from diagnosis was 9·6 years (IQR 5·9-16·7). RaDaR participants had significantly higher 5-year cumulative incidence of kidney failure than 2·81 million UK patients with all-cause chronic kidney disease (28% vs 1%; p<0·0001), but better survival rates (standardised mortality ratio 0·42 [95% CI 0·32-0·52]; p<0·0001). Median age at kidney failure, median age at death, time from start of dialysis to death, time from diagnosis to eGFR thresholds, and therapeutic trial window all varied substantially between rare diseases. INTERPRETATION: Patients with rare kidney diseases differ from the general population of individuals with chronic kidney disease: they have higher 5-year rates of kidney failure but higher survival than other patients with chronic kidney disease stages 3-5, and so are over-represented in the cohort of patients requiring kidney replacement therapy. Addressing unmet therapeutic need for patients with rare kidney diseases could have a large beneficial effect on long-term kidney replacement therapy demand. FUNDING: RaDaR is funded by the Medical Research Council, Kidney Research UK, Kidney Care UK, and the Polycystic Kidney Disease Charity.

Randomized Trial on the Effect of an Oral Spleen Tyrosine Kinase Inhibitor in the Treatment of IgA Nephropathy.

Introduction: We reported increased spleen tyrosine kinase (SYK) expression in kidney biopsies of patients with IgA nephropathy (IgAN) and that inhibition of SYK reduces inflammatory cytokines production from IgA stimulated mesangial cells. Methods: This study was a double-blind, randomized, placebo-controlled phase 2 trial of fostamatinib (an oral SYK inhibitor) in 76 patients with IgAN. Patients were randomized to receive placebo, fostamatinib at 100 mg or 150 mg twice daily for 24 weeks on top of maximum tolerated dose of renin-angiotensin system inhibitors. The primary end point was reduction of proteinuria. Secondary end points included change from baseline in estimated glomerular filtration rate (eGFR) and kidney histology. Results: Although we could not detect significant reduction in proteinuria with fostamatinib overall, in a predetermined subgroup analysis, there was a trend for dose-dependent reduction in median proteinuria (from baseline to 24 weeks by 14%, 27%, and 36% in the placebo, fostamatinib 100 mg, and 150 mg groups, respectively) in patients with baseline urinary protein-to-creatinine ratios (UPCR) more than 1000 mg/g. Kidney function (eGFR) remained stable in all groups. Fostamatinib was well-tolerated. Side effects included diarrhea, hypertension, and increased liver enzymes. Thirty-nine patients underwent repeat biopsy showing reductions in SYK staining associated with therapy at low dose (-1.5 vs. 1.7 SYK+ cells/glomerulus in the placebo group, P < 0.05). Conclusions: There was a trend toward reduction in proteinuria with fostamatinib in a predefined analysis of high risk patients with IgAN despite maximal care, as defined by baseline UPCR greater than 1000 mg/g. Further study may be warranted.

Association and progression of multi-morbidity with Chronic Kidney Disease stage 3a secondary to Type 2 Diabetes Mellitus, grouped by albuminuria status in the multi-ethnic population of Northwest London: A real-world study.

INTRODUCTION: The prevalence of Diabetic Kidney Disease (DKD) secondary to Type 2 Diabetes Mellitus (T2DM) is rising worldwide. However, real-world data linking glomerular function and albuminuria to the degree of multi-morbidity is lacking. We thus utilised the Discover dataset, to determine this association. METHOD: Patients with T2DM diagnosed prior to 1st January 2015 with no available biochemical evidence of CKD were included. Patients subsequently diagnosed and coded for CKD3a in 2015, were grouped by the degree of albuminuria. Baseline and 5-year co-morbidity was determined, as were prescribing practices with regards to prognostically beneficial medication. RESULTS: We identified 56,261 patients with T2DM, of which 1082 had CKD stage 3a diagnosed in 2015 (224-CKD3aA1,154-CKD3aA2,93-CKD3aA1; 611 patients with CKD3a but no uACR available in 2015 were excluded from follow up). No statistically significant difference was observed in the degree of co-morbidities at baseline. A significant difference in the degree of hypertension, retinopathy, ischaemic heart disease and vascular disease from baseline compared to study end point was observed for all 3 study groups. Comparing co-morbidities developed at study end point, highlighted a statistical difference between CKD3aA1 Vs CKD3aA3 for retinopathy alone and for hypertension and heart failure between CKD3aA2 Vs CKD3aA3. 40.8% of patients with CKD3aA2 or A3 were prescribed Renin Angiotensin Aldosterone inhibitors (RAASi) therapy between June-December 2021. Survival analysis showed 15% of patients with CKD3aA3 developed CKD stage 5 within 5 years of diagnosis. DISCUSSION: CKD3a secondary to DKD is associated with significant multimorbidity at baseline and 5 years post diagnosis, with CKD3aA3 most strongly associated with CKD progression to CKD 5, heart failure, hypertension and retinopathy compared to CKD3aA1 or CKD3aA2 at 5 years post diagnosis. The lack of uACR testing upon diagnosis and poor prescribing of RAASi, in those with CKD3aA2/A3, raises significant cause for concern. CONCLUSION: DKD is associated with significant multimorbidity. Significant work is needed to be done to ensure patients undergo testing for uACR, to allow for future risk stratification and ability to be started on prognostically beneficial medication.

Non-crystalline light chain proximal tubulopathy, a morphologically protean entity.

BACKGROUND: Light chain proximal tubulopathy (LCPT) is a rare form of paraprotein-related disease, occurring in two main histopathological forms: crystalline and non-crystalline. The clinicopathological features, treatment strategies and outcomes, especially of the non-crystalline form, are not well described. METHODS: We conducted a single-centre retrospective case series of 12 LCPT patients, 5 crystalline and 7 non-crystalline, between 2005 and 2021. RESULTS: The median age was 69.5 years (range 47-80). Ten patients presented with CKD and significant proteinuria (median estimated glomerular filtration rate of 43.5 ml/min/1.73 m2; urine protein:creatinine ratio 328 mg/mmol). Only six patients had known haematological disease at the time of renal biopsy. Multiple myeloma (MM) was diagnosed in seven patients cases and monoclonal gammopathy of renal significance (MGRS) in five patients. A clone was detected in all cases combining serum/urine electrophoresis and free light chain (LC) assays. Crystalline and non-crystalline variants had similar clinical presentations. For the non-crystalline variant, a diagnosis was reached based on a combination of CKD without another cause, haematological workup, LC restriction on immunofluorescence and abnormalities on electron microscopy (EM). Nine of 12 patients received clone-directed treatment. Patients who achieved haematological response (including all non-crystalline LCPT) had improved renal outcomes over a median follow-up of 79 months. CONCLUSIONS: The non-crystalline variant may go unrecognised because of its subtle histopathological features and requires EM to distinguish it from 'excessive LC resorption without tubular injury'. Clone-directed treatment with good haematological response improves renal outcomes in both variants but limited data exist in MGRS. Multicentre prospective studies are needed to better define the clinicopathological characteristics associated with poor outcomes and optimize treatment strategies in patients with MGRS.

Syk Activation in Circulating and Tissue Innate Immune Cells in Antineutrophil Cytoplasmic Antibody-Associated Vasculitis.

OBJECTIVE: Syk is a cytoplasmic protein tyrosine kinase that plays a role in signaling via B cell and Fc receptors (FcR). FcR engagement and signaling via Syk is thought to be important in antineutrophil cytoplasm antibody (ANCA) IgG-mediated neutrophil activation. This study was undertaken to investigate the role of Syk in ANCA-induced myeloid cell activation and vasculitis pathogenesis. METHODS: Phosphorylation of Syk in myeloid cells from healthy controls and ANCA-associated vasculitis (AAV) patients was analyzed using flow cytometry. The effect of Syk inhibition on myeloperoxidase (MPO)-ANCA IgG activation of cells was investigated using functional assays (interleukin-8 and reactive oxygen species production) and targeted gene analysis with NanoString. Total and phosphorylated Syk at sites of tissue inflammation in patients with AAV was assessed using immunohistochemistry and RNAscope in situ hybridization. RESULTS: We identified increased phosphorylated Syk at critical activatory tyrosine residues in blood neutrophils and monocytes from patients with active AAV compared to patients with disease in remission or healthy controls. Syk was phosphorylated in vitro following MPO-ANCA IgG stimulation, and Syk inhibition was able to prevent ANCA-mediated cellular responses. Using targeted gene expression analysis, we identified up-regulation of FcR- and Syk-dependent signaling pathways following MPO-ANCA IgG stimulation. Finally, we showed that Syk is expressed and phosphorylated in tissue leukocytes at sites of organ inflammation in AAV. CONCLUSION: These findings indicate that Syk plays a critical role in MPO-ANCA IgG-induced myeloid cell responses and that Syk is activated in circulating immune cells and tissue immune cells in AAV; therefore, Syk inhibition may be a potential therapeutic option.

Predictors of iron versus erythropoietin responsiveness in anemic hemodialysis patients.

Anemia protocols for hemodialysis patients usually titrate erythropoietin (ESA) according to hemoglobin and iron according to a threshold of ferritin, with variable response seen. A universally optimum threshold for ferritin may be incorrect, and another view is that ESA and iron are alternative anemia treatments, which should be selected based on the likely response to each. Hemodialysis patients developing moderate anemia were randomised to treatment with either an increase in ESA or a course of intravenous iron. Over 2423 patient-months in 197 patients, there were 133 anemia episodes with randomized treatment. Treatment failure was seen in 20/66 patients treated with ESA and 20/67 patients treated with iron (30.3 vs. 29.9%, p = 1.0). Successful ESA treatment was associated with lower C-reactive protein (13.5 vs. 28.6 mg/L, p = 0.038) and lower previous ESA dose (6621 vs. 9273 µg/week, p = 0.097). Successful iron treatment was associated with lower reticulocyte hemoglobin (33.8 vs. 35.5 pg, p = 0.047), lower hepcidin (91.4 vs. 131.0 µg/ml, p = 0.021), and higher C-reactive protein (29.5 vs. 12.6 mg/L, p = 0.085). A four-variable iron preference score was developed to indicate the more favorable treatment, which in a retrospective analysis reduced treatment failure to 17%. Increased ESA and iron are equally effective, though treatment failure occurs in almost 30%. Baseline variables including hepcidin can predict treatment response, and a four-variable score shows promise in allowing directed treatment with improved response rates.

Impact of kidney size on the outcome of diabetic patients receiving hemodialysis.

INTRODUCTION: Diabetic patients normally have enlarged or normal-sized kidneys throughout their lifetime, but some diabetic uremic patients have small kidneys. It is uncertain if kidney size could have any negative impact on outcome in hemodialysis patients. METHODS: This longitudinal, observational cohort study recruited 301 diabetic hemodialysis patients in 2015, and followed until 2019. Patients were stratified into two subgroups according to their kidney sizes before dialysis, as small (n = 32) or enlarged or normal (n = 269). Baseline demographic, hematological, biochemical, nutritional, inflammatory and dialysis related data were collected for analysis. RESULTS: Patients with small kidney size were not only older (P<0.001) and had lower body mass index (P = 0.016), but had also higher blood uric acid concentration (P<0.001) compared with patients with enlarged or normal kidney size. All patients received adequate doses of hemodialysis since the Kt/V and urea reduction ratio was 1.7±0.3 and 0.7±0.1, respectively. Patients with small size kidneys received higher erythropoietin dose than patients with enlarged or normal kidney size (P = 0.031). At the end of analysis, 92 (30.6%) patients expired. Kaplan-Meier analysis revealed no survival difference between both groups (P = 0.753). In a multivariate logistic regression model, it was demonstrated that age (P<0.001), dialysis duration (P<0.001), as well as blood albumin (P = 0.012) and low-density lipoprotein (P = 0.009) concentrations were significantly correlated with mortality. CONCLUSIONS: Small kidney size on starting hemodialysis was not related with an augmented risk for death in diabetic patients receiving hemodialysis. Further studies are necessary.

Inhibition of spleen tyrosine kinase decreases donor specific antibody levels in a rat model of sensitization.

Antibody mediated rejection is a major cause of renal allograft loss. Circulating preformed donor specific antibodies (DSA) can result as a consequence of blood transfusion, pregnancy or prior transplantation. Current treatment strategies are limited due to partial or transient efficacy, adverse side-effects or patient unsuitability. Previous in vivo studies exploring autoimmune diseases have shown that spleen tyrosine kinase (SYK) signalling is involved in the development of pathogenic autoantibody. The role of SYK in allogenic antibody production is unknown, and we investigated this in a rodent model of sensitization, established by the transfusion of F344 whole blood into LEW rats. Two-week treatment of sensitized rats with selective SYK inhibitor fostamatinib strongly blocked circulating DSA production without affecting overall total immunoglobulin levels, and inhibition was sustained up to 5 weeks post-completion of the treatment regimen. Fostamatinib treatment did not affect mature B cell subset or plasma cell levels, which remained similar between non-treated controls, vehicle treated and fostamatinib treated animals. Our data indicate fostamatinib may provide an alternative therapeutic option for patients who are at risk of sensitization following blood transfusion while awaiting renal transplant.

A novel model of nephrotic syndrome results from a point mutation in Lama5 and is modified by genetic background.

Nephrotic syndrome is characterized by severe proteinuria, hypoalbuminaemia, edema and hyperlipidaemia. Genetic studies of nephrotic syndrome have led to the identification of proteins playing a crucial role in slit diaphragm signaling, regulation of actin cytoskeleton dynamics and cell-matrix interactions. The laminin α5 chain is essential for embryonic development and, in association with laminin ß2 and laminin γ1, is a major component of the glomerular basement membrane, a critical component of the glomerular filtration barrier. Mutations in LAMA5 were recently identified in children with nephrotic syndrome. Here, we have identified a novel missense mutation (E884G) in the uncharacterized L4a domain of LAMA5 where homozygous mice develop nephrotic syndrome with severe proteinuria with histological and ultrastructural changes in the glomerulus mimicking the progression seen in most patients. The levels of LAMA5 are reduced in vivo and the assembly of the laminin 521 heterotrimer significantly reduced in vitro. Proteomic analysis of the glomerular extracellular fraction revealed changes in the matrix composition. Importantly, the genetic background of the mice had a significant effect on aspects of disease progression from proteinuria to changes in podocyte morphology. Thus, our novel model will provide insights into pathologic mechanisms of nephrotic syndrome and pathways that influence the response to a dysfunctional glomerular basement membrane that may be important in a range of kidney diseases.

Combination treatment with rituximab, low-dose cyclophosphamide and plasma exchange for severe antineutrophil cytoplasmic antibody-associated vasculitis.

Antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis can present with life-threatening lung-kidney syndromes. However, many controlled treatment trials excluded patients with diffuse alveolar hemorrhage or severely impaired glomerular filtration rates, and so the optimum treatment in these cases is unclear. In this retrospective cohort study, we report the outcomes of 64 patients with life-threatening disease treated with a combination regimen of rituximab, low-dose intravenous cyclophosphamide, oral glucocorticoids, and plasma exchange. At entry, the median estimated glomerular filtration rate was 9 mL/min, 47% of patients required dialysis, and 52% had diffuse alveolar hemorrhage. All patients received a minimum of seven plasma exchanges, and the median cumulative doses of rituximab, cyclophosphamide, and glucocorticoid were 2, 3, and 2.6 g, respectively, at six months. A total of 94% of patients had achieved disease remission (version 3 Birmingham Vasculitis Activity Score of 0) at this time point, and 67% of patients who required dialysis recovered independent kidney function. During long-term follow-up (median duration 46 months), overall patient survival was 85%, and 69% of patients remained free from end-stage kidney disease, which compares favorably to a historic cohort with severe disease treated with a conventional induction regimen. Combination treatment was associated with prolonged B cell depletion and low rates of relapse; 87% of patients were in continuous remission at month 36. The serious infection rate during total follow-up was 0.28 infections/patient/year, suggesting that combination treatment is not associated with an enduring risk of infection. Thus, we suggest that combination immunosuppressive therapy may permit glucocorticoid avoidance and provide rapid and prolonged disease control in patients with severe ANCA-associated vasculitis.

Impact of kidney size on mortality in diabetic patients receiving peritoneal dialysis.

Although patients with diabetes mellitus mostly present with enlarged or normal-sized kidneys throughout their life, a small proportion of patients have small kidneys. This longitudinal study enrolled 83 diabetic patients treated with peritoneal dialysis (PD) between 2015 and 2019. Patients were stratified into two groups, those with enlarged or normal (n = 67) or small (n = 16) kidneys, based on their kidney sizes before dialysis. Patients with small kidney size were not only older (76.63 ± 10.63 vs. 68.03 ± 11.26 years, P = 0.007), suffered longer duration of diabetes mellitus (272.09 ± 305.09 vs. 151.44 ± 85.31 month, P = 0.006) and predominantly female (75.0 vs. 41.8%, P = 0.017), but also had lower serum levels of creatinine (9.63 ± 2.82 vs. 11.74 ± 3.32 mg/dL, P = 0.022) and albumin (3.23 ± 0.67 vs. 3.60 ± 0.47 g/dL, P = 0.010) than patients with enlarged or normal kidney size. At the end of analysis, 14 (16.9%) patients died. Patients with small kidney size demonstrated higher all-cause (50.0 vs. 9.0%, P < 0.001) and infection-related (43.8 vs. 7.5%, P < 0.001) mortality than patients with enlarged or normal kidney size. In a multivariate-logistic-regression model, small kidney size was a powerful predictor of mortality (odds ratio 6.452, 95% confidence interval 1.220-34.482, P = 0.028). Diabetic patients with small kidney size at the beginning of PD carry a substantial risk for mortality.

A High-Content Screen for Mucin-1-Reducing Compounds Identifies Fostamatinib as a Candidate for Rapid Repurposing for Acute Lung Injury.

Drug repurposing has the advantage of identifying potential treatments on a shortened timescale. In response to the pandemic spread of SARS-CoV-2, we took advantage of a high-content screen of 3,713 compounds at different stages of clinical development to identify FDA-approved compounds that reduce mucin-1 (MUC1) protein abundance. Elevated MUC1 levels predict the development of acute lung injury (ALI) and acute respiratory distress syndrome (ARDS) and correlate with poor clinical outcomes. Our screen identifies fostamatinib (R788), an inhibitor of spleen tyrosine kinase (SYK) approved for the treatment of chronic immune thrombocytopenia, as a repurposing candidate for the treatment of ALI. In vivo, fostamatinib reduces MUC1 abundance in lung epithelial cells in a mouse model of ALI. In vitro, SYK inhibition by the active metabolite R406 promotes MUC1 removal from the cell surface. Our work suggests fostamatinib as a repurposing drug candidate for ALI.

Global microRNA profiling in human urinary exosomes reveals novel disease biomarkers and cellular pathways for autosomal dominant polycystic kidney disease.

MicroRNAs (miRNAs) play an important role in regulating gene expression in health and disease but their role in modifying disease expression in Autosomal Dominant Polycystic Kidney Disease (ADPKD) remains uncertain. Here, we profiled human urinary exosome miRNA by global small RNA-sequencing in an initial discovery cohort of seven patients with ADPKD with early disease (eGFR over 60ml/min/1.73m2), nine with late disease (eGFR under 60ml/min/1.73m2), and compared their differential expression with six age and sex matched healthy controls. Two kidney-enriched candidate miRNA families were identified (miR-192/miR-194-2 and miR-30) and selected for confirmatory testing in a 60 patient validation cohort by quantitative polymerase chain reaction. We confirmed that miR-192-5p, miR-194-5p, miR-30a-5p, miR-30d-5p and miR-30e-5p were significantly downregulated in patient urine exosomes, in murine Pkd1 cystic kidneys and in human PKD1 cystic kidney tissue. All five miRNAs showed significant correlations with baseline eGFR and ultrasound-determined mean kidney length and improved the diagnostic performance (area under the curve) of mean kidney length for the rate of disease progression. Finally, inverse correlations of these two miRNA families with increased expression in their predicted target genes in patient PKD1 cystic tissue identified dysregulated pathways and transcriptional networks including novel interactions between miR-194-5p and two potentially relevant candidate genes, PIK3R1 and ANO1. Thus, our results identify a subset of urinary exosomal miRNAs that could serve as novel biomarkers of disease progression and suggest new therapeutic targets in ADPKD.

A High Content Screen for Mucin-1-Reducing Compounds Identifies Fostamatinib as a Candidate for Rapid Repurposing for Acute Lung Injury during the COVID-19 pandemic.

Drug repurposing is the only method capable of delivering treatments on the shortened time-scale required for patients afflicted with lung disease arising from SARS-CoV-2 infection. Mucin-1 (MUC1), a membrane-bound molecule expressed on the apical surfaces of most mucosal epithelial cells, is a biochemical marker whose elevated levels predict the development of acute lung injury (ALI) and respiratory distress syndrome (ARDS), and correlate with poor clinical outcomes. In response to the pandemic spread of SARS-CoV-2, we took advantage of a high content screen of 3,713 compounds at different stages of clinical development to identify FDA-approved compounds that reduce MUC1 protein abundance. Our screen identified Fostamatinib (R788), an inhibitor of spleen tyrosine kinase (SYK) approved for the treatment of chronic immune thrombocytopenia, as a repurposing candidate for the treatment of ALI. In vivo , Fostamatinib reduced MUC1 abundance in lung epithelial cells in a mouse model of ALI. In vitro , SYK inhibition by Fostamatinib promoted MUC1 removal from the cell surface. Our work reveals Fostamatinib as a repurposing drug candidate for ALI and provides the rationale for rapidly standing up clinical trials to test Fostamatinib efficacy in patients with COVID-19 lung injury.

Spleen tyrosine kinase inhibition is an effective treatment for established vasculitis in a pre-clinical model.

The anti-neutrophil cytoplasm antibody (ANCA)-associated vasculitides (AAV) are a group of life-threatening multi-system diseases characterized by necrotising inflammation of small blood vessels and crescentic glomerulonephritis. ANCA are thought to play a direct pathogenic role. Previous studies have shown that spleen tyrosine kinase (SYK) is phosphorylated during ANCA-induced neutrophil activation in vitro. However, the role of SYK in vivo is unknown. Here, we studied its role in the pathogenesis of experimental autoimmune vasculitis, a pre-clinical model of myeloperoxidase-ANCA-induced pauci-immune systemic vasculitis in the Wistar Kyoto rat. Up-regulation of SYK expression in inflamed renal and pulmonary tissue during early autoimmune vasculitis was confirmed by immunohistochemical and transcript analysis. R406, the active metabolite of fostamatinib, a small molecule kinase inhibitor with high selectivity for SYK, inhibited ANCA-induced pro-inflammatory responses in rat leucocytes in vitro. In an in vivo study, treatment with fostamatinib for 14 days after disease onset resulted in rapid resolution of urinary abnormalities, significantly improved renal and pulmonary pathology, and preserved renal function. Short-term exposure to fostamatinib did not significantly affect circulating myeloperoxidase-ANCA levels, suggesting inhibition of ANCA-induced inflammatory mechanisms in vivo. Finally, SYK expression was demonstrated within inflammatory glomerular lesions in ANCA-associated glomerulonephritis in patients, particularly within CD68+ve monocytes/macrophages. Thus, our data indicate that SYK inhibition warrants clinical investigation in the treatment of AAV.

P2X7 Receptor Stimulation Is Not Required for Oxalate Crystal-Induced Kidney Injury.

Oxalate crystal-induced renal inflammation is associated with progressive kidney failure due to activation of the NLRP3/CASP-1 inflammasome. It has been suggested previously that purinergic P2X7 receptor signaling is critical for crystal-induced inflammasome activation and renal injury. Therefore, we investigated the role of the P2X7 receptor in response to crystal-induced cytokine release, inflammation, and kidney failure using in vitro and in vivo models. Dendritic cells and macrophages derived from murine bone marrow and human peripheral blood mononucleated cells stimulated with calcium-oxalate crystals, monosodium urate crystals, or ATP lead to the robust release of interleukin-1beta (IL-1ß). Treatment with the P2X7 inhibitor A740003 or the depletion of ATP by apyrase selectively abrogated ATP-induced, but not oxalate and urate crystal-induced IL-1ß release. In line with this finding, dendritic cells derived from bone marrow (BMDCs) from P2X7-/- mice released reduced amounts of IL-1ß following stimulation with ATP, while oxalate and urate crystal-induced IL-1ß release was unaffected. In sharp contrast, BMDCs from Casp1-/- mice exhibited reduced IL-1ß release following either of the three stimulants. In addition, P2X7-/- mice demonstrated similar degrees of crystal deposition, tubular damage and inflammation when compared with WT mice. In line with these findings, increases in plasma creatinine were no different between WT and P2X7-/- mice. In contrast to previous reports, our results indicate that P2X7 receptor is not required for crystal-induced CKD and it is unlikely to be a suitable therapeutic target for crystal-induced progressive kidney disease.