The sGC Activator Runcaciguat Has Kidney Protective Effects and Prevents a Decline of Kidney Function in ZSF1 Rats.

Chronic kidney disease (CKD) progression is associated with persisting oxidative stress, which impairs the NO-sGC-cGMP signaling cascade through the formation of oxidized and heme-free apo-sGC that cannot be activated by NO. Runcaciguat (BAY 1101042) is a novel, potent, and selective sGC activator that binds and activates oxidized and heme-free sGC and thereby restores NO-sGC-cGMP signaling under oxidative stress. Therefore, runcaciguat might represent a very effective treatment option for CKD/DKD. The potential kidney-protective effects of runcaciguat were investigated in ZSF1 rats as a model of CKD/DKD, characterized by hypertension, hyperglycemia, obesity, and insulin resistance. ZSF1 rats were treated daily orally for up to 12 weeks with runcaciguat (1, 3, 10 mg/kg/bid) or placebo. The study endpoints were proteinuria, kidney histopathology, plasma, urinary biomarkers of kidney damage, and gene expression profiling to gain information about relevant pathways affected by runcaciguat. Furthermore, oxidative stress was compared in the ZSF1 rat kidney with kidney samples from DKD patients. Within the duration of the 12-week treatment study, kidney function was significantly decreased in obese ZSF1 rats, indicated by a 20-fold increase in proteinuria, compared to lean ZSF1 rats. Runcaciguat dose-dependently and significantly attenuated the development of proteinuria in ZSF1 rats with reduced uPCR at the end of the study by -19%, -54%, and -70% at 1, 3, and 10 mg/kg/bid, respectively, compared to placebo treatment. Additionally, average blood glucose levels measured as HbA1C, triglycerides, and cholesterol were increased by five times, twenty times, and four times, respectively, in obese ZSF1 compared to lean rats. In obese ZSF1 rats, runcaciguat reduced HbA1c levels by -8%, -34%, and -76%, triglycerides by -42%, -55%, and -71%, and cholesterol by -16%, -17%, and -34%, at 1, 3, and 10 mg/kg/bid, respectively, compared to placebo. Concomitantly, runcaciguat also reduced kidney weights, morphological kidney damage, and urinary and plasma biomarkers of kidney damage. Beneficial effects were accompanied by changes in gene expression that indicate reduced fibrosis and inflammation and suggest improved endothelial stabilization. In summary, the sGC activator runcaciguat significantly prevented a decline in kidney function in a DKD rat model that mimics common comorbidities and conditions of oxidative stress of CKD patients. Thus, runcaciguat represents a promising treatment option for CKD patients, which is in line with recent phase 2 clinical study data, where runcaciguat showed promising efficacy in CKD patients (NCT04507061).

Treatment effects of soluble guanylate cyclase modulation on diabetic kidney disease at single-cell resolution.

Diabetic kidney disease (DKD) is the most common cause of renal failure. Therapeutics development is hampered by our incomplete understanding of animal models on a cellular level. We show that ZSF1 rats recapitulate human DKD on a phenotypic and transcriptomic level. Tensor decomposition prioritizes proximal tubule (PT) and stroma as phenotype-relevant cell types exhibiting a continuous lineage relationship. As DKD features endothelial dysfunction, oxidative stress, and nitric oxide depletion, soluble guanylate cyclase (sGC) is a promising DKD drug target. sGC expression is specifically enriched in PT and stroma. In ZSF1 rats, pharmacological sGC activation confers considerable benefits over stimulation and is mechanistically related to improved oxidative stress regulation, resulting in enhanced downstream cGMP effects. Finally, we define sGC gene co-expression modules, which allow stratification of human kidney samples by DKD prevalence and disease-relevant measures such as kidney function, proteinuria, and fibrosis, underscoring the relevance of the sGC pathway to patients.

Prevalence, phenotypic characteristics and prognostic role of apparent treatment resistant hypertension in the German Chronic Kidney Disease (GCKD) study.

Treatment resistant hypertension (TRH) appears of particular relevance in patients with chronic kidney disease (CKD). However, causes and consequences of TRH in CKD patients remain incompletely understood. Therefore, we analyzed the prevalence of apparent TRH (aTRH), and phenotypic characteristics and prognosis associated with aTRH among participants of the German Chronic Kidney Disease (GCKD) study. As insufficient medication adherence has been shown to be a frequent cause of pseudoresistance, we also assessed treatment adherence. Study participants were classified as having aTRH, controlled hypertension and uncontrolled hypertension based on study visit blood pressure and self-reported medication intake. Drug adherence was assessed by comparing self-reported antihypertensive medication with detectable urinary drug metabolites measured by mass spectroscopy. Out of 4901 individuals included in this study, 38% were classified as having aTRH. Male sex, older age, lower estimated glomerular filtration rate (eGFR), higher body mass index (BMI), higher urine albumin-to-creatinine ratio (UACR) and presence of diabetes mellitus were independently associated with higher prevalence of aTRH in a multivariable adjusted regression model. Patients classified as aTRH had higher risk for major adverse cardiovascular events and worsening of kidney disease compared to patients with no aTRH after multivariate adjustment for potential confounders. There was a high agreement between self-reported medication and detectable urinary drug metabolites. In conclusion, in a cohort of Caucasian patients with moderately severe CKD, aTRH was highly prevalent and, in most cases, likely not caused by low medication adherence. Furthermore, aTRH was linked to cardio-renal endpoints, emphasizing the need for improved management.

BAY-7081: A Potent, Selective, and Orally Bioavailable Cyanopyridone-Based PDE9A Inhibitor.

Despite advances in the treatment of heart failure in recent years, options for patients are still limited and the disease is associated with considerable morbidity and mortality. Modulating cyclic guanosine monophosphate levels within the natriuretic peptide signaling pathway by inhibiting PDE9A has been associated with beneficial effects in preclinical heart failure models. We herein report the identification of BAY-7081, a potent, selective, and orally bioavailable PDE9A inhibitor with very good aqueous solubility starting from a high-throughput screening hit. Key aspect of the optimization was a switch in metabolism of our lead structures from glucuronidation to oxidation. The switch proved being essential for the identification of compounds with improved pharmacokinetic profiles. By studying a tool compound in a transverse aortic constriction mouse model, we were able to substantiate the relevance of PDE9A inhibition in heart diseases.

Genetic regulation of serum IgA levels and susceptibility to common immune, infectious, kidney, and cardio-metabolic traits.

Immunoglobulin A (IgA) mediates mucosal responses to food antigens and the intestinal microbiome and is involved in susceptibility to mucosal pathogens, celiac disease, inflammatory bowel disease, and IgA nephropathy. We performed a genome-wide association study of serum IgA levels in 41,263 individuals of diverse ancestries and identified 20 genome-wide significant loci, including 9 known and 11 novel loci. Co-localization analyses with expression QTLs prioritized candidate genes for 14 of 20 significant loci. Most loci encoded genes that produced immune defects and IgA abnormalities when genetically manipulated in mice. We also observed positive genetic correlations of serum IgA levels with IgA nephropathy, type 2 diabetes, and body mass index, and negative correlations with celiac disease, inflammatory bowel disease, and several infections. Mendelian randomization supported elevated serum IgA as a causal factor in IgA nephropathy. African ancestry was consistently associated with higher serum IgA levels and greater frequency of IgA-increasing alleles compared to other ancestries. Our findings provide novel insights into the genetic regulation of IgA levels and its potential role in human disease.

TRPM2 Plays a Minor Role in AKI and Kidney Fibrosis.

TRPM2 is a Ca2+-permeable cationic channel and serves as an oxidative stress sensor.TRPM2 deletion was harmful in renal ischemia-reperfusion injury, whereas TRPM2 deletion mitigated kidney fibrosis.Our findings suggest the role of TRPM2 in kidney diseases is context dependent.

Novel soluble guanylyl cyclase activators increase glomerular cGMP, induce vasodilation and improve blood flow in the murine kidney.

BACKGROUND AND PURPOSE: Generation of cGMP via NO-sensitive soluble guanylyl cyclase (sGC) has been implicated in the regulation of renal functions. Chronic kidney disease (CKD) is associated with decreased NO bioavailability, increased oxidative stress and oxidation of sGC to its haem-free form, apo-sGC. Apo-sGC cannot be activated by NO, resulting in impaired cGMP signalling that is associated with chronic kidney disease progression. We hypothesised that sGC activators, which activate apo-sGC independently of NO, increase renal cGMP production under conditions of oxidative stress, thereby improving renal blood flow (RBF) and kidney function. EXPERIMENTAL APPROACH: Two novel sGC activators, runcaciguat and BAY-543, were tested on murine kidney. We measured cGMP levels in real time in kidney slices of cGMP sensor mice, vasodilation of pre-constricted glomerular arterioles and RBF in isolated perfused kidneys. Experiments were performed at baseline conditions, under L-NAME-induced NO deficiency, and in the presence of oxidative stress induced by ODQ. KEY RESULTS: Mouse glomeruli showed NO-induced cGMP increases. Under baseline conditions, sGC activator did not alter glomerular cGMP concentration or NO-induced cGMP generation. In the presence of ODQ, NO-induced glomerular cGMP signals were markedly reduced, whereas sGC activator induced strong cGMP increases. L-NAME and ODQ pretreated isolated glomerular arterioles were strongly dilated by sGC activator. sGC activator also increased cGMP and RBF in ODQ-perfused kidneys. CONCLUSION AND IMPLICATION: sGC activators increase glomerular cGMP, dilate glomerular arterioles and improve RBF under disease-relevant oxidative stress conditions. Therefore, sGC activators represent a promising class of drugs for chronic kidney disease treatment. LINKED ARTICLES: This article is part of a themed issue on cGMP Signalling in Cell Growth and Survival. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v179.11/issuetoc.

Sirolimus in renal transplant recipients with malignancies in Germany.

BACKGROUND: Renal transplant recipients have an increased cancer risk. The mammalian target of rapamycin inhibitor sirolimus (SRL) has immunosuppressive and antitumour activities but knowledge about its use in recipients with cancer is limited. METHODS: We retrospectively analysed 726 renal allograft recipients converted to SRL from 10 German transplant centres. Patient and graft survival were analysed depending on malignancy status prior to conversion and tumour entity. RESULTS: Malignancy before conversion to SRL was reported in 230 patients, with 137 patients having skin cancers and 101 having solid cancers. Cancer occurred 4.6 ± 9.4 (median 3.0) years after transplantation. Basal cell carcinoma, squamous cell carcinoma and Bowen's disease were the most prevalent skin cancers, while carcinomas of the kidney, colon and breast were the most prevalent solid cancers before conversion. Patients with prior malignancy were older and had better renal function at conversion compared with patients without a history of cancer. After conversion to SRL, cancer incidence rates (IRs) of all tumours were lower compared with rates before conversion. Cancer IRs after conversion were higher in patients with malignancy before conversion compared with those without. Patient survival was worse in patients with solid cancers compared with patients with skin cancers or without malignancies. Biopsy-proven acute rejections in the first year after conversion were less frequent in patients with malignancy compared with those without. Graft survival and renal function in all cancer types were better than in patients converted to SRL without cancers. CONCLUSIONS: Conversion to SRL in patients with a history of cancer is safe regarding renal function and graft survival, while patient survival is largely dependent on tumour entity.

Runcaciguat, a novel soluble guanylate cyclase activator, shows renoprotection in hypertensive, diabetic, and metabolic preclinical models of chronic kidney disease.

Chronic kidney diseaQueryse (CKD) is associated with oxidative stress which can interrupt the nitric oxide (NO)/soluble guanylyl cyclase (sGC) signaling and decrease cyclic guanosine monophosphate (cGMP) production. Low cGMP concentrations can cause kidney damage and progression of CKD. The novel sGC activator runcaciguat targets the oxidized and heme-free form of sGC, restoring cGMP production under oxidative stress. The purpose of this study is to investigate if runcaciguat could provide an effective treatment for CKD. Runcaciguat was used for the treatment not only in rat CKD models with different etiologies and comorbidities, namely of hypertensive rats, the renin transgenic (RenTG) rat, and angiotensin-supplemented (ANG-SD) rat, but also in rats with diabetic and metabolic CKD, the Zucker diabetic fatty (ZDF) rat. The treatment duration was 2 to 42 weeks and runcaciguat was applied orally in doses from 1 to 10 mg/kg/bid. In these different rat CKD models, runcaciguat significantly reduced proteinuria (urinary protein to creatinine ratio; uPCR). These effects were also significant at doses which did not or only moderately decrease systemic blood pressure. Moreover, runcaciguat significantly decreased kidney injury biomarkers and attenuated morphological kidney damages. In RenTG rats, runcaciguat improved survival rates and markers of heart injury. These data demonstrate that the sGC activator runcaciguat exhibits cardio-renal protection at doses which did not reduce blood pressure and was effective in hypertensive as well as diabetic and metabolic CKD models. These data, therefore, suggest that runcaciguat, with its specific mode of action, represents an efficient treatment approach for CKD and associated CV diseases.

Direct Blood Pressure-Independent Anti-Fibrotic Effects by the Selective Nonsteroidal Mineralocorticoid Receptor Antagonist Finerenone in Progressive Models of Kidney Fibrosis.

INTRODUCTION: The nonsteroidal mineralocorticoid receptor (MR) antagonist finerenone and sodium-glucose cotransporter-2 (SGLT2) inhibitors have demonstrated clinical benefits in chronic kidney disease patients with type 2 diabetes. Precise molecular mechanisms responsible for these benefits are incompletely understood. Here, we investigated potential direct anti-fibrotic effects and mechanisms of nonsteroidal MR antagonism by finerenone or SGLT2 inhibition by empagliflozin in 2 relevant mouse kidney fibrosis models: unilateral ureter obstruction and sub-chronic ischemia reperfusion injury. METHODS: Kidney fibrosis was induced in mice via unilateral ureteral obstruction or ischemia. In a series of experiments, mice were treated orally with the MR antagonist finerenone (3 or 10 mg/kg), the SGLT2 inhibitor empagliflozin (10 or 30 mg/kg), or in a direct comparison of both drugs. Interstitial myofibroblast accumulation was quantified via alpha-smooth muscle actin and interstitial collagen deposition via Sirius Red/Fast Green staining in both models. Secondary analyses included the assessment of inflammatory cells, kidney mRNA expression of fibrotic markers as well as functional parameters (serum creatinine and albuminuria) in the ischemic model. Blood pressure was measured via telemetry in healthy conscious compound-treated animals. RESULTS: Finerenone dose-dependently decreased pathological myofibroblast accumulation and collagen deposition with no effects on systemic blood pressure and inflammatory markers in the tested dose range. Reduced kidney fibrosis was paralleled by reduced kidney plasminogen activator inhibitor-1 (PAI-1) and naked cuticle 2 (NKD2) expression in finerenone-treated mice. In contrast, treatment with empagliflozin strongly increased urinary glucose excretion in both models and reduced ischemia-induced albuminuria but had no effects on kidney myofibroblasts or collagen deposition. DISCUSSION/CONCLUSION: Finerenone has direct anti-fibrotic properties resulting in reduced myofibroblast and collagen deposition accompanied by a reduction in renal PAI-1 and NKD2 expression in mouse models of progressive kidney fibrosis at blood pressure-independent dosages.

Effects of Finerenone Combined with Empagliflozin in a Model of Hypertension-Induced End-Organ Damage.

INTRODUCTION: The nonsteroidal mineralocorticoid receptor (MR) antagonist finerenone and sodium-glucose cotransporter-2 (SGLT2) inhibitors have demonstrated clinical benefits in CKD patients with type 2 diabetes. Clinical data analyzing the potential value of a combination therapy are currently limited. We therefore investigated cardiorenal protection of respective mono- and combination therapy in a preclinical model of hypertension-induced end-organ damage. METHODS: Cardiovascular (CV) morbidity and mortality were studied in hypertensive, N(ω)-nitro-L-arginine methyl ester-treated, renin-transgenic (mRen2)27 rats. Rats (10- to 11-week-old females, n = 13-17/group) were treated once daily orally for up to 7 weeks with placebo, finerenone (1 and 3 mg/kg), empagliflozin (3 and 10 mg/kg), or a combination of the respective low doses. Key outcome parameters included mortality, proteinuria, plasma creatinine and uric acid, blood pressure, and cardiac and renal histology. RESULTS: Placebo-treated rats demonstrated a 50% survival rate over the course of 7 weeks. Drug treatment resulted in variable degrees of survival benefit, most prominently in the low-dose combination group with a survival benefit of 93%. Monotherapies of finerenone or empagliflozin dose-dependently reduced proteinuria, while low-dose combination revealed an early, sustained, and over-additive reduction in proteinuria. Empagliflozin induced a strong and dose-dependent increase in urinary glucose excretion which was not influenced by finerenone coadministration in the combination arm. Low-dose combination but not respective low-dose monotherapies significantly reduced plasma creatinine and plasma uric acid after 6 weeks. Treatment with finerenone and the low-dose combination significantly decreased systolic blood pressure after 5 weeks. There was a dose-dependent protection from cardiac and kidney fibrosis and vasculopathy with both agents, while low-dose combination therapy was more efficient than the respective monotherapy dosages on most cardiorenal histology parameters. DISCUSSION/CONCLUSIONS: Nonsteroidal MR antagonism by finerenone and SGLT2 inhibition by empagliflozin confer CV protection in preclinical hypertension-induced cardiorenal disease. Combination of these 2 independent modes of action at low dosages revealed efficacious reduction in important functional parameters such as proteinuria and blood pressure, plasma markers including creatinine and uric acid, cardiac and renal lesions as determined by histopathology, and mortality indicating a strong potential for combined clinical use in cardiorenal patient populations.

A flow cytometry approach reveals heterogeneity in conventional subsets of murine renal mononuclear phagocytes.

Mononuclear phagocytes (MNPs) participate in inflammation and repair after kidney injury, reflecting their complex nature. Dissection into refined functional subunits has been challenging and would benefit understanding of renal pathologies. Flow cytometric approaches are limited to classifications of either different MNP subsets or functional state. We sought to combine these two dimensions in one protocol that considers functional heterogeneity in each MNP subset. We identified five distinct renal MNP subsets based on a previously described strategy. In vitro polarization of bone marrow-derived macrophages (BMDM) into M1- and M2-like cells suggested functional distinction of CD86 + MHCII + CD206- and CD206 + cells. Combination of both distinction methods identified CD86 + MHCII + CD206- and CD206 + cells in all five MNP subsets, revealing their heterologous nature. Our approach revealed that MNP composition and their functional segmentation varied between different mouse models of kidney injury and, moreover, was dynamically regulated in a time-dependent manner. CD206 + cells from three analyzed MNP subsets had a higher ex vivo phagocytic capacity than CD86 + MHCII + CD206- counterparts, indicating functional uniqueness of each subset. In conclusion, our novel flow cytometric approach refines insights into renal MNP heterogeneity and therefore could benefit mechanistic understanding of renal pathology.

Discovery of the Soluble Guanylate Cyclase Activator Runcaciguat (BAY 1101042).

Herein we describe the discovery, mode of action, and preclinical characterization of the soluble guanylate cyclase (sGC) activator runcaciguat. The sGC enzyme, via the formation of cyclic guanosine monophoshphate, is a key regulator of body and tissue homeostasis. sGC activators with their unique mode of action are activating the oxidized and heme-free and therefore NO-unresponsive form of sGC, which is formed under oxidative stress. The first generation of sGC activators like cinaciguat or ataciguat exhibited limitations and were discontinued. We overcame limitations of first-generation sGC activators and identified a new chemical class via high-throughput screening. The investigation of the structure-activity relationship allowed to improve potency and multiple solubility, permeability, metabolism, and drug-drug interactions parameters. This program resulted in the discovery of the oral sGC activator runcaciguat (compound 45, BAY 1101042). Runcaciguat is currently investigated in clinical phase 2 studies for the treatment of patients with chronic kidney disease and nonproliferative diabetic retinopathy.

Monitoring transcellular fluid shifts during episodes of intradialytic hypotension using bioimpedance spectroscopy.

BACKGROUND: Transcellular fluid shifts during dialysis treatment could be related to the frequency and severity of intradialytic hypotension (IDH). We investigated that (i) in addition to ultrafiltration, extracellular fluid (ECF) is further depleted by transcellular fluid shifts and (ii) changes in intracellular fluid (ICF), which have been overlooked so far, or if they were considered, are not understood, might be due to these fluid shifts. METHODS: Thirty-six patients were categorized as haemodynamically stable, asymptomatic IDH or unstable (symptomatic IDH) according to their changes in systolic blood pressure and associated clinical symptoms. Their intradialytic changes in body fluids were studied using bioimpedance spectroscopy measurements and compared among groups. RESULTS: For IDH-prone patients, data showed a rapid drop in ECF that was more than expected from the ultrafiltration rate (UFR) profile and was associated with a significant increase in ICF (P = 0.001). Study of accumulative loss profiles of ECF revealed a loss in ECF up to 300 ml, more than that predicted from UFR for unstable patients. CONCLUSIONS: The considerable discrepancy between the expected and measured loss in ECF might provide evidence of transcellular fluid shifts possibly induced by changes in plasma osmolarity due to haemodialysis. Moreover, the results suggest a pattern of fluid removal in IDH-prone patients that significantly differs from that in haemodynamically stable patients.

Pharmacology of Free Fatty Acid Receptors and Their Allosteric Modulators.

The physiological function of free fatty acids (FFAs) has long been regarded as indirect in terms of their activities as educts and products in metabolic pathways. The observation that FFAs can also act as signaling molecules at FFA receptors (FFARs), a family of G protein-coupled receptors (GPCRs), has changed the understanding of the interplay of metabolites and host responses. Free fatty acids of different chain lengths and saturation statuses activate FFARs as endogenous agonists via binding at the orthosteric receptor site. After FFAR deorphanization, researchers from the pharmaceutical industry as well as academia have identified several ligands targeting allosteric sites of FFARs with the aim of developing drugs to treat various diseases such as metabolic, (auto)inflammatory, infectious, endocrinological, cardiovascular, and renal disorders. GPCRs are the largest group of transmembrane proteins and constitute the most successful drug targets in medical history. To leverage the rich biology of this target class, the drug industry seeks alternative approaches to address GPCR signaling. Allosteric GPCR ligands are recognized as attractive modalities because of their auspicious pharmacological profiles compared to orthosteric ligands. While the majority of marketed GPCR drugs interact exclusively with the orthosteric binding site, allosteric mechanisms in GPCR biology stay medically underexploited, with only several allosteric ligands currently approved. This review summarizes the current knowledge on the biology of FFAR1 (GPR40), FFAR2 (GPR43), FFAR3 (GPR41), FFAR4 (GPR120), and GPR84, including structural aspects of FFAR1, and discusses the molecular pharmacology of FFAR allosteric ligands as well as the opportunities and challenges in research from the perspective of drug discovery.

Pharmacological inhibition of Vanin-1 is not protective in models of acute and chronic kidney disease.

Oxidative stress is a key concept in basic, translational, and clinical research to understand the pathophysiology of various disorders, including cardiovascular and renal diseases. Although attempts to directly reduce oxidative stress with redox-active substances have until now largely failed to prove clinical benefit, indirect approaches to combat oxidative stress enzymatically have gained further attention as potential therapeutic strategies. The pantetheinase Vanin-1 is expressed on kidney proximal tubular cells, and its reaction product cysteamine is described to negatively affect redox homeostasis by inhibiting the replenishment of cellular antioxidative glutathione stores. Vanin-1-deficient mice were shown to be protected against oxidative stress damage. The aim of this study was to elucidate whether pharmacological inhibition of Vanin-1 protects mice from oxidative stress-related acute or chronic kidney injury as well. By studying renal ischemia-reperfusion injury in Col4α3-/- (Alport syndrome) mice and in vitro hypoxia-reoxygenation in human proximal tubular cells we found that treatment with a selective and potent Vanin-1 inhibitor resulted in ample inhibition of enzymatic activity in vitro and in vivo. However, surrogate parameters of metabolic and redox homeostasis were only partially and insufficiently affected. Consequently, apoptosis and reactive oxygen species level in tubular cells as well as overall kidney function and fibrotic processes were not improved by Vanin-1 inhibition. We thus conclude that Vanin-1 functionality in the context of cardiovascular diseases needs further investigation and the biological relevance of pharmacological Vanin-1 inhibition for the treatment of kidney diseases remains to be proven.

Allosteric targeting of the FFA2 receptor (GPR43) restores responsiveness of desensitized human neutrophils.

The G protein-coupled free fatty acid receptor 2 (FFA2R) is highly expressed on neutrophils and was previously described to regulate neutrophil activation. Allosteric targeting of G protein-coupled receptors (GPCRs) is increasingly explored to create distinct pharmacology compared to endogenous, orthosteric ligands. The consequence of allosteric versus orthosteric FFA2R activation for neutrophil response, however, is currently largely elusive. Here, different FFA2R desensitization profiles in human neutrophils following allosteric or orthosteric activation are reported. Using a set of neutrophil functional assays to measure calcium flux, pERK1/2, chemotaxis, cellular degranulation, and oxidative burst together with holistic and pathway-unbiased whole cell sensing based on dynamic mass redistribution, it is found that the synthetic positive allosteric modulator agonist 4-CMTB potently activates neutrophils and simultaneously alters FFA2R responsiveness toward the endogenous, orthosteric agonist propionic acid (C3) after homologous and heterologous receptor desensitization. Stimulation with C3 or the hierarchically superior chemokine receptor activator IL-8 led to strong FFA2R desensitization and rendered neutrophils unresponsive toward repeated stimulation with C3. In contrast, stimulation with allosteric 4-CMTB engaged a distinct composition of signaling pathways as compared to orthosteric receptor activation and was able to activate neutrophils that underwent homologous and heterologous desensitization with C3 and IL-8, respectively. Moreover, allosteric FFA2R activation could re-sensitize FFA2 toward the endogenous agonist C3 after homologous and heterologous desensitization. Given the fact that receptor desensitization is critical in neutrophils to sense and adapt to their current environment, these findings are expected to be useful for the discovery of novel pharmacological mechanisms to modulate neutrophil responsiveness therapeutically.

Single versus dual blockade of the renin-angiotensin system in patients with IgA nephropathy.

BACKGROUND: Inhibitors of the renin-angiotensin system (RAS) are cornerstones of supportive therapy in patients with IgA nephropathy (IgAN). We analyzed the effects of single versus dual RAS blockaQueryde during our randomized STOP-IgAN trial. METHODS: STOP-IgAN participants with available successive information on their RAS treatment regimen and renal outcomes during the randomized 3-year trial phase were stratified post hoc into two groups, i.e. patients under continuous single or dual RAS blocker therapy over the entire 3 years of the trial phase. Primary and secondary STOP-IgAN trial endpoints, i.e. frequencies of full clinical remission, eGFR-loss ≥ 15 and ≥ 30 ml/min/1.73 m2 and ESRD onset, were analyzed by logistic regression and linear mixed effects models. RESULTS: Among the 112 patients included in the present analysis, 82 (73%) were maintained on single and 30 (27%) on dual RAS inhibitor therapy throughout the trial. Neither RAS blocker strategy significantly affected full clinical remission, eGFR-loss rates, onset of ESRD. Proteinuria moderately increased in patients under dual RAS blockade by 0.1 g/g creatinine during the 3-year trial phase. This was particularly evident in patients without additional immunosuppression during the randomized trial phase, where proteinuria increased by 0.2 g/g creatinine in the dual RAS blockade group. In contrast, proteinuria decreased in patients under single RAS blocker therapy by 0.3 g/g creatinine. The course of eGFR remained stable and did not differ between the RAS treatment strategies. CONCLUSION: In the STOP-IgAN cohort, neither RAS blocker regimen altered renal outcomes. Patients on dual RAS blockade even exhibited higher proteinuria over the 3-year trial phase.

Determinants of Successful Use of Sirolimus in Renal Transplant Patients.

BACKGROUND: Sirolimus is an established immunosuppressant in renal transplantation with antineoplastic and antiviral features, but side effects like proteinuria limit its use. The aim of this retrospective multicenter observational study is to define predictors for determining which patients most likely benefit from a sirolimus-based therapy. METHODS: All patients from 10 German centers that were switched to a sirolimus-containing maintenance immunosuppression in 2000 to 2008 after 3 months or later post-transplantation were enrolled (n = 726). Observation times after switching to sirolimus ranged from 4 days to 9 years (median: 24.3 months). With multinomial logistic regression, risk factors for the endpoints terminal graft failure and withdrawal of sirolimus therapy compared to successful therapy were identified. RESULTS: Successful sirolimus therapy was observed in 304 patients. Forty patients died with functioning graft. Therapy failures included graft loss (n = 106) and sirolimus-discontinuation for various reasons (n = 276). Successful sirolimus-use was predicted in 83% and graft failure in 65%, whereas prediction of deliberate sirolimus-discontinuation was poor (48%). Most favorable results for sirolimus-use were observed in patients switched in 2006 to 2008. Using ROC analysis, an estimated glomerular filtration rate (eGFR) below 32 mL/min was shown to be the cut-off in patients withdrawing from therapy as a result of renal reasons, as well as in patients with graft loss. Proteinuria above 151 mg/L was shown to be predictive for patients with graft failure. CONCLUSIONS: eGFR and proteinuria are the major determinants for successful sirolimus-therapy. Our findings help stratifying patients who will benefit most from this therapy and avoid toxicities in patients without potential benefits for this therapy.