APOL1 genotype-associated morphologic changes among patients with focal segmental glomerulosclerosis.

BACKGROUND: The G1 and G2 alleles of apolipoprotein L1 (APOL1) are common in the Black population and associated with increased risk of focal segmental glomerulosclerosis (FSGS). The molecular mechanisms linking APOL1 risk variants with FSGS are not clearly understood, and APOL1's natural absence in laboratory animals makes studying its pathobiology challenging. METHODS: In a cohort of 90 Black patients with either FSGS or minimal change disease (MCD) enrolled in the Nephrotic Syndrome Study Network (58% pediatric onset), we used kidney biopsy traits as an intermediate outcome to help illuminate tissue-based consequences of APOL1 risk variants and expression. We tested associations between APOL1 risk alleles or glomerular APOL1 mRNA expression and 83 light- or electron-microscopy traits measuring structural and cellular kidney changes. RESULTS: Under both recessive and dominant models in the FSGS patient subgroup (61%), APOL1 risk variants were significantly correlated (defined as FDR <0.1) with decreased global mesangial hypercellularity, decreased condensation of cytoskeleton, and increased tubular microcysts. No significant correlations were detected in MCD cohort. Independent of risk alleles, glomerular APOL1 expression in FSGS patients was not correlated with morphologic features. CONCLUSIONS: While APOL1-associated FSGS is associated with two risk alleles, both one and two risk alleles are associated with cellular/tissue changes in this study of FSGS patients. Our lack of discovery of a large group of tissue differences in FSGS and no significant difference in MCD may be due to the lack of power but also supports investigating whether machine learning methods may more sensitively detect APOL1-associated changes.

Serially assessed bisphenol A and phthalate exposure and association with kidney function in children with chronic kidney disease in the US and Canada: A longitudinal cohort study.

BACKGROUND: Exposure to environmental chemicals may be a modifiable risk factor for progression of chronic kidney disease (CKD). The purpose of this study was to examine the impact of serially assessed exposure to bisphenol A (BPA) and phthalates on measures of kidney function, tubular injury, and oxidative stress over time in a cohort of children with CKD. METHODS AND FINDINGS: Samples were collected between 2005 and 2015 from 618 children and adolescents enrolled in the Chronic Kidney Disease in Children study, an observational cohort study of pediatric CKD patients from the US and Canada. Most study participants were male (63.8%) and white (58.3%), and participants had a median age of 11.0 years (interquartile range 7.6 to 14.6) at the baseline visit. In urine samples collected serially over an average of 3.0 years (standard deviation [SD] 1.6), concentrations of BPA, phthalic acid (PA), and phthalate metabolites were measured as well as biomarkers of tubular injury (kidney injury molecule-1 [KIM-1] and neutrophil gelatinase-associated lipocalin [NGAL]) and oxidative stress (8-hydroxy-2'-deoxyguanosine [8-OHdG] and F2-isoprostane). Clinical renal function measures included estimated glomerular filtration rate (eGFR), proteinuria, and blood pressure. Linear mixed models were fit to estimate the associations between urinary concentrations of 6 chemical exposure measures (i.e., BPA, PA, and 4 phthalate metabolite groups) and clinical renal outcomes and urinary concentrations of KIM-1, NGAL, 8-OHdG, and F2-isoprostane controlling for sex, age, race/ethnicity, glomerular status, birth weight, premature birth, angiotensin-converting enzyme inhibitor use, angiotensin receptor blocker use, BMI z-score for age and sex, and urinary creatinine. Urinary concentrations of BPA, PA, and phthalate metabolites were positively associated with urinary KIM-1, NGAL, 8-OHdG, and F2-isoprostane levels over time. For example, a 1-SD increase in ∑di-n-octyl phthalate metabolites was associated with increases in NGAL (ß = 0.13 [95% CI: 0.05, 0.21], p = 0.001), KIM-1 (ß = 0.30 [95% CI: 0.21, 0.40], p < 0.001), 8-OHdG (ß = 0.10 [95% CI: 0.06, 0.13], p < 0.001), and F2-isoprostane (ß = 0.13 [95% CI: 0.01, 0.25], p = 0.04) over time. BPA and phthalate metabolites were not associated with eGFR, proteinuria, or blood pressure, but PA was associated with lower eGFR over time. For a 1-SD increase in ln-transformed PA, there was an average decrease in eGFR of 0.38 ml/min/1.73 m2 (95% CI: -0.75, -0.01; p = 0.04). Limitations of this study included utilization of spot urine samples for exposure assessment of non-persistent compounds and lack of specific information on potential sources of exposure. CONCLUSIONS: Although BPA and phthalate metabolites were not associated with clinical renal endpoints such as eGFR or proteinuria, there was a consistent pattern of increased tubular injury and oxidative stress over time, which have been shown to affect renal function in the long term. This raises concerns about the potential for clinically significant changes in renal function in relation to exposure to common environmental toxicants at current levels.

CureGN Study Rationale, Design, and Methods: Establishing a Large Prospective Observational Study of Glomerular Disease.

RATIONALE & OBJECTIVES: Glomerular diseases, including minimal change disease, focal segmental glomerulosclerosis, membranous nephropathy, and immunoglobulin A (IgA) nephropathy, share clinical presentations, yet result from multiple biological mechanisms. Challenges to identifying underlying mechanisms, biomarkers, and new therapies include the rarity of each diagnosis and slow progression, often requiring decades to measure the effectiveness of interventions to prevent end-stage kidney disease (ESKD) or death. STUDY DESIGN: Multicenter prospective cohort study. SETTING & PARTICIPANTS: Cure Glomerulonephropathy (CureGN) will enroll 2,400 children and adults with minimal change disease, focal segmental glomerulosclerosis, membranous nephropathy, or IgA nephropathy (including IgA vasculitis) and a first diagnostic kidney biopsy within 5 years. Patients with ESKD and those with secondary causes of glomerular disease are excluded. EXPOSURES: Clinical data, including medical history, medications, family history, and patient-reported outcomes, are obtained, along with a digital archive of kidney biopsy images and blood and urine specimens at study visits aligned with clinical care 1 to 4 times per year. OUTCOMES: Patients are followed up for changes in estimated glomerular filtration rate, disease activity, ESKD, and death and for nonrenal complications of disease and treatment, including infection, malignancy, cardiovascular, and thromboembolic events. ANALYTICAL APPROACH: The study design supports multiple longitudinal analyses leveraging the diverse data domains of CureGN and its ancillary program. At 2,400 patients and an average of 2 years' initial follow-up, CureGN has 80% power to detect an HR of 1.4 to 1.9 for proteinuria remission and a mean difference of 2.1 to 3.0mL/min/1.73m2 in estimated glomerular filtration rate per year. LIMITATIONS: Current follow-up can only detect large differences in ESKD and death outcomes. CONCLUSIONS: Study infrastructure will support a broad range of scientific approaches to identify mechanistically distinct subgroups, identify accurate biomarkers of disease activity and progression, delineate disease-specific treatment targets, and inform future therapeutic trials. CureGN is expected to be among the largest prospective studies of children and adults with glomerular disease, with a broad goal to lessen disease burden and improve outcomes.

Renal Function and exposure to Bisphenol A and phthalates in children with Chronic Kidney Disease.

RATIONALE AND OBJECTIVE: Exposure to Bisphenol A (BPA) and phthalates is ubiquitous among adults and children in the United States. Among children and adolescents, those with chronic kidney disease (CKD) are potentially at greater risk of adverse effects from BPA and phthalate exposure. The objective of this study was to evaluate BPA and phthalate exposure among children with CKD and evaluate associations with three measures of kidney function. STUDY DESIGN: Cross sectional study. SETTING, PARTICIPANTS, AND MEASUREMENTS: The CKD population was represented by the Chronic Kidney Disease in Children (CKiD) Study, a multicenter, prospective cohort study of children with impaired kidney function in the US. The main outcome was assessment of the relationship between chemical exposures and clinical laboratory findings at enrollment into CKiD. Data collected at baseline from participants 1 to 17 years old (N = 538) were analyzed. Urinary BPA and phthalate levels were evaluated at this time point. Data from the National Health and Nutrition Examination Survey (NHANES), a nationally representative pediatric population, were used for comparison to the CKiD cohort. RESULTS: Urinary BPA and phthalate levels in the CKiD population were consistently lower than levels detected in healthy children. Additionally, BPA was not significantly associated with blood pressure, proteinuria, or estimated glomerular filtration rate (eGFR). Within the CKiD population, for select individual and combined phthalates, there was an inverse relationship with the urinary protein:creatinine ratio (LMW phthalates, - 9.53% change; 95% CI: - 14.21, - 4.21; p = 0.001), and in most cases, a positive relationship with eGFR (LMW phthalates, a 3.46 unit increase in eGFR, 95% CI: 1.85, 5.07; p < 0.001). LIMITATIONS: Lack of longitudinal data, limited assessment of diet and nutritional status. CONCLUSION: In the study cohort, children with CKD did not have increased exposure to BPA and phthalates. Longitudinal studies with repeated measures are likely to be more informative about the possible health effects of prolonged exposure to BPA and phthalates in pediatric patients with CKD.

Exposure to bisphenols and phthalates and association with oxidant stress, insulin resistance, and endothelial dysfunction in children.

BACKGROUND: The health effects of bisphenol A (BPA) and di-(2-ethylhexyl) phthalate (DEHP) have been studied extensively in children. The impact of other chemicals in these two classes has not been investigated as fully. METHODS: We conducted a cross-sectional pilot study of 10-13 y old healthy children. We assessed descriptive, univariable, and multivariable associations of urinary metabolites of bisphenols and phthalates with oxidant stress, insulin resistance, body mass, and endothelial dysfunction. Possible associations with brachial artery distensibility, pulse wave velocity (markers of vascular stiffness), and serum endothelial cell-derived microparticle levels were also assessed. RESULTS: We enrolled 41 participants, 12.1 ± 1.0 y, most of whom were Mexican Americans (42%) or other Hispanics (34%). Increased BPA levels were associated with increased levels of F2-isoprostane (ng/ml) (P = 0.02), with a similar trend for DEHP metabolites. Each log unit increase of high molecular weight (HMW) phthalate metabolites was associated with a 0.550 increase in Homeostatic Model Assessment of insulin resistance (HOMA-IR) units (P = 0.019) and altered circulating levels of activated endothelial cell-derived microparticles (% per ml) (P = 0.026). Bisphenol S (BPS), a replacement for BPA, was associated with increased albumin (mg):creatinine (g) ratio (P = 0.04). Metabolites of HMW phthalates were also associated with decreased brachial artery distensibility (P = 0.047). CONCLUSION: Exposure to bisphenols and phthalates, including a BPA replacement, is associated with increased oxidant stress, insulin resistance, albuminuria, as well as disturbances in vascular function in healthy children.

Efficacy of galactose and adalimumab in patients with resistant focal segmental glomerulosclerosis: report of the font clinical trial group.

BACKGROUND: Patients with resistant focal segmental glomerulosclerosis (FSGS) who are unresponsive to corticosteroids and other immunosuppressive agents are at very high risk of progression to end stage kidney disease. In the absence of curative treatment, current therapy centers on renoprotective interventions that reduce proteinuria and fibrosis. The FONT (Novel Therapies for Resistant FSGS) Phase II clinical trial (NCT00814255, Registration date December 22, 2008) was designed to assess the efficacy of adalimumab and galactose compared to standard medical therapy which was comprised of lisinopril, losartan, and atorvastatin. METHODS: Key eligibility criteria were biopsy confirmed primary FSGS or documentation of a causative genetic mutation, urine protein:creatinine ratio >1.0 g/g, and estimated glomerular filtration rate (eGFR) >40 ml/min/1.73 m(2). The experimental treatments - adalimumab, galactose, standard medical therapy-- were administered for 26 weeks. The primary endpoint was a 50 % reduction in proteinuria with stable eGFR. RESULTS: Thirty-two subjects were screened and 21 were assigned to one of the three study arms. While none of the adalimumab-treated subjects achieved the primary outcome, 2 subjects in the galactose and 2 in the standard medical therapy arm had a 50 % reduction in proteinuria without a decline in eGFR. The proteinuria response did not correlate with serial changes in the serum glomerular permeability activity measured by the Palb assay or soluble urokinase plasminogen activator receptor (suPAR). There were no serious adverse effects related to treatments in the study. CONCLUSIONS: Recruitment into this trial that addressed patients with resistant FSGS fell short of the enrollment goal. Our findings suggest that future studies of novel therapies for rare glomerular diseases such as FSGS may benefit from enrollment of patients earlier in the course of their disease. In addition, better identification of patients who are likely to respond to a new treatment based on biomarkers suggesting involvement of the disease pathway targeted by the experimental agent may reduce the required sample size and increase the likelihood of a favorable outcome.

Urinary exosomal Wilms' tumor-1 as a potential biomarker for podocyte injury.

Renal Wilms' tumor-1 (WT-1) staining is used to detect podocyte loss in kidney biopsies. We aimed to determine if urinary exosomal WT-1 could serve as a noninvasive biomarker of podocyte injury. We examined WT-1 by Western blot in a human podocyte-like cell line, a mouse model of podocyte injury, and human subjects with podocyte disorders. WT-1 was detected in exosomal fraction of the conditioned media from podocytes and increased 48 h after hTGF-ß1 stimulation. Cellular WT-1 decreased in podocytes following hTGF-ß1 incubation. In mice with induced podocyte injury, urinary exosomal WT-1 was detected 1 wk earlier than albuminuria and also tracked the effects of angiotensin receptor blocker (ARB) treatment. In addition, urinary exosomal WT-1 levels at 1 wk post-injury correlated with the severity of glomerular injury at 3 wk later. In human subjects, urinary exosomal WT-1 was significantly increased in focal segmental glomerulosclerosis (FSGS) patients compared with healthy volunteers or steroid-sensitive nephrotic syndrome (SSNS) patients. Urinary exosomal WT-1 was also significantly decreased in patients in remission for either FSGS or SSNS or following steroid treatment in six SSNS subjects. We conclude that urinary exosomal WT-1 is a promising noninvasive biomarker with apparent podocyte specificity that can detect early progression and treatment-induced regression of podocyte injury in FSGS or SSNS. These results warrant longitudinal, prospective studies in a large cohort with a range of podocyte diseases.

Developing an Edema Clinician-Reported Outcome Measure for Nephrotic Syndrome.

Introduction: Edema is a common manifestation of proteinuric kidney diseases, but there is no consensus approach for reliably evaluating edema. The objective of this study was to develop an edema clinician-reported outcome measure for use in patients with nephrotic syndrome. Methods: A literature review was conducted to assess existing clinician-rated measures of edema. Clinical experts were recruited from internal medicine, nephrology, and pediatric nephrology practices to participate in concept elicitation using semi-structured interviews and cognitive debriefing. Qualitative analysis methods were used to collate expert input and inform measurement development. In addition, training and assessment modules were developed using an iterative process that also utilized expert input and cognitive debriefing to ensure interrater reliability. Results: While several clinician-rated measures of edema have been proposed, our literature review did not identify any studies to support the reliability or validity of these measures. Fourteen clinician experts participated in the concept elicitation interviews, and twelve participated in cognitive debriefing. A clinician-reported outcome measure for edema was developed. The measure assesses edema severity in multiple individual body parts. An online training module and assessment tool were generated and refined using additional clinician input and investigative team expertise. Conclusion: The Edema ClinRO (V1) measure is developed specifically to measure edema in nephrotic syndrome. The tool assesses edema across multiple body parts, and it includes a training module to ensure standardized administration across raters. Future examination of this measure is ongoing to establish its reliability and validity.

Clinical trial of focal segmental glomerulosclerosis in children and young adults.

This NIH-funded multicenter randomized study of focal segmental glomerulosclerosis (FSGS) treatment compared the efficacy of a 12-month course of cyclosporine to a combination of oral pulse dexamethasone and mycophenolate mofetil in children and adults with steroid-resistant primary FSGS. Of the 192 patients enrolled, 138 were randomized to cyclosporine (72) or to mycophenolate/dexamethasone (66). The primary analysis compared the levels of an ordinal variable measuring remission during the first year. The odds ratio (0.59) for achieving at least a partial remission with mycophenolate/dexamethasone compared to cyclosporine was not significant. Partial or complete remission was achieved in 22 mycophenolate/dexamethasone- and 33 cyclosporine-treated patients at 12 months. The main secondary outcome, preservation of remission for 26 weeks following cessation of treatment, was not significantly different between these two therapies. During the entire 78 weeks of study, 8 patients treated with cyclosporine and 7 with mycophenolate/dexamethasone died or developed kidney failure. Thus, our study did not find a difference in rates of proteinuria remission following 12 months of cyclosporine compared to mycophenolate/dexamethasone in patients with steroid-resistant FSGS. However, the small sample size might have prevented detection of a moderate treatment effect.

Novel therapies for resistant focal segmental glomerulosclerosis (FONT) phase II clinical trial: study design.

BACKGROUND: The lack of adequate randomized clinical trials (RCT) has hindered identification of new therapies that are safe and effective for patients with primary focal segmental glomerulosclerosis (FSGS), especially in patients who fail to respond to corticosteroids and immunosuppressive therapies. Recent basic science advances have led to development of alternative treatments that specifically target aberrant pathways of fibrosis which are relevant to disease progression in FSGS. There is a need for a flexible Phase II study design which will test such novel antifibrotic strategies in order to identify agents suitable for phase III testing. METHODS/DESIGN: The Novel Therapies for Resistant Focal Segmental Glomerulosclerosis (FONT) project is a multicenter Phase I/II RCT designed to investigate the potential efficacy of novel therapies for resistant FSGS. Adalimumab and galactose will be evaluated against conservative therapy consisting of the combination of lisinopril, losartan and atorvastatin. The sample size is defined to assure that if one of the treatments has a superior response rate compared to that of the other treatments, it will be selected with high probability for further evaluation. Comparison of primary and secondary endpoints in each study arm will enable a choice to be made of which treatments are worthy of further study in future Phase III RCT. DISCUSSION: This report highlights the key features of the FONT II RCT including the two-step outcome analysis that will expedite achievement of the study objectives. The proposed phase II study design will help to identify promising agents for further testing while excluding ineffective agents. This staged approach can help to prevent large expenditures on unworthy therapeutic agents in the management of serious but rare kidney diseases.