Poorichaya Somparn

Pharmacokinetics of mycophenolic acid in severe lupus nephritis

Mycophenolic acid (MPA) is an effective treatment for active lupus nephritis despite its variable efficacy in different ethnic groups. Here we tested whether pharmacokinetic monitoring may help to optimize dosing of MPA in an Asian population. Patients with biopsy-proven class III or IV lupus nephritis (ISN/RPS category) were treated with mycophenolate mofetil or enteric-coated mycophenolate sodium. One month after initiating treatment we measured plasma MPA levels in eight samples taken over a 12-h period after drug administration. The mean area under the time-dependent curve for MPA of responding patients was significantly higher than those not responding. Successful treatment was seen in patients with areas >45 mg h/l. The dosage of the drug was not related to MPA pharmacokinetics. In the mycophenolate mofetil group, however, MPA-area under the curve was positively, and significantly, correlated with trough or 1 h after dose concentrations and associated with a therapeutic response. Thus, our study shows that MPA pharmacokinetics were positively correlated with therapeutic responses of mycophenolate, suggesting that controlling the concentrations may improve its therapeutic efficacy in lupus nephritis. As the absorption and pharmacokinetic peak of enteric-coated tablets is slower, it is important to take different formulations into account when determining optimal MPA concentrations.

APRIL, a proliferation-inducing ligand, as a potential marker of lupus nephritis.

IntroductionBLyS and APRIL are cytokines from the tumor necrosis factor family which play an important role in systemic lupus erythematosus (SLE). Previous works suggested an association between both molecules and SLE disease activity although their correlation with lupus nephritis is not known. We therefore assessed serum BLyS and APRIL in active lupus nephritis patients.MethodsSerum samples from active lupus nephritis and at 6 months post-treatment were obtained. Serum levels of BLyS and APRIL (n = 47) as well as renal mRNA expression were measured. Serum levels of both molecules and clinical data (n = 27) were available at 6 months follow-up. All biopsy-proven lupus nephritis patients were treated with similar immunosuppressive drugs.ResultsSerum levels of APRIL were associated with proteinuria (Rs = 0.44, P value < 0.01) and degree of histological activity (Rs = 0.34; P value < 0.05) whereas BLyS levels were associated with complement levels (Rs = 0.46; P value < 0.01) and dosage of immunosuppressant. Interestingly, serum APRIL as well as its intrarenal mRNA levels were associated with resistance to treatment. From the receiver operating characteristic (ROC) analysis, high levels (> 4 ng/mL) of serum APRIL predicted treatment failure with a positive predictive value of 93 percent.ConclusionAPRIL could be a potential biomarker for predicting difficult-to-treat cases of lupus nephritis.

Exosomes in urine biomarker discovery.

Nanovesicles present in urine the so-called urinary exosomes have been found to be secreted by every epithelial cell type lining the urinary tract system in human. Urinary exosomes are an appealing source for biomarker discovery as they contain molecular constituents of their cell of origin, including proteins and genetic materials, and they can be isolated in a non-invasive manner. Following the discovery of urinary exosomes in 2004, many studies have been performed using urinary exosomes as a starting material to identify biomarkers in various renal, urogenital, and systemic diseases. Here, we describe the discovery of urinary exosomes and address the issues on the collection, isolation, and normalization of urinary exosomes as well as delineate the systems biology approach to biomarker discovery using urinary exosomes.

Urinary proteomics revealed prostaglandin H2D-isomerase, not Zn-α2-glycoprotein, as a biomarker for active lupus nephritis

Although renal histopathology is the gold standard for the diagnosis and prognosis of lupus nephritis (LN), the invasiveness of renal biopsy warrants the discovery of novel non-invasive diagnostic and prognostic biomarkers. In the present study, urine samples from 10 LN patients (5 active and 5 inactive) were analyzed by two-dimensional gel electrophoresis (2-DE) to screen for potential biomarkers of active LN. Quantitative analysis and statistics revealed 16 protein spots whose levels significantly differed between groups. These proteins were successfully identified by electrospray ionization quadrupole time-of-flight tandem mass spectrometry (ESI-Q-TOF MS/MS). Among these potential candidates, differential levels of urinary Zn-α2-glycoprotein (ZA2G) and prostaglandin H(2)D-isomerase (PGDS) were further validated by enzyme-linked immunosorbent assay (ELISA) in another independent group of 78 subjects, including 30 active LN, 26 inactive LN, 14 non-LN glomerular diseases, and 8 healthy normal individuals. Whereas ZA2G levels were elevated in urine of patients with active LN and non-LN glomerular diseases, PGDS was elevated only in the urine of the active LN group. Urinary PGDS, not ZA2G, may serve as a biomarker for active LN and upon validation in larger studies, may become the non-invasive test to evaluate the disease activity in future management of LN.

Therapeutic drug monitoring of mycophenolate mofetil for the treatment of severely active lupus nephritis

Background Plasma mycophenolic acid (MPA) concentrations may predict therapeutic response in active lupus nephritis (LN). We determined the efficacy and safety of a concentration-controlled MPA regime in the treatment of severely active LN. Methods In this prospective study, 19 biopsy-proven class III/IV LN patients were treated with mycophenolate mofetil (MMF) for 48 weeks. The MMF dosage was based on maximal plasma MPA concentration at 1-hour post dose (MPA-C1). All patients had plasma MPA-C1 levels monitored weekly until achieving the targeted level of >13 mg/L. A low-dose steroid protocol was started at 0.5 mg/kg/day and rapidly tapered to 5 mg/day. Therapeutic response was evaluated at week 24 and week 48. MPA area-under-the curve (MPA-AUC0–12h) was measured at week 12 to verify the optimum dosage. Results No death or end-stage kidney disease occurred in this study. Seventeen patients (89%) responded to therapy at week 24 with four (21%) patients having complete response. There was no renal relapse at week 48 and four more patients had converted from partial response to complete response. Seventy eight percent of patients achieved the recommended MPA-AUC0–12h level. No association between plasma MPA concentrations and adverse reactions or infections was found. Conclusions MPA-C1 may be a practical monitoring of MPA levels in patients with LN. It is convenient to monitor and may facilitate an optimum estimate of MPA exposure.

Deubiquitylation of Protein Cargo Is Not an Essential Step in Exosome Formation

Exosomes, derived from multivesicular bodies (MVBs), contain proteins and genetic materials from their cell of origin and are secreted from various cells types, including kidney epithelial cells. In general, it is thought that protein cargo is ubiquitylated but that ubiquitin is cleaved by specific deubiquitylases during the process of cargo incorporation into MVBs. Here, we provide direct evidence that, in vivo, deubiquitylation is not essential. Ubiquitin was detected within human MVBs and urinary exosomes by electron microscopy. Of the >6000 proteins identified in human urinary exosomes was mass spectrometry, 15% were ubiquitylated with various topologies (Lys63>Lys48> Lys11>Lys6>Lys29>Lys33>Lys27). A significant preference for basic amino acids upstream of ubiquitylation sites suggests specific ubiquitylation motifs. The current studies demonstrate that, in vivo, deubiquitylation of proteins is not necessary for their incorporation into MVBs and highlight that urinary exosomes are an enriched source for studying ubiquitin modifications in physiological or disease states.

Serum miRNA-122 in acute liver injury induced by kidney injury and sepsis in CD-1 mouse models.

miRNA‐122 (miR‐122) is a new, interesting liver injury biomarker but little is known about its effects when there is an indirect acute liver injury.

Population pharmacokinetics of mycophenolate mofetil in Thai lupus nephritis patients.

OBJECTIVE Mycophenolic acid (MPA) has become the first-line drug therapy for proliferative lupus nephritis, a common and serious complication of systemic lupus erythematosus. Although a sufficient MPA exposure is required, a high interindividual variability in the pharmacokinetics of MPA has been observed. The knowledge of MPA pharmacokinetics in lupus nephritis patients is limited, especially in Asian patients. This study aimed to develop a population pharmacokinetic model for MPA and determine the population pharmacokinetic parameters and their interindividual variability in Thai patients with lupus nephritis. METHODS A total of 112 MPA plasma concentrations from 14 adult lupus nephritis patients (International Society of Nephrology/Renal Pathology Society Class III/IV) receiving mycophenolate mofetil were included in this study. The data was analyzed using NONMEM. The model evaluation was performed by the bootstrap approach and visual predictive check. RESULTS A two-compartment model with a lag time best described the data. The estimated mean apparent clearance (CL/F) was 14.5 l/h with an interindividual variability of 45.2%. The estimated mean CL/F was found to be lower than the values previously reported. The estimated mean apparent volume of the central compartment was 12.2 l with an interindividual variability of 166%. None of the covariates were found to significantly influence MPA pharmacokinetics. CONCLUSION In this study, a population pharmacokinetic model of MPA in severe lupus nephritis patients was successfully developed. The mean pharmacokinetic parameters were estimated and a high interindividual variability of MPA in this population was observed. This provides evidence to show that individualizing dosage regimens in this population is crucial. The model developed in this study could be used to obtain initial information for MPA dose adjustments in Thai and Asian patients with lupus nephritis. Further studies are required to validate the results and clarify the influence of covariates on MMF pharmacokinetics.

Autophagic degradation of aquaporin-2 is an early event in hypokalemia-induced nephrogenic diabetes insipidus

Hypokalemia (low serum potassium level) is a common electrolyte imbalance that can cause a defect in urinary concentrating ability, i.e., nephrogenic diabetes insipidus (NDI), but the molecular mechanism is unknown. We employed proteomic analysis of inner medullary collecting ducts (IMCD) from rats fed with a potassium-free diet for 1 day. IMCD protein quantification was performed by mass spectrometry using a label-free methodology. A total of 131 proteins, including the water channel AQP2, exhibited significant changes in abundance, most of which were decreased. Bioinformatic analysis revealed that many of the down-regulated proteins were associated with the biological processes of generation of precursor metabolites and energy, actin cytoskeleton organization, and cell-cell adhesion. Targeted LC-MS/MS and immunoblotting studies further confirmed the down regulation of 18 selected proteins. Electron microscopy showed autophagosomes/autophagolysosomes in the IMCD cells of rats deprived of potassium for only 1 day. An increased number of autophagosomes was also confirmed by immunofluorescence, demonstrating co-localization of LC3 and Lamp1 with AQP2 and several other down-regulated proteins in IMCD cells. AQP2 was also detected in autophagosomes in IMCD cells of potassium-deprived rats by immunogold electron microscopy. Thus, enhanced autophagic degradation of proteins, most notably including AQP2, is an early event in hypokalemia-induced NDI.

Gastrointestinal Leakage Detected by Serum (1→3)-β-d-glucan in Mouse Models and a Pilot Study in Patients with Sepsis

ABSTRACT Gastrointestinal (GI) leakage is believed to exacerbate sepsis and new, validated markers of GI barrier performance might benefit clinical decision-making. Serum (1→3)-&bgr;-D-glucan (BG) was evaluated as a potential GI leakage marker. Serum BG was tested in several mouse models of GI leakage, including dextran sulfate solution (DSS) administration, endotoxin (LPS) injection, and cecal ligation and puncture sepsis (CLP). Serum BG titer was also evaluated in patients with sepsis and septic shock, for comparison. With 0.75% DSS administration, BG increased only after oral administration of heat-killed C. albicans, but increased spontaneously with 1.5% DSS. In the LPS and CLP models, BG increased as early as 1 h and at 12 h after LPS administration and surgery, respectively. GI leakage was confirmed by orthogonal validation methods including FITC-dextran oral administration in the DSS, LPS, and CLP models and, in the DSS model, with urine sucralose after oral administration and serum endotoxemia. IL-6 increased in parallel with serum BG. Serum BG or IL-6, at 18 h, anticipated sepsis mortality in the CLP model. Analysis of serum BG from patients with febrile neutropenic sepsis (N = 49) and febrile non-neutropenic sepsis (N = 39) demonstrated BG elevation. Patients with bacterial septic shock had serum BG titers similar to levels observed in invasive fungal disease, regardless of febrile neutropenia. Serum BG was lower in less severe cases of bacterial sepsis. Elevated serum IL-6 was associated with GI leakage and elevated serum BG. Serum BG may have potential as a sepsis/septic shock biomarker and further study in this context is warranted.