Free mycophenolic acid should be monitored in renal transplant recipients with hypoalbuminemia

The current approach for therapeutic drug monitoring in renal transplant recipients receiving mycophenolate mofetil (MMF) is measurement of total mycophenolic acid (MPA) concentration. Because MPA is highly bound, during hypoalbuminemia the total concentration no longer reflects the free (pharmacologically active) concentration. The authors investigated what degree of hypoalbuminemia causes a significant change in protein binding and thus percentage free MPA. Forty-two renal transplant recipients were recruited for the study. Free and total concentrations of MPA (predose, and 1, 3, and 6 hours post-MMF dose samples) and plasma albumin concentrations were determined on day 5 posttransplantation. Six-hour area under the concentration-time curve (AUC(0-6)) values were calculated for free and total MPA, and percentage free MPA was determined for each patient. The authors found a significant relationship between low albumin concentrations and increased percentage free MPA (Spearman correlation = -0.54, P < 0.0001). Receiver operating characteristic (ROC) curve analysis was performed on the albumin versus percentage free MPA data. The cutoff value of albumin determined from the ROC analysis that differentiated normal from elevated percentage free MPA (defined as greater than or equal to3%) in this patient population was 31 g/L. At this cutoff value albumin was found to be a good predictor of altered free MPA percentage, with a sensitivity and specificity of 0.75 and 0.80, respectively, and an area under the ROC curve of 0.79. To rationalize MMF dosing regimens in hypoalbuminemic patients (plasma albumin less than or equal to 31 g/L), clinicians should consider monitoring the free MPA concentration.

Relationships between sirolimus dosing, concentration and outcomes in renal transplant recipients

Aim To explore relationships between sirolimus dosing, concentration and clinical outcomes. Methods Data were collected from 25 kidney transplant recipients (14 M/11 F), median 278 days after transplantation. Outcomes of interest were white blood cell (WBC) count, platelet (PLT) count, and haematocrit (HCT). A naive pooled data analysis was performed with outcomes dichotomized (Mann-Whitney U-tests). Results Several patients experienced at least one episode when WBC (n = 9), PLT (n = 12), or HCT (n = 21) fell below the lower limits of the normal range. WBC and HCT were significantly lower (P < 0.05) when sirolimus dose was greater than 10 mg day(-1), and sirolimus concentration greater than 12 mu g l(-1). No relationship was shown for PLT and dichotomized sirolimus dose or concentration. Conclusions Given this relationship between sirolimus concentration and effect, linked population pharmacokinetic-pharmacodynamic modelling using data from more renal transplant recipients should now be used to quantify the time course of these relationships to optimize dosing and minimize risk of these adverse outcomes.

Impacto de los Niveles de Tacrolimus en el Rechazo al Injerto y la Tasa de Filtración Glomerular en Pacientes con Trasplante Hepático realizado en la FCI-IC entre 2009 y 2014

Introducción: El tacrolimus es el medicamento de elección para evitar el rechazo al injerto hepático. Su dosis se ajusta a partir de los niveles séricos que se toman periódicamente para asegurar rango terapéutico. Además, niveles elevados se asocian con disfunción renal postrasplante. Sin embargo, no hay consenso frente a los niveles adecuados para pacientes con trasplante hepático. Objetivo: Determinar la relación entre los niveles de tacrolimus y la presencia de rechazo agudo al injerto hepático en pacientes con trasplante hepático realizado en la Fundación Cardioinfantil – Instituto de Cardiología (FCI-IC). Determinar la relación entre los niveles de tacrolimus y la TFG en pacientes con trasplante hepático realizado en la FCI-IC. Métodos: Estudio observacional tipo cohorte histórica en pacientes adultos con trasplante hepático realizado en la FCI-IC entre 2009-2014. Resultados: No se encontró una asociación estadísticamente significativa entre los niveles de tacrolimus y la presencia de rechazo agudo, en sus diferentes definiciones (OR=1,02, p=0,14 y OR=1,01, p=0,29) incluso al ajustar por otras covariables (OR=1,03, p=0,10 y OR=1,02, p=0,25). No fue posible corroborar el diagnóstico con biopsia porque no todos la tenían. Si bien la relación entre los niveles de tacrolimus y la TFG fue estadísticamente significativa (p≤0,001), tiene bajo impacto clínico, pues la TFG disminuyó menos de un punto por cada incremento en 1 ng/ml en los niveles de tacrolimus. Conclusiones: Se necesitan más estudios para establecer la relación entre la exposición a tacrolimus y estos desenlaces para definir si es seguro disminuir su dosis con el fin de reducir los eventos adversos.

Malattie immunomediate del cane e del gatto: aspetti diagnostici e terapeutici

La presente tesi di dottorato affronta alcune delle più comuni malattie immunomediate del cane e del gatto. Il manoscritto è incentrato sugli aspetti diagnostici e terapeutici in corso di: anemia emolitica immunomediata (Immune-mediated hemolytic anemia, IMHA), trombocitopenia immunomediata (Immune-mediated thrombocytopenia, ITP) e poliartrite immunomediata (Immune-mediated polyarthritis, IMP). Il capitolo 1 costituisce un’introduzione all’argomento delle malattie immunologiche; vengono sottolineati alcuni aspetti patogenetici delle singole malattie immunomediate e riassunte le difficoltà diagnostiche e terapeutiche. Il capitolo 2 riporta uno studio riguardante una popolazione di gatti con diagnosi, o sospetto diagnostico, di IMHA che evidenziava una discrepanza tra i test diagnostici per il virus della leucemia felina (Feline Leukemia Virus, FeLV). La positività FeLV al test point of care, non confermata dalla PCR del DNA provirale, lascia spazio a diverse interpretazioni. Il capitolo 3 mostra i dati relativi al confronto tra tre diversi protocolli immunosoppressivi (glucocorticoidi, glucocorticoidi+ciclosporina, glucocorticoidi+micofenolato mofetile) in una popolazione di cani con IMHA non associativa. Il confronto verteva, principalmente, sulla risposta ematologica dei pazienti, che non si è dimostrata differente tra i tre gruppi terapeutici. Il capitolo 4 riporta una revisione della letteratura riguardante l’ITP del cane e del gatto. Si tratta di una malattia eterogenea in cui le manifestazioni cliniche appaiono variabili: alcuni pazienti sono asintomatici, altri presentano dei sanguinamenti spontanei. La mancanza di criteri diagnostici standardizzati, porta il clinico a considerare l’ITP una diagnosi “ad esclusione”. Le strategie terapeutiche non si basano purtroppo su linee guida condivise, pertanto il target della terapia rimane, ad oggi, sconosciuto. Nel capitolo 5 viene posta l’attenzione su alcuni interrogativi diagnostici e terapeutici che riguardano l’IMP del cane e del gatto. La sintomatologia clinica, caratterizzata da zoppia, febbre e riluttanza al movimento, talvolta può essere subdola. Anche in questa malattia, non vi sono evidenze scientifiche circa il regime immunosoppressivo più corretto ed indicato.

Pharmacokinetic/pharmacodynamic relationships of FTY720 in kidney transplant recipients

FTY720 is a new and effective immunosuppressive agent, which produces peripheral blood lymphopenia through a lymphocyte homing effect. We investigated the relationship between the dose of FTY720 or blood concentration (pharmacokinetics, PK) and peripheral lymphopenia (pharmacodynamics, PD) in 23 kidney transplant recipients randomized to receive FTY720 (0.25-2.5 mg/day) or mofetil mycophenolate (2 mg/day) in combination with cyclosporine and steroids. FTY720 dose, blood concentrations and lymphocyte counts were determined weekly before and 4 to 12 weeks after transplantation. The effect of PD was calculated as the absolute lymphocyte count or its reductions. PK/PD modeling was used to find the best-fit model. Mean FTY720 concentrations were 0.36 ± 0.05 (0.25 mg), 0.73 ± 0.12 (0.5 mg), 3.26 ± 0.51 (1 mg), and 7.15 ± 1.41 ng/ml (2.5 mg) between 4 and 12 weeks after transplantation. FTY720 PK was linear with dose (r² = 0.98) and showed low inter- and intra-individual variability. FTY720 produced a dose-dependent increase in mean percent reduction of peripheral lymphocyte counts (38 vs 42 vs 56 vs 77, P < 0.01, respectively). The simple Emax model [E = (Emax * C)/(C + EC50)] was the best-fit PK/PD modeling for FTY720 dose (Emax = 87.8 ± 5.3% and ED50 = 0.48 ± 0.08 mg, r² = 0.94) or concentration (Emax = 78.3 ± 2.9% and EC50 = 0.59 ± 0.09 ng/ml, r² = 0.89) vs effect (% reduction in peripheral lymphocytes). FTY720 PK/PD is dose dependent and follows an Emax model (EC50 = 0.5 mg or 0.6 ng/ml). Using lymphopenia as an FTY720 PD surrogate marker, high % reductions (~80%) in peripheral lymphocytes are required to achieve best efficacy to prevent acute allograft rejection.

Avaliação da tolerabilidade do micofenolato sódico com revestimento entérico versus micofenolato mofetil em receptores de transplante renal

ResumoIntrodução:O micofenolato mofetil (MMF), pró-droga do ácido micofenólico (MPA), é um tratamento imunossupressor eficaz na profilaxia da rejeição aguda, mas associado a eventos adversos gastrointestinais. O micofenolato sódico (MPS) com revestimento entérico foi desenvolvido com a intenção de reduzir tais eventos associados ao MPA.Objetivo:Avaliar a tolerabilidade de EC-MPS e MMF em receptores de transplante renal.Métodos:Estudo retrospectivo, multicêntrico, com pacientes submetidos a transplante renal entre 07/01/2004 e 31/07/2007 em 18 centros brasileiros.Resultados:1380 pacientes incluídos, 702 receberam EC-MPS e 678 receberam MMF. A idade média de 42,3 anos, 60% masculino e 62,5% de etnia caucasiana. A incidência de eventos avaliados no desfecho composto de eficácia não foi diferente entre os grupos ao final de 24 meses de acompanhamento (22,9% para EC-MPS versus 19,9% para MMF, p = 0,203). Os pacientes tratados com EC-MPS apresentaram maior incidência de eventos adversos gastrointestinais comparados com os tratados com MMF (57,7% vs. 52,5%). Infecções virais foram mais frequentes no grupo EC-MPS (38,2%) comparado com MMF (32,6%). Não houve diferença nos valores médios tolerados no final do primeiro (1187 ± 344 mg vs. 1209 ± 426 mg, p = 0,294) e segundo ano (1172,3 ± 347mg vs. 1197,4 ± 430,6 mg, p = 0,241) pós-transplante.Conclusão:Não houve diferença estatística na incidência de rejeição aguda, função tardia e eventos gastrointestinais entre os tratamentos. A dose média tolerada de MPA foi semelhante entre os grupos, mas pacientes tratados com MMF foram submetidos a mais reduções de doses e descontinuações do tratamento.

Clinicopathologic features and outcome of mycophenolate-induced colitis in renal transplant recipients

Reports on the clinical course of mycophenolic acid (MPA)-related colitis in kidney transplant recipients are scarce. This study aimed at assessing MPA-related colitis incidence, risk factors, and progression after kidney transplantation. All kidney transplant patients taking MPA who had colonic biopsies for persistent chronic diarrhea, between 2000 and 2012, at the Kidney Transplantation Unit of Botucatu Medical School Hospital, Brazil, were included. Cytomegalovirus (CMV) immunohistochemistry was performed in all biopsy specimens. Data on presenting symptoms, medications, immunosuppressive drugs, colonoscopic findings, and follow-up were obtained. Of 580 kidney transplant patients on MPA, 34 underwent colonoscopy. Colonoscopic findings were associated with MPA usage in 16 patients. The most frequent histologic patterns were non-specific colitis (31.3%), inflammatory bowel disease (IBD)-like colitis (25%), normal/near normal (18.8%), graft-versus-host disease-like (18.8%), and ischemia-like colitis (12.5%). All patients had persistent acute diarrhea and weight loss. Six of the 16 MPA-related diarrhea patients (37.5%) showed acute dehydration requiring hospitalization. Diarrhea resolved when MPA was switched to sirolimus (50%), discontinued (18.75%), switched to azathioprine (12.5%), or reduced by 50% (18.75%). No graft loss occurred. Four patients died during the study period. Late-onset MPA was more frequent, and no correlation with MPA dose or formulation was found.

Mycophenolate acid inhibits endothelial NAD(P)H oxidase activity and superoxide formation by a Rac1-dependent mechanism

Endothelial dysfunction precedes hypertension and atherosclerosis and predicts cardiac allograft vasculopathy and death in heart transplant recipients. Endothelial overproduction of reactive oxygen species, such as superoxide anions produced by NAD(P)H oxidase, induces endothelial dysfunction. Because immunosuppressive drugs have been associated with increased reactive oxygen species production and endothelial dysfunction, we sought to elucidate the underlying mechanisms. Reactive oxygen species, release of superoxide anions, and NAD(P)H oxidase activity were studied in human umbilical vein endothelial cells and in polymorphonuclear neutrophils. Gp91ds-tat was used to specifically block NAD(P)H oxidase. Transcriptional activation of different subunits of NAD(P)H oxidase was assessed by real-time RT-PCR. Rac1 subunit translocation and activation were studied by membrane fractionation and pull-down assays. Calcineurin inhibitors significantly increased endothelial superoxide anions production because of NAD(P)H oxidase, whereas mycophenolate acid (MPA) blocked it. MPA also attenuated the respiratory burst induced by neutrophil NAD(P)H oxidase. Because transcriptional activation of NAD(P)H oxidase was not affected, but addition of guanosine restored endothelial superoxide anions formation after MPA treatment, we speculate that the inhibitory effect of MPA was mediated by depletion of cellular guanosine triphosphate content. This prevented activation of Rac1 and, thus, of endothelial NAD(P)H oxidase. Because all heart transplant recipients are at risk for cardiac allograft vasculopathy development, these differential effects of immunosuppressants on endothelial oxidative stress should be considered in the choice of immunosuppressive drugs.

Quantification of free mycophenolic acid by high-performance liquid chromatography-atmospheric pressure chemical ionisation tandem mass spectrometry

To facilitate the investigation of free mycophenolic acid concentrations we developed a high-performance liquid chromatography tandem mass spectrometry method using indomethacin as an internal standard. Free drug was isolated from plasma samples (500 mul) using ultrafiltration, The analytes were extracted from the ultrafiltrate (200 mul) using C-18 solid-phase extraction. Detection was by selected reactant monitoring of mycophenolic acid (m/z 318.9-->190.9) and the internal standard (m/z 356.0-->297.1) with an atmospheric pressure chemical ionisation interface. The total chromatographic analysis time was 12 min. The method was found to be linear over the range investigated, 2.5-200 mug/l (r>0.990, n=6). The relative recovery of the method for the control samples studied (7.5, 40.0 and 150 mug/l) ranged from 95 to 104%. The imprecision of the method, expressed in terms of intra- and inter-day coefficients of variation, was

Impact of Renal Dysfunction on Liver Transplantation: a Retrospective Study in 708 Orthotopic Liver Transplant Recipients

Renal dysfunction often complicates the course of orthotopic liver transplant recipients and is associated with increased morbid -mortality. The aims of this study were to determine the incidence of chronic renal disease and its impact on patient survival. Clinical data included age, gender and weight,aetiology of hepatic failure, presence of diabetes,hypertension, hepatitis B and C infection, renal dysfunction pretransplant and immunosuppression. Laboratory data included serum creatinine at days 1, 7, 21, month 6, 12 and yearly. The glomerular filtration rate was determined by Cockcroft-Gault equation. We studied retrospectively from September 1992 to March 2007 708 orthotopic liver transplant recipients. Mean age 44±12.6 years, 64% males, 17% diabetic, 18.8% hypertensive, 19.9% with hepatitis C and 3.8% hepatitis B. Renal dysfunction pretransplant was known in 21.6%. Mean follow-up was 3.6 years. Mean transplant survival 75% at 12 months. 154 patients died. Univariate and multivariate analyses were performed and a p<0.05 was considered significant. Acute kidney injury occurred in 33.2%. Chronic kidney disease stage 3 was observed in 34.3%,stage 4 in 6.2% and stage 5 in 5.1%. At the time of this study, 46.4% were on Cyclosporine A, 44.7% on tacrolimus and 8.9% on sirolimus. Using multivariate analysis, renal dysfunction was correlated with renal dysfunction pre -orthotopic liver transplant (p<0.001), acute kidney injury (p<0.001), haemodialysis development (p<0.001), and inversely correlated with the use of mycophenolate mophetil (p<0.001); mortality was positively correlated with renal dysfunction pretransplant (p=0.03),chronic kidney disease stage 4 (p=0.001), chronic kidney disease stage 5 (p<0.001) and inversely correlated with the use of tacrolimus (p=0.006). In conclusion orthotopic liver transplant recipients are disposed to renal complications that have a negative impact on survival of these patients.

Transplantation in Highly Sensitised Patients Treated with Intravenous Immunoglobulin and Rituximab

Renal transplant in highly sensitised patients is associated with increased morbidity. The aim of this retrospective study was to evaluate the clinical evolution of 30 highly sensitised deceased donor kidney transplants and the influence of different timing of B cell directed treatment and its importance in the outcome of these patients. All recipients had negative complement dependent lymphocytotoxicity cytotoxic T cell crossmatch and no identified anti human leucocyte antigen class I donor specific antibodies. T cell flow crossmatch was performed within 24h of transplantation with serum obtained pretransplant (historic, recent or baseline). Posttransplant flow crossmatch were performed prospectively starting on the 3rd posttransplantation day. The immunosuppressive regime included thymoglobulin, tacrolimus, mycofenolate mofetil and steroids. Positive flow crossmatch occurred in 20/29 patients by the 3rd posttransplantation day, and in 17/27 patients after the 3rd posttransplantation day. All patients were started on intravenous immunoglobulin before transplantation: in nine patients (group A) at 400mg/kg/day for five days; in the remaining 21 patients (group B), as a continued infusion of 2g/kg during 48h. In group A, Rituximab was added only in the presence of antibody mediated rejection; in group B, introduced on the 3rd posttransplantation day whenever a positive flow crossmatch (with serum obtained pre or posttransplant) was reported. Antibody mediated rejection was observed in 44.4% of patients in group A, and 19% of those in group B. Mean follow-up was 12.2±5.5 months. Overall allograft survival was 76.6%, 81% in group B, and 66.6% in group A. At last follow up, mean serum creatinine was 1.3±0.6 mg/dl. Renal transplantation with pretransplant positive flow crossmatch is highly associated with antibody mediated rejection, despite introduction of intravenous immunoglobulin pretransplantation. However high dose intravenous immunoglobulin for 48h plus Rituximab by the 3rd posttransplantation day reduce the incidence of antibody mediated rejection by more than 50% and allowed for allograft survival of 81% at one year, with an excellent renal function.

Leflunomide and Polyomavirus-Associated Nephropathy in Renal Transplant

Polyomavirus nephropathy is a major complication in renal transplantation, associated with renal allograft loss in 14 to 80% of cases. There is no established treatment, although improvement has been reported with a variety of approaches. The authors report two cases of polyomavirus infection in renal allograft recipients. In the first case, a stable patient presented with deterioration of renal function, worsening hypertension and weight gain following removal of ureteral stent placed routinely at the time of surgery. Ultrasound examination and radiology studies revealed hydronephrosis due to ureteral stenosis. A new ureteral stent was placed, but renal function did not improve. Urinary cytology revealed the presence of decoy cells and polyomavirus was detected in blood and urine by qualitative polymerase chain reaction. Renal biopsy findings were consistent with polyomavirus -associated nephropathy. In the second case, leucopaenia was detected in an asymptomatic patient 6 months after transplantation. Mycophenolate mophetil dosage was reduced but renal allograft function deteriorated, and a kidney biopsy revealed polyomavirus -associated nephropathy, also with SV40 positive cells. In both patients immunosuppression with tacrolimus was reduced, mycophenolate mophetil stopped and intravenous immune globulin plus ciprofloxacin started. As renal function continued to deteriorate, therapy with leflunomide (40 mg/day) was associated and maintained during 5 and 3 months respectively. In the first patient, renal function stabilised within one month of starting leflunomide and polymerase chain reaction was negative for polyomavirus after 5 months. A repeated allograft biopsy 6 months later showed no evidence of polyomavirus nephropathy. In the second patient, polyomavirus was undetectable in blood and urine by polymerase chain reaction after 3 months of leflunomide treatment, with no evidence of polyomavirus infection in a repeated biopsy 6 months after beginning treatment.

Provas Epicutâneas sob Imunossupressão Sistémica - Contra-Indicação Absoluta?

Introdução: As provas epicutâneas (PE) são o exame complementar de diagnóstico indicado para avaliação de suspeita de dermite de contacto alérgica. Idealmente, devem ser realizadas sem que o doente se encontre sob imunossupressores. Existem contudo situações clínicas em que tal não é possível, não havendo informação disponível acerca de como realizar e valorizar os resultados das PE nestes doentes. O objectivo do presente trabalho é rever a literatura no que concerne à realização de PE sob imunossupressão iatrogénica. Material e Métodos: Revisão da literatura relevante para o tema publicada até Janeiro de 2015 e indexada à Medline. Resultados: De acordo com o reportado na literatura, foram realizadas PE em 77 doentes sob corticóide sistémico, 78 doentes sob ciclosporina (CyA), 6 sob azatioprina, 10 sob metotrexato (MTX), 4 sob micofenolato de mofetil (MMF), 11 sob fármacos anti-factor de necrose tumoral e 7 sob fármaco anti-IL-12/23. Foram ainda descritos 15 casos de realização de PE sob associação de imunossupressores. Verificaram-se reacções positivas em todos os grupos. Conclusão: O tratamento concomitante com imunossupressores não deve ser uma contra-indicação para realização de PE, estando descritas reacções positivas em doentes sob prednisolona, azatioprina, CyA, MTX, MMF, infliximab, etanercept, adalimumab e ustecinumab. Os resultados negativos ou duvidosos devem, contudo, ser interpretados de forma cautelosa.