A plasmatic score using a miRNA signature and CXCL-10 for accurate prediction and diagnosis of liver allograft rejection.

Introduction: The use of noninvasive biomarkers may avoid the need for liver biopsy (LB) and could guide immunosuppression adjustment in liver transplantation (LT). The aims of this study were: to confirm the predictive and diagnostic capacity of plasmatic expression of miR-155-5p, miR-181a-5p, miR-122-5p and CXCL-10 for assessing T-cell mediated rejection (TCMR) risk; to develop a score based on a panel of noninvasive biomarkers to predict graft rejection risk and to validate this score in a separate cohort. Methods: A prospective, observational study was conducted with a cohort of 79 patients followed during the first year after LT. Plasma samples were collected at predetermined time points for the analysis of miRNAs and the CXCL-10. Patients with LFTs abnormalities were submitted to a LB to rule out rejection, assessing previous and concurrent expression of the biomarkers to evaluate their predictive and diagnostic ability. Information from 86 patients included in a previous study was collected and used as a validation cohort. Results: Twenty-four rejection episodes were diagnosed in 22 patients. Plasmatic CXCL-10 concentration and the expression of the three miRNAs were significantly elevated prior to and at the moment of the diagnosis of rejection. We developed a logistic model for rejection prediction and diagnosis, which included CXCL-10, miR-155-5p and miR-181a-5p. The area under the ROC curve (AUROC) for rejection prediction was 0.975 (79.6% sensitivity, 99.1% specificity, 90,7% PPV; 97.7% NPV; 97.1% correctly classified) and 0.99 for diagnosis (87.5% sensitivity, 99.5% specificity, 91.3% PPV; 99.3% NPV; 98.9% correctly classified). In the validation cohort (n=86; 14 rejections), the same cut-off points were used obtaining AUROCs for rejection prediction and diagnosis of 0.89 and 0.92 respectively. In patients with graft dysfunction in both cohorts the score could discriminate those with rejection regarding other causes with an AUROC of 0.98 (97.3% sensitivity, 94.1%specificity). Conclusion: These results suggest that the clinical implementation of the monitoring of this noninvasive plasmatic score may allow the prediction and diagnosis of rejection and identify patients with graft dysfunction due to rejection, helping with a more efficient guide for immunosuppressive therapy adjustment. This finding warrants the development of prospective biomarker-guided clinical trials.

Dried Blood Spot (DBS) as a useful tool to improve clozapine, aripiprazole and paliperidone treatment: From adherence to efficiency.

INTRODUCTION: Therapeutic Drug Monitoring (TDM) of antipsychotics in schizophrenia is a powerful tool that allows tailoring the treatment in an individualized approach. Our goals are to develop and validate a Dried Blood Spot (DBS) method for monitoring some commonly used antipsychotics (aripiprazole, clozapine, and paliperidone) and to evaluate its usefulness as a compliance biomarker, as well as in drug-dose adjustment to personalize the antipsychotic treatment to improve its efficacy and safety. METHODS: 31 first-psychotic episode (FEP) and schizophrenia patients were included; 5 refer to naïve FEP who started antipsychotic treatment; 26, to patients with more than one episode and under antipsychotic treatment: aripiprazole (7 cases), clozapine (17), paliperidone (11). For DBS sample collection, 25µl of capillary blood were placed in the spot of a FTA™DMPK-C-card. After completely dryness, antipsychotics were extracted and analyzed by a validated UHPLC-MS/MS-method. DBS antipsychotic results were compared with those obtained in venous blood/plasma. RESULTS: Aripiprazole, paliperidone and clozapine showed from good to excellent correlations between concentrations in venous blood and DBS capillary blood (r2, from 0.500 to 0.721). The correlation between conventional plasma and DBS concentrations for paliperidone, aripiprazole, clozapine, and their metabolites were moderate, suggesting that optimal drug target concentrations should be established for DBS. CONCLUSIONS: In this study, for aripiprazole, dehydroaripiprazole, paliperidone, clozapine and desmethylclozapine, DBS has provided good analytical performance for TDM. Thus, DBS sampling can offer a great alternative over conventional sampling for plasma measurement. The assay provides good analytical performances for TDM and clinical research applicability, suggesting that DBS is a promising clinical application in TDM in psychiatry.

Dried Blood Spot (DBS) as a useful tool to improve clozapine, aripiprazole and paliperidone treatment: From adherence to efficiency / Análisis de sangre seca (Dried Blood Spot [DBS]) como herramienta útil para mejorar el tratamiento con clozapina, aripiprazol y paliperidona: de la adherencia a la eficacia

Introduction: Therapeutic Drug Monitoring (TDM) of antipsychotics in schizophrenia is a powerful tool that allows tailoring the treatment in an individualized approach. Our goals are to develop and validate a Dried Blood Spot (DBS) method for monitoring some commonly used antipsychotics (aripiprazole, clozapine, and paliperidone) and to evaluate its usefulness as a compliance biomarker, as well as in drug-dose adjustment to personalize the antipsychotic treatment to improve its efficacy and safety. Methods: 31 first-psychotic episode (FEP) and schizophrenia patients were included; 5 refer to naïve FEP who started antipsychotic treatment; 26, to patients with more than one episode and under antipsychotic treatment: aripiprazole (7 cases), clozapine (17), paliperidone (11). For DBS sample collection, 25μl of capillary blood were placed in the spot of a FTA™DMPK-C-card. After completely dryness, antipsychotics were extracted and analyzed by a validated UHPLC-MS/MS-method. DBS antipsychotic results were compared with those obtained in venous blood/plasma. Results: Aripiprazole, paliperidone and clozapine showed from good to excellent correlations between concentrations in venous blood and DBS capillary blood (r2, from 0.500 to 0.721). The correlation between conventional plasma and DBS concentrations for paliperidone, aripiprazole, clozapine, and their metabolites were moderate, suggesting that optimal drug target concentrations should be established for DBS. (AU)

Introducción: La monitorización terapéutica de medicamentos (Therapeutic Drug Monitoring [TDM]) de los antipsicóticos en la esquizofrenia es una potente herramienta que permite adaptar el tratamiento de forma individualizada y personalizada. Los objetivos del presente estudio son desarrollar y validar un método de análisis en sangre seca (Dried Blood Spot [DBS]) para la monitorización de algunos antipsicóticos de uso común (aripiprazol, clozapina y paliperidona) y evaluar su utilidad como biomarcador de cumplimiento y adherencia terapéutica, así como en el ajuste de la dosis del fármaco para personalizar el tratamiento antipsicótico para mejorar su eficacia y su seguridad. Metodología: Se incluyeron 31 pacientes con un primer episodio psicótico (PEP) o un diagnóstico de esquizofrenia; 5 eran PEP que iniciaron tratamiento antipsicótico, y 26 eran pacientes con más de un episodio y que seguían tratamiento con antipsicóticos: aripiprazol (7 casos), clozapina (17), paliperidona (11). Para la recogida de muestras de DBS se colocaron 25μl de sangre capilar en el punto de una tarjeta FTATMDMPK-C. Tras secarse completamente, se extrajeron los antipsicóticos y se analizaron mediante un método UHPLC-MS/MS validado. Los resultados de los antipsicóticos según la DBS se compararon con los obtenidos en sangre venosa/plasma. Resultados: El aripiprazol, la paliperidona y la clozapina mostraron de buenas a excelentes correlaciones entre las concentraciones en sangre venosa y en sangre capilar del DBS (r2, de 0,500 a 0,721). La correlación entre las concentraciones plasmáticas convencionales y las del DBS para la paliperidona, el aripiprazol, la clozapina y sus metabolitos fue moderada, lo que sugiere que se deberían definir y establecer concentraciones óptimas del fármaco para el DBS. (AU)

Early prognostic performance of miR155-5p monitoring for the risk of rejection: Logistic regression with a population pharmacokinetic approach in adult kidney transplant patients.

Previous results from our group and others have shown that urinary pellet expression of miR155-5p and urinary CXCL-10 production could play a key role in the prognosis and diagnosis of acute rejection (AR) in kidney transplantation patients. Here, a logistic regression model was developed using NONMEM to quantify the relationships of miR155-5p urinary expression, CXCL-10 urinary concentration and tacrolimus and mycophenolic acid (MPA) exposure with the probability of AR in adult kidney transplant patients during the early post-transplant period. Owing to the contribution of therapeutic drug monitoring to achieving target exposure, neither tacrolimus nor MPA cumulative exposure was identified as a predictor of AR in the studied population. Even though CXCL-10 urinary concentration showed a trend, its effect on AR was not significant. In contrast, urinary miR155-5p expression was prognostic of clinical outcome. Monitoring miR155-5p urinary pellet expression together with immunosuppressive drug exposure could be very useful during routine clinical practice to identify patients with a potential high risk of rejection at the early stages of the post-transplant period. This early risk assessment would allow for the optimization of treatment and improved prevention of AR.

Synthetic oligodeoxynucleotides induce MAP kinases activation in murine TIB-73 hepatocytes.

AIMS: In this work we aimed to investigate the expression of TLR9 protein in the murine hepatocyte cell line TIB-73, compared to macrophage-like J774 cells, by Western blot analysis, and the role played by ERK 1/2 MAP kinase in the intracellular signals triggered by stimulation with CpG and non-CpG phosphodiester-ODN, and their more stable phosphorothioate-modified analogues. RESULTS: TIB-73 hepatocytes express TLR9 protein. CpG and non-CpG ODN stimulation activated ERK 1/2 MAPK signal pathway in both hepatocytes and J774 murine macrophages. As expected, their phosphorothioate-CpG and non-CpG ODN analogues induced higher levels of ERK1/2 phosphorylation in TIB-73 cells, even higher than that induced in J774 cells under the same conditions. Phosphorylation of ERK 1/2 induced by synthetic ODN is dose-response dependent, being maximal at 100 microg/ml. Pretreatment of hepatocytes with an inhibitor of MEK-1 abrogated phosphorylation of ERK1/2 kinase. CONCLUSIONS: TIB-73 hepatocytes constitutively express TLR9 and respond to synthetic ODN stimulation through a high ERK1/2 phosphorylation independent of CpG motifs. Slight differences were found on ERK1/2 activation when using phosphorothioate versus phosphodiester oligonucleotides.