Monitoring of NFAT-regulated gene expression in the peripheral blood of allograft recipients: a novel perspective toward individually optimized drug doses of cyclosporine A

Giese, Thomas; Zeier, Martin; Schemmer, Peter; Uhl, Waldemar; Schoels, Margarita; Dengler, Thomas J.; Buechler, Markus W.; Meuer, Stefan

doi:10.1097/01.tp.0000109260.00094.01

Cited by 87 publications

(116 citation statements)

References 17 publications

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“…Previous articles have studied NFAT translocation to the nucleus by means of multispectral image stream flow cytometer (13,14 ), but this is the first time that NFAT1 nuclear translocation was measured by conventional flow cytometry, with overall low to moderate intraassay variability, consistent with previous reports (4,6,7,9,15,16 ), although comparisons are difficult owing to different markers, cell types, and experimental schemes. The other biomarkers were also measured with good intraassay precision, except for IL-2 in CD8 ϩ cells at 5 ng/mL TAC.…”

Section: Cd8supporting

confidence: 72%

Tacrolimus Pharmacodynamics and Pharmacogenetics along the Calcineurin Pathway in Human Lymphocytes

et al. 2014

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BACKGROUND Although therapeutic drug monitoring has improved the clinical use of immunosuppressive drugs, there is still interpatient variability in efficacy and toxicity that pharmacodynamic monitoring may help to reduce. To select the best biomarkers of tacrolimus pharmacodynamics, we explored the strength and variability of signal transduction and the influence of polymorphisms along the calcineurin pathway. METHODS Peripheral blood mononuclear cells from 35 healthy volunteers were incubated with tacrolimus (0.1–50 ng/mL) and stimulated ex vivo. Inhibition of NFAT1 (nuclear factor of activated T cells 1) translocation to the nucleus and intracellular expression of interleukin-2 in CD4+ and CD8+ T cells and the surface activation marker CD25 in CD3+ cells were measured by flow cytometry. We sequenced the promoter regions of immunophilins and calcineurin subunits and characterized selected single nucleotide polymorphisms in the genes of the calcineurin pathway with allelic discrimination assays. RESULTS All responses closely fitted an I/Imax sigmoid model. Large interindividual variability (n = 30) in I0 and IC50 was found for all biomarkers. Moreover, strong and statistically significant associations were found between tacrolimus pharmacodynamic parameters and polymorphisms in the genes coding cyclophilin A, the calcineurin catalytic subunit α isoenzyme, and CD25. CONCLUSIONS This study demonstrates the consistency and large interindividual variability of signal transduction along the calcineurin pathway, as well as the strong influence of pharmacogenetic polymorphisms in the calcineurin cascade on both the physiological activity of this route and tacrolimus pharmacodynamics.

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Section: Cd8supporting

confidence: 72%

Tacrolimus Pharmacodynamics and Pharmacogenetics along the Calcineurin Pathway in Human Lymphocytes

et al. 2014

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“…The most promising approaches for the development of CsA bioassays are based on whole blood stimulation and the subsequent measurement of direct or indirect targets of CsA action in T cells. Among them are assays determining calcineurin activity (27,28), cytokine, in particular IL-2 expression (16,(29)(30)(31)(32), proliferation (11,33), and adenosine triphosphate (ATP) production (34)(35)(36).…”

mentioning

confidence: 99%

Whole blood flow cytometric measurement of NFATc1 and IL‐2 expression to analyze cyclosporine A‐mediated effects in T cells

et al. 2010

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The calcineurin inhibitor Cyclosporine A (CsA) is one of the crucial immunosuppressive drugs given after organ transplantation. The small therapeutic window of CsA generates the dilemma that efficient and toxic drug doses differ only slightly. Moreover, these threshold concentrations differ considerably between individuals; therefore, functional assays are urgently needed. We explored whether the transcription factor NFATc1, a direct as well as indirect target of CsA, can be used as a potential biomarker to determine the individual immunosuppressive activity of CsA. First, in isolated human T cells we showed that flow cytometry is practicable to measure NFATc1, the most abundant NFATc isoform in activated T cells. Second, for whole blood we developed a flow cytometric assay to determine in parallel the inducible transcription factor NFATc1 and the cytokine IL-2 in stimulated T cells. We found that added CsA inhibits both the expression of NFATc1 and IL-2 in T cells of stimulated whole blood samples with IC 50 values of 200 and 150 nM, respectively. The intra-and inter-assay variability was low, and clinical practicability was good. Further experiments have to demonstrate whether the parallel cytometric measurement of NFATc1 and IL-2 in whole blood is a good predictor of individual CsA efficacy and toxicity in CsA-treated patients. '

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“…Dephosphorylated NFAT translocates into the nucleus and switches on certain genes, such as IL-2, INF-γ and GM-CSF, which are required for T-cell activation. The group around Zeier has studied intensively the expression of these genes in recent years as pharmacodynamic biomarkers of the calcineurin inhibitor effect [98] . Using qPCR, the expression of the three genes is measured in stimulated leukocytes by means of the LightCycler instrument (Roche Diagnostics, Mannheim, Germany) before and 2 h or 1.5 h after taking cyclosporine or tacrolimus.…”

Section: Expression Of Nfat-regulated Genesmentioning

confidence: 99%

Biomarkers in blood for individualization of the pharmacotherapy with immunosuppressive drugs after transplantation of solid organs

Wieland

Shipkova

2015

LaboratoriumsMedizin

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Abstract:The success of transplantation medicine is closely associated with the introduction of potent immunosuppressive drugs. Therapy guidance of the calcineurin inhibitors cyclosporine and tacrolimus as well as the mammalian target of rapamycin (mTOR) inhibitors sirolimus and everolimus is achieved by therapeutic drug monitoring (TDM). For other immunosuppressive drugs such as mycophenolic acid, TDM is not established. It has been suggested that a better individualization of pharmacotherapy could be helpful in avoiding over-and underimmunosuppression, which have been associated with poor long-term outcome of grafts and patients. Therefore, a search for biomarkers that could complement TDM, thereby allowing a better individualization of therapy, is ongoing worldwide. Pharmacodynamic biomarkers in transplantation medicine can be either non-drug-specific, aiming at assessing the global effect on the immune system, or drug-specific, aiming at recording the pharmacologic effect on a drug target. Non-specific pharmacodynamic biomarkers can reflect the degree of immune activation by measuring T-or B-cell activation, the development of operational tolerance by monitoring, e.g., regulatory T-cells, or the graft damage by measuring cell-free DNA released by the graft in response to injury. Drug-specific pharmacodynamic biomarkers have been published for mycophenolic acid, calcineurin, and mTOR inhibitors. The field of biomarker research to complement TDM for a better individualization of immunosuppression is still in its infancy, and controlled prospective clinical trials are needed to unravel or dismiss the potential of particular biomarkers or biomarker combinations in various patient cohorts with different grafts.

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Monitoring of NFAT-regulated gene expression in the peripheral blood of allograft recipients: a novel perspective toward individually optimized drug doses of cyclosporine A

Abstract: Quantitative measurement of the inhibition of NFAT-regulated gene expression 2 hr after CsA intake represents a novel approach to assess the biologic effectiveness of CsA therapy and has the potential to enable individualized immunosuppressive regimens.

Cited by 87 publications

References 17 publications

Tacrolimus Pharmacodynamics and Pharmacogenetics along the Calcineurin Pathway in Human Lymphocytes

Tacrolimus Pharmacodynamics and Pharmacogenetics along the Calcineurin Pathway in Human Lymphocytes

Whole blood flow cytometric measurement of NFATc1 and IL‐2 expression to analyze cyclosporine A‐mediated effects in T cells

Biomarkers in blood for individualization of the pharmacotherapy with immunosuppressive drugs after transplantation of solid organs

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