Noceti Ofelia M, Woillard Jean-Baptiste, Boumediene Ahmed, Esperón Patricia, Taupin Jean-Luc, Gerona Solange, Valverde Marcelo, Touriño Cristina, Marquet Pierre
INSERM U850 and Liver Diseases Department, National Center for Liver Transplantation, Hospital Central de las Fuerzas Armadas, Montevideo, Uruguay; Molecular Biology Unit, Clinical Biochemistry Department, School of Chemistry, and.
INSERM U850 and.
Clin Chem. 2014 Oct;60(10):1336-45. doi: 10.1373/clinchem.2014.223511. Epub 2014 Aug 20.
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.
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.
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.
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.
尽管治疗药物监测改善了免疫抑制药物的临床应用，但疗效和毒性在患者间仍存在差异，而药效学监测可能有助于减少这种差异。为了选择他克莫司药效学的最佳生物标志物，我们探究了信号转导的强度和变异性以及钙调神经磷酸酶途径中多态性的影响。
将35名健康志愿者的外周血单个核细胞与他克莫司（0.1 - 50 ng/mL）一起孵育并进行体外刺激。通过流式细胞术测量CD4(+)和CD8(+) T细胞中NFAT1（活化T细胞核因子1）向细胞核的转位抑制以及白细胞介素-2的细胞内表达，以及CD3(+)细胞表面活化标志物CD25。我们对亲免蛋白和钙调神经磷酸酶亚基的启动子区域进行测序，并通过等位基因鉴别分析对钙调神经磷酸酶途径基因中选定的单核苷酸多态性进行表征。
所有反应均紧密拟合I/Imax S形模型。所有生物标志物的I0和IC50均存在较大的个体间变异性（n = 30）。此外，在编码亲环蛋白A、钙调神经磷酸酶催化亚基α同工酶和CD25的基因多态性与他克莫司药效学参数之间发现了强烈且具有统计学意义的关联。
本研究证明了钙调神经磷酸酶途径信号转导的一致性和较大的个体间变异性，以及钙调神经磷酸酶级联反应中的药物遗传学多态性对该途径的生理活性和他克莫司药效学的强烈影响。
