F REIMANN1,2, P GALETTIS1, J JOHNSTONE1, J SCHNEIDER1, P TREVILLIAN1,3, J MARTIN1,3
1School of Medicine and Public Health, University of Newcastle, Newcastle, Australia, 2Calvary Mater Hospital, Newcastle, Australia, 3John Hunter Hospital, Newcastle, Australia
Background: Tacrolimus, ciclosporin, sirolimus and everolimus blood concentrations inform dosing in organ transplant recipients. Quantification with automated immunoassays is common, but liquid chromatography mass spectrometry (LCMS) has higher analyte specificity and allows processing of microsamples.
Aim: To validate a LCMS method for quantification of immunosuppressant drugs in whole blood and compare results of regular samples with volumetric absorptive microsamples (VAMS).
Methods: For LCMS, internal standards are added to blood, cells lysed and centrifuged. Supernatant is injected for chromatographic separation on a Kinetex C18 column prior to quantification in a Shimadzu 8060 mass spectrometer. Analyte sensitivity is optimised at ion source, and detection of at least two product ions for each drug ensures specificity. Quality controls are included in all batches. Patient samples were then analysed by LCMS and Abbott Architect® immunoassay. VAMS of 10 μL, collected on Neoteryx Mitra®, were also processed by LCMS.
Results:The LCMS method is linear over a range of 1.3 to 46 μg/L for tacrolimus, 1.5 to 55 μg/L for both sirolimus and everolimus, and 26 to 1300 μg/L for ciclosporin. Bias and imprecision do not exceed 15% for any drug and analysis time is 3 min per sample. Results of regular samples and VAMS processed by LCMS correlated well with the immunoassay in 35 patients treated with tacrolimus: [LCMS] = 0.9801*[immunoassay] + 0.1779 and [LCMS] = 0.9564*[immunoassay] – 0.4683 for regular samples and VAMS, respectively.
Conclusions: LCMS allows drug quantification over the range of peak and trough concentrations expected in organ transplantation. Determination of drug concentrations in VAMS, such as those obtained by fingerprick, can be further explored.
Dr Reimann completed his medical degree in Germany and obtained a medical doctorate investigating intracellular signal transduction of G protein-coupled receptors. He completed adult nephrology training in the Hunter New England Training Network in New South Wales in 2017 and undertakes further training in clinical pharmacology.