A-344 A therapeutic drug monitoring testing strategy for immunosuppressant drugs from microsampled dried whole blood

Abstract

Therapeutic drug monitoring (TDM) of immunosuppressant drugs (ISD) following solid organ transplantation minimizes a patient’s risk of toxicity and rejection. Frequent blood sampling is required to appropriately quantify drug exposure and adjust dosage. Specimen collection via microsampling represents an emergent alternative to traditional venipuncture. Such minimally invasive techniques permit decentralized or home specimen collections for pediatric patients requiring TDM of ISD. We describe the validation of a volumetric absorptive microsampling (VAMS) testing strategy for the analysis of the ISD cyclosporine, sirolimus and tacrolimus in dried whole blood. 30 µL of fluid whole blood were volumetrically sampled from commercially available calibration standards, quality control (QC) or remnant patient specimens collected by venipuncture and dried on the VAMS tip of the Mitra device (Neoteryx). An ultra-sonication based solvent extraction procedure was optimized to remove the ISD from the dried whole blood. Each were respectively quantified using an established turbulent flow liquid chromatography tandem mass spectrometry (TFC-MS/MS) assay for fluid whole blood ISD testing. Performance of the VAMS strategy was determined through an assessment of linearity, precision, accuracy and the effects of ground transit. A 98:2 (v/v) water:methanol extraction solvent provided the highest average recovery for all ISD. Matrix-matched dried external calibration standards (Waters) recovered within 20% of their respective target concentrations (0 to 1500 ng/mL for cyclosporine and 0 to 30 ng/mL sirolimus and tacrolimus). The average calibration curves (N=5) for cyclosporine (R^2: 0.9977, slope: 0.842, intercept: 3.36), sirolimus (R^2: 0.9996, slope: 0.984, intercept: -0.18) and tacrolimus (R^2: 0.9828, slope: 1.007, intercept: -0.03) were linear. The average recovery of dried matrix-matched external QC (N=3 concentrations, BioRad) was 93±13%, 105±4% and 109±7% for cyclosporine, sirolimus and tacrolimus, respectively. Average inter-day (N=20 days) precision (%CV) of the dried external QC was 17.3%, 12.8% and 13.1% for cyclosporine, respectively. Accurate ISD results (N=3) were obtained from dried microsamples of archived College of American Pathology (CAP) proficiency testing material. Dried whole blood specimens were shipped from different locations in Massachusetts to our laboratory for testing. The maximum transit duration, temperature and relative humidity (%RH) ranges experienced were 6 days 19 hours 21 minutes, -8.4 to 22.9°C and 24.7 to 74.8%RH, respectively. All ISD were stable in the microsampled dried whole blood under all shipping conditions, with average concentration biases of 24.0, 0.2 and 0.5 ng/mL for cyclosporine, respectively. Concentrations of ISD derived from paired testing of dried and fluid whole blood (N=30) were highly correlative based on Deming regression: cyclosporine (R: 0.9344, slope: 1.10; bias: 9.9 ng/mL or 5.8%), sirolimus (R: 0.9340; slope: 0.845; bias: 0.1 ng/mL or 1.8%) and tacrolimus (R: 0.9151; slope: 0.0.958; bias: 0.6 ng/mL or 6.2%). Cyclosporine, sirolimus and tacrolimus results from the microsampled dried whole blood TFC-MS/MS testing strategy were accurate relative to traditional analyses of fluid whole blood specimens. Testing dried whole blood from home collected specimens may be appropriate for pediatric patients requiring TDM of ISD.