Comparison of the Innofluor® certican assay with HPLC-UV for the determination of everolimus concentrations in heart transplantation

Objectives: Therapeutic monitoring of everolimus with chromatographic methods (HPLC) enabled effective immunosuppression while limiting the incidence of drug-related adverse events. A fluorescence polarization immunoassay (FPIA) has been recently developed for the assessment of everolimus levels. The present study was designed to evaluate FPIA performance and to compare it to HPLC. Design and methods: The performance of HPLC and FPIA was initially tested using drug-free whole blood spiked with different amount of everolimus concentrations and, subsequently, by analyzing 113 trough blood samples from heart transplant recipients chronically given everolimus as part of their immunosuppressive regimen. Results: Inaccuracy and imprecision of both methods were below 15%. The correlation between everolimus concentrations and measured FPIA and HPLC was good, with a Pearson coefficient of 0.9118. The FPIA gave a mean overestimation of 24.3% as compared with HPLC. As additional analysis, cross-reactivity ranging from 85.4% to 138.0% was found with sirolimus, an immunosuppressant with a chemical structure close to everolimus. Conclusion: The FPIA demonstrated acceptable performance for therapeutic drug monitoring of everolimus, and is a viable alternative to HPLC-based methods.