Introduction to high-performance liquid chromatography-tandem mass spectrometry: The gold standard for monitoring immunosuppressive drugs

Many of the immunosuppressive drugs require monitoring to provide optimal and individualised therapy for patients who have undergone solid organ transplantation. Clinical laboratories have the choice of methodology used for these drug analyses and this choice may impact on the quality of pharmacokinetic data provided to the transplant physician. The majority of laboratories use either high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) or antibody-based immunoassays. Most of the measured immunosuppressive drugs are neutral in nature and would appear unsuited to the ionisation processes of HPLC-MS/MS. However, ionisation of these compounds can be achieved by the addition of modifiers, such as
ammonium acetate, to the mobile phase which promote adduct formation. The use of successive (tandem) mass analysers provides for high selectivity and sensitivity which is required for the measurement of the more potent drugs (i.e. tacrolimus, sirolimus and everolimus) and simplifies the chromatographic separation. Thus making this technique highly suited to the clinical requirements for monitoring these drugs. While immunoassays are widely used for analysis of immunosuppressive drugs, there are several drawbacks to their use. Firstly, these methods tend to suffer from over-estimation in results due to the non-specific binding (to varying degrees) of metabolites with the assay antibody and have been shown to be influenced by the biochemical status of the patient, such as hematocrit. Secondly, some immunoassays have been shown to have poor accuracy and imprecision at the critical lower end of the therapeutic range. The aims of this presentation are to provide an overview on HPLC-MS/MS instrumentation, to discuss its use in the therapeutic drug monitoring of immunosuppressive drugs, and to provide a critical comparison with immunoassays.

Keywords: immunosuppressive drugs, Organ Transplantation, Bladder, cystofix, knotting.