Development and validation of a methotrexate adherence assay.

Area of Science:

• Pharmacology
• Analytical Chemistry
• Rheumatology

Background:

• Methotrexate (MTX) is a first-line therapy for rheumatoid arthritis (RA).
• Approximately 40% of RA patients exhibit non-adherence to oral MTX therapy.
• Non-adherence can lead to treatment failure and the need for more expensive biological therapies.

Purpose of the Study:

• To develop and validate a novel assay for accurately measuring MTX adherence.
• To determine the diagnostic accuracy of a high-performance liquid chromatography-selected reaction monitoring-mass spectrometry (HPLC-SRM-MS) method for MTX analysis.
• To provide an objective tool for monitoring MTX adherence in RA patients.

Main Methods:

• Developed and validated a two-compartment pharmacokinetic model for MTX.
• Utilized HPLC-SRM-MS for plasma MTX concentration analysis over a 6-day period.
• Validated the assay using independent real-world patient samples (n=50) and an observational cohort (n=138).

Main Results:

• The developed pharmacokinetic model accurately predicted MTX adherence.
• Assay optimization confirmed correct identification of MTX adherers.
• The validated HPLC-SRM-MS assay demonstrated 95% sensitivity in detecting MTX adherence in real-world samples.

Conclusions:

• Non-adherence to MTX is prevalent and impacts RA disease activity.
• HPLC-SRM-MS plasma analysis provides an accurate and objective method for detecting MTX adherence.
• This validated test can be implemented in clinical settings to identify patients requiring adherence support.