Highly sensitive quantification of pemetrexed in human plasma using UPLC-MS/MS to support microdosing studies

Area of Science:

• Analytical Chemistry
• Pharmacology
• Clinical Chemistry

Background:

• Pemetrexed is a key antifolate drug for non-small-cell lung cancer and mesothelioma.
• Microdosing pemetrexed (100 μg) allows for safe drug-drug interaction and dose individualization studies.
• A highly sensitive bioanalytical assay is crucial for accurate microdose pharmacokinetic assessments.

Purpose of the Study:

• To develop and validate a sensitive bioanalytical assay for determining pemetrexed concentrations in human plasma after microdosing.
• To facilitate future pharmacokinetic studies involving pemetrexed microdosing.

Main Methods:

• Developed a reversed-phase ultra-high performance liquid chromatography (RP-UHPLC) method.
• Utilized [13C5]-Pemetrexed as the internal standard.
• Employed solid-phase extraction (SPE) for sample preparation and MS/MS for detection, with a total run time of 9.5 minutes.

Main Results:

• The assay was validated for concentrations ranging from 0.0250–25.0 μg/L.
• Average accuracies were 96.5%, with within-day and between-day precision <8.8% CV.
• Extraction recovery was 59 ± 1% for pemetrexed and 55 ± 5% for the internal standard; samples remained stable for 2 days.

Conclusions:

• A validated, sensitive RP-UHPLC-MS/MS assay for pemetrexed microdosing in human plasma was successfully developed.
• The assay is suitable for application in pharmacokinetic studies, including those within ongoing clinical trials.
• This method supports safer drug interaction and personalized dosing strategies for pemetrexed therapy.