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A Validated Assay to Quantify Osimertinib and Its Metabolites, AZ5104 and AZ7550, from Microsampled Dried Blood Spots
Bharat Venkatesh1, Alex Yuile2, Matthew J McKay1
1Bowel Cancer and Biomarker Laboratory, School of Medical Sciences, Kolling Institute, The University of Sydney, Sydney, NSW, Australia; and.
Background:
Osimertinib is an oral small-molecule tyrosine kinase receptor inhibitor used to treat non-small cell lung cancer (NSCLC) with a sensitizing epidermal growth factor receptor mutation. Patients may experience drug toxicity and require dose deescalation. The study aimed to quantitate osimertinib and its 2 active metabolites, AZ5104 and AZ7550, in microsampled dried blood spots (DBS) collected from patients with NSCLC using a hemaPEN device and compare them with plasma drug levels.
Methods:
A 6-min ultrahigh-performance liquid chromatography-tandem mass spectrometry method was developed and validated using plasma and DBS. The accuracy, selectivity, matrix effect, recovery, and stability were assessed using bioanalytical validation criteria. The hematocrit effect was investigated in DBS. Drug levels were measured in 15 patients with NSCLC, and the Bland-Altman method was used to compare measurements between plasma and DBS.
Results:
The validated assay determined accurate and precise quantities, respectively, for osimertinib in both plasma (93.2%-99.3%; 0.2%-2.3%) and DBS (96.7%-99.6%; 0.5%-10.3%) over a concentration of 1-729 ng/mL. The osimertinib metabolites, AZ5104 and AZ7550, were similarly validated in accordance with bioanalytical guidelines. For 30%-60% patient hematocrit, no hematocrit bias was observed with DBS for all analytes. The Bland-Altman method showed high concordance between plasma and DBS analyte levels. Stability experiments revealed that osimertinib and its metabolites were poorly stable in plasma at room temperature, whereas all analytes were stable in DBS for 10 days at room temperature.
Conclusions:
The measurement of osimertinib, AZ5104, and AZ7550 from hemaPEN microsampled DBS is a convenient and reliable approach for therapeutic drug monitoring that produces measurements consistent with plasma drug levels.
Insights
Dried blood spot (DBS) sampling using a hemaPEN device offers a reliable method for measuring osimertinib and its metabolites in non-small cell lung cancer (NSCLC) patients. This approach provides results consistent with plasma levels, aiding therapeutic drug monitoring.
Area of Science:
- Pharmacokinetics and Drug Metabolism
- Analytical Chemistry
- Oncology
Background:
- Osimertinib is a crucial tyrosine kinase inhibitor for EGFR-mutated non-small cell lung cancer (NSCLC).
- Drug toxicity necessitates dose adjustments, highlighting the need for precise therapeutic drug monitoring.
- Quantifying osimertinib and its active metabolites (AZ5104, AZ7550) is essential for optimizing patient treatment.
Purpose of the Study:
- To develop and validate a method for quantifying osimertinib and its metabolites in dried blood spots (DBS) using a hemaPEN device.
- To compare drug levels measured in DBS with traditional plasma measurements.
- To assess the stability and reliability of DBS samples for therapeutic drug monitoring in NSCLC patients.
Main Methods:
- Development and validation of a 6-minute ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) assay for plasma and DBS.
- Assessment of accuracy, selectivity, matrix effect, recovery, and stability according to bioanalytical guidelines.
- Comparison of plasma and DBS measurements using the Bland-Altman method, including hematocrit effect investigation.
Main Results:
- The validated UHPLC-MS/MS assay accurately quantified osimertinib (93.2%-99.6%) and its metabolites in both plasma and DBS.
- No significant hematocrit bias was observed in DBS samples across a 30%-60% hematocrit range.
- Analyte stability was superior in DBS (10 days at room temperature) compared to plasma, which showed poor stability.
Conclusions:
- Microsampling of DBS with a hemaPEN provides a convenient and reliable method for measuring osimertinib and its metabolites.
- DBS measurements demonstrate high concordance with plasma drug levels, supporting its use in therapeutic drug monitoring.
- The stability of analytes in DBS enhances its practicality for clinical applications in NSCLC treatment.
