Development and validation of an LC-MS/MS method for the comprehensive quantification of tamoxifen and its metabolites in volumetric absorptive microsampling: clinical application in breast cancer patients.

Tamoxifen requires biotransformation into metabolites to exert antiestrogenic activity, and variability in metabolite exposure contributes to interindividual differences in clinical outcomes. Volumetric absorptive microsampling (VAMS) provides a convenient and minimally invasive alternative for therapeutic drug monitoring (TDM). However, analytical methods enabling comprehensive quantification of tamoxifen and its metabolites in VAMS remain limited, and translation of VAMS-derived concentrations into plasma-equivalent values has not been well established. In this study, an LC-MS/MS method was developed and fully validated for simultaneous quantification of tamoxifen and six metabolites in VAMS samples. Water prerinsing prior to methanol extraction improved analyte recoveries to 89.2-111.2%. Stability evaluation of VAMS samples revealed poor room-temperature stability for most analytes, whereas all analytes remained stable for at least 100 days at -80 °C. Experimentally derived plasma-to-blood concentration ratios (Cp/Cb, 1.41-1.83) were further applied to estimate plasma-equivalent concentrations from VAMS samples. Clinical applicability was evaluated using paired VAMS and plasma samples collected from ten tamoxifen-treated breast cancer patients. Strong correlations between VAMS-derived and measured plasma concentrations were observed (R2 = 0.77-0.98). Bland-Altman analysis further demonstrated good agreement, with more than 90% of values falling within the ±1.96 SD limits. These results demonstrate that the proposed workflow provides a reliable approach for comprehensive tamoxifen metabolite quantification in VAMS samples and supports plasma-equivalent concentration estimation for comparison with existing plasma-based therapeutic data. The proposed strategy may facilitate the clinical implementation of VAMS-based TDM for personalized tamoxifen therapy.