Development and Validation of UHPLC-MS/MS Assay for Therapeutic Drug Monitoring of Highdose Methotrexate in Children with Acute Lymphoblastic Leukemia

• Published in: Drug design, development and therapy development and therapy, 2020-11, Vol.14, p.4835
• Main Authors: Lian, Le-jing, Lin, Bin, Cui, Xiao, He, Jie, Wang, Zhe, Lin, Xiao-dong, Ye, Wei-jian, Chen, Rui-jie, Sun, Wei
• Format: Article
• Language: English
• Subjects: Acute lymphocytic leukemia; Analysis; Complications and side effects; Formic acid; High performance liquid chromatography; Ionization; Methotrexate; Nitriles; Organic acids
• ISSN: 1177-8881; 1177-8881
• DOI: 10.2147/DDDT.S271568

Purpose: Precise and timely detection of methotrexate (MTX) concentration played a key role in high-dose MTX individualization therapy in acute lymphoblastic leukemia (ALL) children to avoid serious adverse effects or nonresponse. This report described a sensibility and validation of ultra-high performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) method for therapeutic drug monitoring (TDM) of methotrexate concentration in children's plasma. Methods: One-step protein precipitation of samples was accomplished by adding 200 [micro]L of acetonitrile to 100 [micro]L of plasma sample. The separation of plasma samples was carried out on a ZORBAX Eclipse Plus C18 Rapid Resolution HD column with gradient elution using a mobile phase constituted of acetonitrile and 1% formic acid. The detection was executed by electrospray ionization (ESI) of triple quadrupole tandem mass spectrometer (TQMS) in the multiple reaction monitoring (MRM) mode with the transitions m/z 455.2 [right arrow] 307.9 for methotrexate and m/z 458.2 [right arrow] 311.2 for IS, separately. Linear concentration range of the calibration curve was 44-11,000 nmol/L and 44 nmol/L was the lower limit of quantification. Results: The methotrexate elution time was at 1.577 min, and the overall running time was only 3.3 min. The intra- and interday precision for all the analysis results was within 11.24%, and mean recoveries rate of methotrexate exceeded 87.98%. Conclusion: The described and fully validated UHPLC-MS/MS method was successfully applied in clinical TDM after infusion of high-dose methotrexate 1-5 g/[m.sup.2] to 41 childpatients. Keywords: high-dose methotrexate, UHPLC-MS/MS, therapeutic drug monitoring, children, acute lymphoblastic leukemia