A Simple and Sensitive LC-MS/MS Method for the Simultaneous Determination of Cyclophosphamide and Doxorubicin Concentrations in Human Plasma

Background: The combination of cyclophosphamide and doxorubicin is commonly used in the adjuvant treatment of breast cancer in women with a high risk of recurrent disease. It is essential to devise dosing strategies for such drugs in order to minimise their in-use toxicity and to maximise their efficacy. Data have shown that concentration directed rather than dose targeted therapy provides better clinical outcomes. Therefore, analytical methods that permit the simple, sensitive and accurate determination of such compounds in human plasma will assist in drug dosing strategies Methods: We report herein a method for the LC-MS/MS determination of cyclophosphamide and doxorubicin concentrations in patient plasma, following intravenous chemotherapy using protein precipitation without time-consuming supernatant drying and reconstitution steps, at the same time using a HILIC pre-column to further remove residual phospholipids. Results: The method has a less than 15% RE and less than 5% RSD both intra- and inter- day. It has following principal advantages over recently published LC-MS/MS methods for cyclophosphamide and doxorubicin in human plasma: 1) the sample preparation method is very simple and rapid; 2) the method has a very low LLOQ (cyclophosphamide 0.5 ng/mL and doxorubicin 1.0 ng/mL) at the same time the precision and accuracy meeting the primary requirements established by the FDA; and 3) the procedure only requires a very small plasma volume (30 μL). Conclusion: Such a methodology, when applied in close temporal and geographical proximity to patients will permit a short turn-around time and thereby facilitate the clinical interpretation and feedback to improve patient outcomes.