Improving the Pharmacokinetic Analysis of the Once Daily IV Busulfan Regimen for Allogeneic Stem-Cell Transplants: A Scientific Method Modification.

Derivitization, a chemical technique used to transform a compound into a similar product with a different structure required by certain detection methods, was carried out for the detection of Busulfan (Bu) in human plasma. Using a proficient scientific method and enhancing its clinical association our method furthered the pre-column derivitization technique based on isocratic high performance liquid chromatography with ultraviolet detection (HPLC-UV). In-house adjustments have been made over the course of the last two years in order to increase the efficiency and turn-around of the method assay for clinical purposes. The concentration of drug in human plasma was measured with UV detection at 278nm by employing a Waters Nov-Pak C-18 analytical column heated to 40°C. The mobile phase, run at 1.5mL/min, consisted of a mixture of 20:80 deionized water and methanol respectively. All concentration-time plasma Bu data were analyzed by a non-compartmental analysis employing linear kinetics using WinNonlin Version 4.1 software (Pharsight Corporation, Mountain View, CA, USA). Analyses were performed on 155 allogeneic Stem-Cell Transplant (SCT) patients (pts). All pts received fludarabine 50mg/m² on days -6 to -2 and IV Bu at a “myeloablative” dose of 3.2mg/kg daily, on days -5 to -2 inclusive. Additional TBI 200cGy × 2 was given to 74 pts. Graft-vs.-host disease prophylaxis for all pts comprised cyclosporine A, “short course” methotrexate with folinic acid and Thymoglobulin (Genzyme) 4.5mg/kg administered in divided doses over 3 consecutive days pre-transplant finishing on D0. The range of area under the curve (AUC) per dose was 2184 to 7513μM·min (median 4509μM·min). Twenty nine pts (19%) had AUC < 3600μM·min and 17 (11%) had AUC > 6000μM·min. The remaining 109 (70%) were between 3600 and 6000μM·min, equivalent to an exposure of 900 – 1500μM·min per dose established as desirable from studies of qid oral Bu given with cyclophosphamide. We found that IV Bu resulted in a 3–4 fold variability in AUC with a significant number of pts with exposures deemed either too high or low. High exposures (> 6000uM*min) increase the risk of transplant-related death and there is justification for therapeutic monitoring. Our predicted adjustments obtained from a streamlined method are based on day -5 Bu doses. The adjusted doses are administered on days -3 and -2, allowing for a 24 hour turn around time between days -5 and -3.