A rapid & sensitive liquid chromatography- tandem mass spectrometry method for the quantitation of busulfan levels in plasma & application for routine therapeutic monitoring in haematopoietic stem cell transplantation.

BACKGROUND & OBJECTIVES: Busulfan (Bu) in combination with cyclophosphamide is widely used in myeloablative conditioning regimen prior to haematopoietic stem cell transplantation (HSCT). Its narrow therapeutic range and toxic side effects at high systemic exposure and graft rejection at low exposure emphasize the need for busulfan dose optimization using targeted dose adjustment prior to HSCT. We report here a rapid and sensitive method to quantitate busulfan plasma levels in patients receiving busulfan as part of pre-transplant conditioning. METHODS: The method involves simple protein precipitation of the plasma followed by analysis using a high performance liquid chromatography (HPLC) with tandem mass spectrometry - electrospray ionization technique (LC-ESI MS/MS) in positive ionization mode and quantified using multiple reaction monitoring (MRM). Deuterated busulfan (d8-busulf`an) was used as the internal standard. RESULTS: The method was linear for the concentration ranging from 0 to 4000 ng/ml of busulfan with a limit of detection of 2 ng/ml and limit of quantitation of 5 ng/ml. The assay was accurate for serial concentrations of Bu in plasma for five consecutive days and the CV was less than 10 per cent. CONCLUSION: Using this rapid and sensitive method, busulfan levels were targeted and subsequent doses adjusted at our center in 26 patients receiving high dose busulfan in combination with cyclophosphamide or fludarabine.

Frequency of TPMT alleles in Indian patients with acute lymphatic leukemia and effect on the dose of 6-mercaptopurine.

Functional polymorphisms in the thiopurine methyl transferase (TPMT) gene have been associated with varying levels of enzyme activity and the occurrence of toxicity related to thiopurines. A total of 98 patients (66 pediatric and 32 adults) with precursor B acute lymphoblastic leukemia (Pre-B ALL) were evaluated for TPMT gene polymorphisms. The inability to tolerate 6-mercaptopurine (6-MP) at conventional doses was considered as a surrogate marker of hematologic toxicity. The allele frequency of TPMT*2, *3A, *3B and *3C in the study population was 0.5, 0, 0 and 2.6%, respectively, similar to the frequency observed in other Asian populations. Five patients were heterozygous for TPMT*3C variant allele, and one of these patient's was compound heterozygous with TPMT*2 variant as the other allele. The impact of TPMT polymorphisms on the toxicity and treatment outcome was assessed in 66 pediatric patients only, as there was no variant TPMT detected in the adult patients. Three of the 5 patients (60%) heterozygous for TPMT*2 or TPMT*3C polymorphisms and 12/61 patients (20%) with wild type TPMT genotype had more than 10% of reduction of 6-MP dose (P=0.07). The presence of TPMT polymorphisms did not seem to completely explain the variation in 6-MP toxicity in this small group of patients. Other novel variants in TPMT or variations in the genes involved in transport and biotransformation of 6-MP need to be evaluated in the Indian population.

Single-agent arsenic trioxide in the treatment of newly diagnosed acute promyelocytic leukemia: long-term follow-up data.

PURPOSE We previously reported our results with a single-agent arsenic trioxide (ATO) -based regimen in newly diagnosed cases of acute promyelocytic leukemia (APL). The concern remained about the long-term outcome of this well-tolerated regimen. We report our long-term follow-up data on the same cohort. PATIENTS AND METHODS From January 1998 to December 2004, 72 patients with PML/RARalpha+ APL were enrolled. All patients were treated with a single-agent ATO regimen. Results Overall 62 (86.1%) achieved a hematologic remission (complete remission). After the initial report, an additional seven patients have relapsed for a total of 13 relapses. There were no additional toxicities to report on follow-up. At a median follow-up 60 months, the 5-year Kaplan-Meier estimate (+/- SE) of event-free survival, disease-free survival, and overall survival (OS) was 69% +/- 5.5%, 80% +/- 5.2%, and 74.2% +/- 5.2%, respectively. The OS in the good risk group as defined by us remains 100% over this period. CONCLUSION Single-agent ATO as used in this study in the management of newly diagnosed cases of APL is safe and is associated with durable responses. Results in the low-risk group are comparable to that reported with conventional therapy while additional interventions would probably be required in high-risk cases.