EVALUATION OF MINIMAL RESIDUAL DISEASE IN ACUTE MYELOID LEUKEMIA WITH NPM1 MARKER

Background: Minimal residual disease (MRD ) tests provide early identification of hematologic relapse and timely management of AML patients. Molecular markers are useful in the dissection of cytogenetically normal AML patients into prognostically different subgroups. About 60% of adult CN-AML have a mutation in exon 12 of the NPM1 gene. This mutation is specific for malignant clone and potentially is a good marker of MRD.we set up a quantitative test for quantifying NPM1 type A mutation and then AML patients carrying this mutation at the time of diagnosis, were followed up.Methods: we prepared plasmids containing a cDNA fragment of the NPM1 and ABL genes by PCR cloning. ABL served as a control gene. The plasmids were used to construct standard curves and quantitation of each of the genes. Eleven patients were analyzed using established method. Serial PB and/or BM samples ( 71 samples ) were taken in 1-3 month intervals (mean 1.5-month intervals) and median follow up duration after chemotherapy was 11 months ( 5- 28.5 months ).Results: we developed RNA-based RQ-PCR to quantitation of NPM1 mutation A with sensitivities of 10(-5). The percent of NPMmut/ ABL level showed a range between 132 and 757 with a median of 383.5 in samples at diagnosis. The median NPMmut transcript level log reduction was 3 log. Relapse occurred in 54.5% of patients ( 6 cases), All cases at diagnosis demonstrated the same mutation at relapse. In patients who experienced relapse, log reduction levels of NPM1 mRNA transcript after therapy were 4(2 patients), 3(two patients) and 1 log(2 patient). Totaly, NPMmut level showed less than 5 log reduction in all of them, Whereas this reduction was 5 - 6 log in other patients. Conclusion: Despite the limitations of this study in terms of sample size and duration of follow up , it showed the accuracy of set up for detection of mutation and this marker has worth for following up at different stages of disease. Because of high frequency, stability, specificity to abnormal clone and high sensitivity, NPM1 is a suitable marker for monitoring of NPMc+ AML patients.

Keywords:

Acute myeloid leukemia , Minimal residual disease , NPM1 mutation , Quantitative reverse transcriptase-polymerase chain reaction