Significance Of Study On Plasma Free DNA FLT3-ITD Mutation And ITD Features Of AML Patients For Individualized Diagnosis

Objective:The purpose of our research is to detect FLT3-ITD(internal tandem duplication, ITD)mutation and ITDs (FLT3-ITD positive patients) from plasma circulation cell free DNA(ccfDNA),and comparing it with the DNA test results of bone marrow cells, further to provides a new strategy for individualized detection, diagnosis, and molecular target therapy efficacy monitoring with acute myelocytic leukemia(AML) patients.Methods:1. Polymerase chain reaction (PCR) amplification the housekeeper of88newly diagnosed patients with AML and the40controls (20heathlty persons and20patients with the benign diseases of the blood system) plasma ccfDNA and the DNA from bone marrow cells to exam the success of extraction.The DNA concentration in plasma from88patients newly diagnosed with AML and the40controls were analyzed by nucleic acid concentration measuring instrument to detact the difference.2.PCR is used for the DNA products from both the cell and plasma to detect FLT3-ITD mutation of the88newly diagnosed patients,then10μl PCR products were separated on the2%agarose gel and viewed by ethidium bromide (EB)staining. By using tanon image analysis software to explain the final results.3.We use the capillary electrophoresis (CE) method to detect the ITDs of the patient with FLT3-ITD mutation from the plasma ccfDNA and DNA from bone marrow cells. The two groups of test results will be compared.Results:1. Plasma concentration of88AML patients in experimental group is significantly higher than the control groups (P<0.05),28times of normal people,and23times of the patients with benign diseases of the blood system.There is no statistical differences between the normal people and the patients with benign diseases of the blood system (P>0.05).Among the patients of different FAB classification,we couldn’t see the differences.2. FLT3-ITD was detected in18(20.5%,18/88)AML sample in both cellular and plasma.3.18patients who with FLT3-ITD capillary electrophoresis results showed that13cases (72.22%,13/18) have only one ITD band in both cellular and plasma DNA,5cases (27.78%,5/18) showed ITDs in plasma but not in cellular.Conclusions:1.The concentration of ccfDNA from peripheral blood(PB) plasma sample of newly diagnosed patients with AML is significantly higher than normal people and patients with benign diseases.2.The detection of FLT3-ITD mutation in plasma consistent with the test in bone marrow cells, which tip to us that we can be through the detection of specificity mutations from ccfDNA to diagnose leukemia. For the noninvasive diagnosis and monitoring of prognosis of leukemia has great significance.3.18patients who with FLT3-ITD mutation capillary electrophoresis results showed that15cases which have only one ITD band (13cases) and ITDs(2cases) in both cellular and plasma DNA,3cases showed ITDs in plasma but not in cellular,which hint us that to detect ccfDNA from plasma maybe a better way to diagnose AML than from bone marrow cells.