| Objective:The C-KIT gene in core binding factor acute myeloid leukemia(CBF-AML)patients in the incidence rate of 30-50% with C-KIT gene mutation in core bindingfactor leukemia(represented by CBF-AML-CK in this study)patients with poor prognosis,relapse is likely to occour after achieving complete remission(CR)undertaking normalized treatment,therefore,MRD monitoring in remission period is of great significance to patients,the main means is to detect the level of RUNX1-RUNX1T1 or CBFβ /MYH11 fusion gene by real time fluorescence quantitative PCR(RT-qPCR),the abnormal phenotypic leukemia cell(LAIP)proportions by flow cytometry(FCM).The purpose of this study is to explore the relationship between the level of C-KIT gene mutation detected by droplet digital PCR(ddPCR)and prognosis CBF-AML-CK in CR.C-KIT gene as a molecular marker monitoring MRD could provide clinical guidance for predicting recurrence,prognosis,timely adjustment of treatment programs.Methods:1.Collect bone marrow samples of 20 patients with CBF-AML-CK and extract DNA and RNA for preservation.2.The ddPCR technique based on MGB probe was used to quantify C-KIT gene mutation load at DNA level in 20 cases of CBF-AML-CK patients.3.Using RT-qPCR technology based on TaqMan probe,we detected the level of RUNX1-RUNX1T1 or CBFβ /MYH11 fusion gene in 20 cases of CBF-AML-CK patients at cDNA level.4.The C-KIT gene was amplified by the common PCR method and the mutation was detected by one generation Sanger sequencing in 20 cases of CBF-AML-CK patients.5.The proportion of LAIPs in 20 cases of CBF-AML-CK patients was detected by FCM.Results:1.20 patients with CBF-AML-CK were divided into continuous remission group and relapse group according to whether the patient relapsed during follow-up.The mean mutation level of the C-KIT gene was 0.07324% ± 0.32116% in the continuous remission group(six cases)and 8.79605% ± 4.44686% in the recurrence group(14 cases).The difference between the two groups was statistically significant(P = 0.005).2.The correlation coefficient between the mutation level of the C-KIT gene and the corresponding expression level of the RUNX1-RUNX1T1 or CBFβ/MYH11 fusion gene was 0.618 in CR,and the correlation between the two indexes was statistically significant(P = 0.004).3.Grouped according to the mutation level of C-KIT gene in CR.Compared with the low-level group,the high-level group has a high one-year recurrence rate(P= 0.004)and a low one-year survival rate(P = 0.039).The difference between the two groups has a statistically significant difference.4.The correlation coefficients between the mutation level of the C-KIT gene and annual recurrence rate,survival rate,overall survival,event-free survival,and complete response duration were higher than those of the RUNX1-RUNX1T1 or CBFβ/MYH11 fusion gene at the time of CR.The dynamic results showed mutation level of the C-KIT gene increased more evidently and earlier before recurrence.5.The MRD results from four methods of ddPCR,RT-qPCR,FCM,Sanger sequencing showed that the positive rate of ddPCR is highest,and compared with other methods had significant difference;prediction of Sanger sequencing of recurrence was statistically significant compared with the other three methods(P =0.008),the prediction of ddPCR and RT-qPCR on survival was statistically different(P=0.003,P=0.021).6.The comparison of the relationship between the results of the four methods and the prognosis showed that 11.5 years of age or older,male patients,a high proportion of LAIP and C-KIT gene mutation,SWOG group did not reach the high level of CRm were unfavorable prognostic factor.Conclusions:1.The mutation level of the C-KIT gene in CR is an important factor affecting the prognosis of CBF-AML-CK patients,and C-KIT gene can be used as MRD molecular marker.2.The ddPCR technique is more sensitive than other techniques to reflect the patient’s MRD status. |
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