MS-PCR监测儿童急性非淋巴细胞白血病骨髓与外周血的微小残留病

MS-PCR Monitoring the Minimal Residual Disease of Children Acute Non-Lymphocytic Leukemia in Bone Marrow and Peripheral Blood

目的检验用外周血监测AML患儿MRD的可行性。方法选择确诊的AML患儿为研究对象。以甲基化的降钙素(calcitonin,CT)基因作为检测AMLMRD的目的基因。提取骨髓液、外周血的DNA,用MS-PCR方法扩增甲基化的CT基因,产物用电泳的方法检测,为定性检测。结果在所有患者中,治疗前骨髓液组CT甲基化阳性14例,阳性率77.7%,外周血组阳性14例,阳性率77.7%,两组阳性率无统计学差异;诱导28天骨髓液组CT甲基化阳性4例,阳性率22.2%,外周血组阳性3例,阳性率16.7%,阳性率无统计学差异。18例中13例(非M37例,M36例)完成巩固治疗,巩固结束后骨髓液组与外周血组CT甲基化阳性均有2例,阳性率均为15.4%,无统计学差异。另外,12例非M3组在诱导15天的CT甲基化检测,骨髓液组阳性11例,阳性率91.7%,外周血组阳性9例,阳性率75%,阳性率无统计学差异。结论①在治疗前、诱导15天、诱导28天和巩固结束分别用MS-PCR检测甲基化CT基因,骨髓组和外周血组的阳性率无统计学差别,提示外周血可能可以替代骨髓液用定性MS-PCR方法监测儿童AML的MRD。②用定性MS-PCR检测甲基化CT的方法监测AML的MRD,灵敏度为10-3。

Objective To explore the feasibility of detecting MRD of peripheral blood in children with AML. Methods Children with AML were selected in this study. Methylation ofCT gene as the purpose testing gene ofAML MRD, extracted the DNA of bone marrow, peripheral blood, then increased the methylation calcitonin （CT） gene with MS-PCR and detected the product with electrophoretic methods which was qualitation detection. Results During the 18 AML patients, the bone marrow and peripheral blood CT gene＇s methylation was both positive in 14 patients before treatment, and the positive rate was both 77.7%. There was no significant difference in these groups about the total positive rate before treatment; the bone marrow calcitonin gene＇s methylation was positive in 4 patients on inductive-28th-day and the positive rate was 22.2%. The peripheral blood was positive in 3 patients, and the positive rate was 16.7%. There was no significant difference in their total positive rates; the bone marrow group and peripheral positive rates were both 15.4% in the 13 patients （including 6 M3 and 7 non-M3） after solidified treatment, and there was no significant difference in their total positive rates. During the 12 non-M3 patients, the positive rate of bone marrow was 91.7% and the positive rate of peripheral blood was 75%, there was no significant difference in their total positive rates. Conclusions ① There is no significant difference between MRD from peripheral blood and MRD from bone marrow before treatment, on inductive-28th-day, after solidified treatment. Peripheral blood may possiblely replace bone marrow to detect the MRD before treatment, on inductive-28th-day, after solidified treatment.②The sensitivity of MS-PCR is 10-3 .