摘要
本研究目的旨在构建实时荧光定量RT-PCR(RQ-RT-PCR)方法检测用的am l1/eto融合基因的RNA标准品,并以此作为工具监测急性髓系白血病M2患者微小残留病(MRD)的状态。应用定性的RT-PCR筛选患者标本中的am l1/eto融合基因,经体外扩增转录,得到1010拷贝数RNA标准品,并对37例患者骨髓/外周血标本进行了检测。结果表明:构建的RNA标准品具有线形良好的标准曲线,扩增效率较高,特异性较好;患者初诊组和复发组的目的基因am l1/eto的相对值的平均值高于完全缓解组(CR)(p<0.05);随访的患者中初诊时目的基因am l1/eto的相对值很高,完全缓解时降低,复发时明显升高。初诊和CR两个时间点目的基因am l1/eto的相对值相差2个数量级以上时,病人的复发危险性相对较小,如果am l1/eto基因的相对值持续升高,则提示患者的预后较差。结论:RQ-RT-PCR方法检测am l1/eto融合基因的敏感性和特异性比较高,可信度和稳定性较好;对am l1/eto融合基因阳性患者MRD定量动态监测有助于初步评价治疗效果,估计患者的复发危险度,因此有望成为确定强化治疗的时机、延长病人完全缓解期的重要手段。
This study was aimed to construct the standard product for detecting the aml1/eto fusion gene by real-time quantitative reverse transcription polymerase chain reaction （RQ-RT-PCR）, monitoring minimal residual disease （MRD） in the patents with AML-M2. Having obtained the aml1/eto fusion gene from the patients by the RT-PCR with a pair of specific primers, the RNA standard product as 10^10 copies was gained after amplifying and transcribing in vitro and was used for detecting bone marrow and peripheral blood samples from 37 patients. The results indicated that the standard product constructed above displayed a standard curve which showed a linear correlation of the Ct with the log of the RNA concentration of each standard dilution. The average relative levels of aml1/eto fusion gene in the patients at diagnosis and the patients in relapse were higher than those in patients ongoing complete remission （CR） （p 〈 0.05 ）. The relative level of aml1/eto fusion gene of the follow-up patients was higher at diagnosis, and lower in patients ongoing CR, then went up again at relapse. The patients whose relative level of aml1/eto fusion gene in CR decreased by 2 log even lower than at diagnosis had a lower risk of relapse. If the relative level of aml1/eto fusion gene kept increasing, the patients had a poor prognosis. It is concluded that RQ-RT-PCR is a sensitive, specific, reliable and reproducible methed for monitoring aml1/eto fusion gene. Application of RQ-RT-PCR to detect aml1/eto fusion gene for monitoring MRD in AML-M2 is helpful to assess the response of therapy and estimate the risk of relapse, RQ-RT-PCR may become an important method to decide the time for intensified therapy and prolong CR for patients.
出处
《中国实验血液学杂志》
CAS
CSCD
2009年第1期17-22,共6页
Journal of Experimental Hematology
