中国实用儿科杂志

中国实用儿科杂志

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自动片段分析的多重PCR检测
儿童急性淋巴细胞白血病微小残留病探索

黄礼彬1，柯志勇1，张晓莉1，胡昌明2，朱晓霞2，罗学群1   

  1. 1.中山大学附属第一医院儿科，广东  广州   510080；2.广州金域医学检验中心，广东  广州   510330
  • 出版日期:2013-04-06 发布日期:2013-03-28
  • 基金资助:

    广东省科技计划项目（2008B030301104）

Detection of minimal residual disease in childhood acute lymphoblastic leukemia by multiplex PCR and automated fragment analysis. 

HUANG Li-bin**， KE Zhi-yong， ZHANG Xiao-li， HU Chang-ming， ZHU Xiao-xia， LUO Xue-qun.   

  1. Department of Pediatrics， The First Affiliated Hospital of Sun Yat-sen University， Guangzhou 510080， China
  • Online:2013-04-06 Published:2013-03-28

摘要：

摘要：目的    探讨运用自动片段分析的多重PCR检测IgH和（或）TCRγ基因单克隆重排作为检测儿童急性淋巴细胞白血病（ALL）微小残留病（MRD）方法的可行性。方法    采用1∶3匹配设计的病例-对照研究方法，收集2009年1月至2011年12月中山大学附属第一医院4例极早期复发的非高危儿童ALL（病例组）和危险度匹配的12例非高危儿童ALL（对照组）在初诊时、诱导化疗结束（距开始治疗5周，W5）、再诱导化疗前（距开始治疗23周，W23）及维持第3个月（距开始治疗44周，W44）的骨髓样本，治疗方案均按照广东2008 ALL协作组方案进行。用多重PCR方法定性检测各时间两组骨髓样本是否存在IgH和（或）TCRγ基因单克隆重排，PCR产物采用片段分析软件进行分析。检出单克隆重排的IgH和（或）TCRγ被认为是MRD阳性。结果    复发组4例和对照组12例ALL患儿在初诊时均监测到IgH单克隆重排，TCRγ单克隆重排阳性率56%。在W5时，病例组和对照组的MRD阳性率的差异无统计学意义（P＜0.05）。在W23和W44这两个时间点，病例组的MRD阳性率显著高于对照组，差异有统计学意义（P＜0.05）。结论    多重PCR联合自动片段分析检测IgH和（或）TCRγ基因单克隆重排的方法可以用于儿童ALL治疗过程中的MRD监测。

关键词: 急性淋巴细胞白血病, 微小残留病, 多聚合酶链反应

Abstract:

Abstract：Objective    To explore the feasibility of monitoring minimal residual disease （MRD） in childhood acute lymphoblastic leukemia （ALL） by detection of cloned IgH and TCRγ gene rearrangements using multiplex polymerase chain-reaction （PCR） and automated fragment analysis. Methods    In this 1∶3 matched case-control study， 4 cases of very early bone marrow relapsed non-high risk ALL （relapsed group） and 12 cases of non-relapsed non-high risk ALL as control （control group） were enrolled in the First Affiliated Hospital of Sun Yat-sen University from Jan. 2009 to Dec.2011. All patients were treated with Guangdong 2008 ALL protocol. Bone marrow samples were collected at four time points: at diagnosis， the end of induction， the beginning of  reinduction and the third month of maintenace treatment. Cloned IgH and TCRγ gene rearrangements were amplified by multiplex PCR. The clonality of PCR production was analyzed by GENEMAPPERID softwares. Detectable clonality of IgH/TCRγ was defined as MRD positive. Results       At diagnosis， the frequency of cloned IgH and TCRγ in all patients was 100% and 56%. The positive rate of MRD was found to have no statistical difference between two groups at the end of induction， while the difference of the MRD positive proportion between the two groups was statistically significant at the beginning of reinduction and the third month of maintenane therapy， which was much more higher in relapse group than that of control group. Conclusion    Detection of monoclonal IgH/TCRγ gene rearrangements by multiplex PCR with automated fragment analysis can be used as a method to monitor MRD during treatment for childhood ALL.

Key words: acute lymphoblastic leukemia, minimal residual disease, polymerase chain reaction