Nakao M, Yokota S, Horiike S, Taniwaki M, Kashima K, Sonoda Y, Koizumi S, Takaue Y, Matsushita T, Fujimoto T, Misawa S
Third Department of Internal Medicine, Kyoto Prefectural University of Medicine, Japan.
Leukemia. 1996 Sep;10(9):1463-70.
We investigated TEL/AML1 fusion mRNA in 108 children with acute lymphoblastic leukemia (ALL) (86 B-lineage ALL, 15 T-ALL, two mixed lineage ALL, and five other phenotypes) using reverse transcriptase-polymerase chain reaction (RT-PCR). TEL/AML1 transcripts were found in 14 patients (13%) including three relapsed patients, and were unexceptionally limited to B-lineage ALL patients. The incidence of TEL/AML1 transcripts among B-lineage ALL was 16% (14/86). The reciprocal AML1/TEL transcripts were detected in 12 (86%) of the 14 cases expressing a TEL/AML1 transcript. In three cases, the TEL gene was fused to exon 3 of the AML1 gene, and to exon 2 in the remaining cases. To evaluate the amount of TEL/AML1 molecules for the quantification of a minimal residual disease (MRD), a plasmid vector which contained either a long TEL/AML1 PCR product (464 bp) or a short one (425 bp) was used as a competitor. We amplified RNAs obtained from bone marrow (BM) at complete remission or from peripheral blood stem cell (PBSC) harvests in two representative cases. For one PBSC harvest showing a positive result, a competitive PCR was carried out to quantify the amount of MRD. A 1:4 dilution series of competitor vectors was constructed, and each vector was added to a PCR reaction which contain a constant amount of cDNA obtained from the PBSC harvest. An equivalent point was compared to that of corresponding samples at diagnosis. Using this method, MRD in the PBSC harvest was 3.9:10(3). Our results elucidated the incidence, lineage-specificity, and variant forms of TEL/AML1 fusion transcripts in childhood ALL. Since the percentage of other chromosomal translocations in childhood ALL is not more than 5%, TEL/AML1 transcript would be the most feasible clone-specific marker for these patients. In addition, our method could be a powerful tool for quantification of the TEL/AML1 transcript and for the detection of MRD.
我们采用逆转录聚合酶链反应(RT-PCR)对108例急性淋巴细胞白血病(ALL)患儿(86例B系ALL、15例T-ALL、2例混合系ALL和5例其他表型)进行了TEL/AML1融合mRNA检测。在14例患者(13%)中发现了TEL/AML1转录本,其中包括3例复发患者,且无一例外均局限于B系ALL患者。B系ALL中TEL/AML1转录本的发生率为16%(14/86)。在表达TEL/AML1转录本的14例病例中的12例(86%)检测到了相互的AML1/TEL转录本。在3例病例中,TEL基因与AML1基因的外显子3融合,其余病例中与外显子2融合。为了评估用于微小残留病(MRD)定量的TEL/AML1分子数量,使用了一种包含长TEL/AML1 PCR产物(464 bp)或短产物(425 bp)的质粒载体作为竞争物。我们扩增了来自两例代表性病例完全缓解期骨髓(BM)或外周血干细胞(PBSC)采集物的RNA。对于一份呈阳性结果的PBSC采集物,进行了竞争性PCR以定量MRD的数量。构建了竞争物载体的1:4稀释系列,并将每个载体加入到含有从PBSC采集物中获得的恒定数量cDNA的PCR反应中。将等效点与诊断时相应样本的等效点进行比较。使用该方法,PBSC采集中的MRD为3.9:10(3)。我们的结果阐明了儿童ALL中TEL/AML1融合转录本的发生率、谱系特异性和变异形式。由于儿童ALL中其他染色体易位的百分比不超过5%,TEL/AML1转录本将是这些患者最可行的克隆特异性标志物。此外,我们的方法可能是定量TEL/AML1转录本和检测MRD的有力工具。