Characteristics of TCR β gene rearrangements in adult patients with T-lineage acute lymphoblastic leukemia and its significance in quantitation of minimal residual disease / 中华检验医学杂志

ABSTRACT

Objective To develop allele specific oligonucleotide(ASO) -PCR assay based on TCR βgene rearrangements and provide a screening method for minimal residual disease (MRD) in adult patients with T-lineage acute lymphoblastic leukemia (T-ALL).Methods DNA samples from newly diagnosed 20 adult T-ALL patients were obtained.The TCR β gene rearrangements were detected by multiplex PCR,which included 38 paired of primers in 3 reaction tubes.Gel electrophoresis and two-color Gene Scanning was also applied for clonality analysis of TCR β followed by sequencing and subsequent blasting for monoclonal PCR products in four patients.ASO primers were designed based on the sequence of junction regions.MRD were detected in the bone marrow by RQ-PCR with ASO upstream primers, consensus Jβprobes and downstream primers.Results The detection rate of the clonal TCR β rearrangements was 85.0% (17/20).At least one complete Vβ-Jβ rearrangement could be detected at the time of diagnosis in 16 out of 17 patients(94.1%, 16/17).Incomplete Dβ-Jβ rearrangement could be detected in 7 patients (41.2% ,7/17).The positivitity rate of Vβ-Jβ to Dβ-Jβ was 2∶1 (94.1% versus 41.2% ).Two-color Gene Scanning analysis showed the Jβ2 family was used more frequently than the Jβ1 family (73% versus 27% ).The slopes of the standard curves ranged from - 3.60 to - 3.27.The correlation coefficients of all four standard curves were more than 0.99.The detection sensitivity of ASO-PCR was 4 × 10 -5 μg/μl.The fluorescence background were detected at a low level.Quantitative MRD values of TCR β rearrangement in sequential BM specimens of 4 adult T-ALL patients were monitored during the treatment, including complete remission after induction and after consolidation therapy. RQ-PCR showed the MRD values of TCR β rearrangement were gradually decreased in response to the treatment.Conclusions The quantification of TCR β rearrangement by ASO-PCR approach is sensitive, specific and reliable for the accurate evaluation of malignant clones.It is suitable for the monitoring of minimal residual disease of adult T-ALL patients.