Quantitative assessment of minimal residual disease in childhood lymphoid malignancies using an allele-specific oligonucleotide real-time quantitative polymerase chain reaction.
We developed an assay using a real-time quantitative polymerase chain reaction (RQ-PCR) for the quantitative assessment of minimal residual disease (MRD) in childhood lymphoid malignancies by using a consensus V-region probe combining a allele-specific oligonucleotide (ASO) reverse primer. Our strategy employs a set consisting of a consensus V-region probe, an ASO reverse primer, and a patient-specific forward primer for clonal antigen-receptor (IgH, immunoglobulin heavy chain; TCR, T-cell receptor) gene rearrangements (IgH-ASO and TCR-ASO RQ-PCR assays). The limit of detection in both assays was 5 copies of the target/10(5) cell equivalents. We tested the assays in 17 childhood malignancies (14 cases of acute lymphoblastic leukemia and 3 of non-Hodgkin's lymphoma). High correlation coefficients of the standard curves (>0.980) and PCR efficiency (>0.95) were achieved with all primer/probe sets. In 2 (12%) of the 17 patients, ASO primers could not be designed because there was no junctional N-sequence. The quantitative data suggest that the copy number of clonal antigen receptors markedly decreased after induction therapy in 15 of 17 patients and that 1 patient relapsed and died of the disease. Consensus probes make it possible to examine a large number of patients with only a limited number of probes. The strategy used for IgH-ASO and TCR-ASO RQ-PCR assays is accurate and reliable in the clinical prospective study of MRD in childhood lymphoid malignancies.
Tarusawa
Mitsu
M
Department of Pathology, Iwate Medical University School of Medicine, Morioka, Japan.
Yashima
Akiko
A
Endo
Mikiya
M
Maesawa
Chihaya
C
D000293:Adolescent; D000483:Alleles; D002648:Child; D002675:Child, Preschool; D015342:DNA Probes / Q000235:genetics / Q000592:standards; D005260:Female; D006801:Humans; D007223:Infant; D008228:Lymphoma, Non-Hodgkin / Q000175:diagnosis* / Q000235:genetics; D008297:Male; D018365:Neoplasm, Residual / Q000175:diagnosis / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods* / Q000592:standards; D054198:Precursor Cell Lymphoblastic Leukemia-Lymphoma / Q000175:diagnosis* / Q000235:genetics; D012015:Reference Standards; D012680:Sensitivity and Specificity
D015342:DNA Probes