Published online before print March 26, 2009

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Journal of Molecular Diagnostics 2009, Vol. 11, No. 3
Copyright © 2009 American Society for Investigative Pathology & Association for Molecular Pathology
DOI: 10.2353/jmoldx.2009.080048

Sensitive and Specific Measurement of Minimal Residual Disease in Acute Lymphoblastic Leukemia

Alexander A. Morley^*, Sue Latham^*, Michael J. Brisco^*, Pamela J. Sykes^*, Rosemary Sutton, Elizabeth Hughes^*, Vicki Wilczek^*, Bradley Budgen^*, Katrina van Zanten^*, Bryone J. Kuss^*, Nicola C. Venn, Murray D. Norris, Catherine Crock, Colin Storey, Tamas Revesz and Keith Waters

From the Department of Haematology and Genetic Pathology, * Flinders University and Medical Centre, Bedford Park, Adelaide; the Children’s Cancer Institute Australia for Medical Research, University of New South Wales, Sydney; the Department of Clinical Haematology and Oncology, Royal Children’s Hospital, Parkville; and the Department of Haematology/Oncology, Women’s and Children’s Hospital, Adelaide, Australia

A sensitive and specific quantitative real-time polymerase chain reaction method, involving three rounds of amplification with two allele-specific oligonucleotide primers directed against an rearrangement, was developed to quantify minimal residual disease (MRD) in B-lineage acute lymphoblastic leukemia (ALL). For a single sample containing 10 µg of good quality DNA, MRD was quantifiable down to approximately 10^–6, which is at least 1 log more sensitive than current methods. Nonspecific amplification was rarely observed. The standard deviation of laboratory estimations was 0.32 log units at moderate or high levels of MRD, but increased markedly as the level of MRD and the number of intact marker gene rearrangements in the sample fell. In 23 children with ALL studied after induction therapy, the mean MRD level was 1.6 x 10^–5 and levels ranged from 1.5 x 10^–2 to less than 10^–7. Comparisons with the conventional one-round quantitative polymerase chain reaction method on 29 samples from another 24 children who received treatment resulted in concordant results for 22 samples and discordant results for seven samples. The sensitivity and specificity of the method are due to the use of nested polymerase chain reaction, one segment-specific and two allele-specific oligonucleotide primers, and the use of a large amount of good quality DNA. This method may improve MRD-based decisions on treatment for ALL patients, and the principles should be applicable to DNA-based MRD measurements in other disorders.

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				M. J. Brisco, S. Latham, R. Sutton, E. Hughes, V. Wilczek, K. van Zanten, B. Budgen, A. Y. Bahar, M. Malec, P. J. Sykes, et al. Determining the Repertoire of IGH Gene Rearrangements to Develop Molecular Markers for Minimal Residual Disease in B-Lineage Acute Lymphoblastic Leukemia J. Mol. Diagn., May 1, 2009; 11(3): 194 - 200. [Abstract] [Full Text] [PDF]