Abstract
Minimal residual disease (MRD) is a powerful prognostic factor in acute lymphoblastic leukemia (ALL) and is used for patient stratification and treatment decisions, but its precise role in Philadelphia chromosome positive ALL is less clear. This uncertainty results largely from methodological differences relating to the use of real-time quantitative PCR (qRT-PCR) to measure BCR-ABL1 transcript levels for MRD analysis. We here describe the first results by the EURO-MRD consortium on standardization of qRT-PCR for the e1a2 BCR-ABL1 transcript in Ph + ALL, designed to overcome the lack of standardisation of laboratory procedures and data interpretation. Standardised use of EAC primer/probe sets and of centrally prepared plasmid standards had the greatest impact on reducing interlaboratory variability. In QC1 the proportion of analyses with BCR-ABL1/ABL1 ratios within half a log difference were 40/67 (60%) and 52/67 (78%) at 10−3 and 36/67 (53%) and 53/67 (79%) at 10−4BCR-ABL1/ABL1. Standardized RNA extraction, cDNA synthesis and cycler platforms did not improve results further, whereas stringent application of technical criteria for assay quality and uniform criteria for data interpretation and reporting were essential. We provide detailed laboratory recommendations for the standardized MRD analysis in routine diagnostic settings and in multicenter clinical trials for Ph + ALL.
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14 January 2020
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
References
Bassan R, Spinelli O, Oldani E, Intermesoli T, Tosi M, Peruta B, et al. Improved risk classification for risk-specific therapy based on the molecular study of minimal residual disease (MRD) in adult acute lymphoblastic leukemia (ALL). Blood. 2009;113:4153–62.
Brisco J, Hughes E, Neoh SH, Sykes PJ, Bradstock K, Enno A, et al. Relationship between minimal residual disease and outcome in adult acute lymphoblastic leukemia. Blood. 1996;87:5251–6.
Bruggemann M, Raff T, Flohr T, Gokbuget N, Nakao M, Droese J, et al. Clinical significance of minimal residual disease quantification in adult patients with standard-risk acute lymphoblastic leukemia. Blood. 2006;107:1116–23.
Cave H, van der Werff ten Bosch J, Suciu S, Guidal C, Waterkeyn C, Otten J, et al. Clinical significance of minimal residual disease in childhood acute lymphoblastic leukemia. European Organization for Research and Treatment of Cancer—Childhood Leukemia Cooperative Group. N Engl J Med. 1998;339:591–8.
Conter V, Bartram CR, Valsecchi MG, Schrauder A, Panzer-Grumayer R, Moricke A, et al. Molecular response to treatment redefines all prognostic factors in children and adolescents with B-cell precursor acute lymphoblastic leukemia: results in 3184 patients of the AIEOP-BFM ALL 2000 study. Blood. 2010;115:3206–14.
Gokbuget N, Kneba M, Raff T, Trautmann H, Bartram CR, Arnold R, et al. Adult patients with acute lymphoblastic leukemia and molecular failure display a poor prognosis and are candidates for stem cell transplantation and targeted therapies. Blood. 2012;120:1868–76.
Mortuza FY, Papaioannou M, Moreira IM, Coyle LA, Gameiro P, Gandini D, et al. Minimal residual disease tests provide an independent predictor of clinical outcome in adult acute lymphoblastic leukemia. J Clin Oncol. 2002;20:1094–104.
van der Velden VH, Joosten SA, Willemse MJ, van Wering ER, Lankester AW, van Dongen JJ, et al. Real-time quantitative PCR for detection of minimal residual disease before allogeneic stem cell transplantation predicts outcome in children with acute lymphoblastic leukemia. Leukemia. 2001;15:1485–7.
Borowitz MJ, Pullen DJ, Shuster JJ, Viswanatha D, Montgomery K, Willman CL, et al. Minimal residual disease detection in childhood precursor-B-cell acute lymphoblastic leukemia: relation to other risk factors. a Children’s Oncology Group study. Leukemia. 2003;17:1566–72.
Conter V, Valsecchi MG, Parasole R, Putti MC, Locatelli F, Barisone E, et al. Childhood high-risk acute lymphoblastic leukemia in first remission: results after chemotherapy or transplant from the AIEOP ALL 2000 study. Blood. 2014;123:1470–8.
Bader P, Hancock J, Kreyenberg H, Goulden NJ, Niethammer D, Oakhill A, et al. Minimal residual disease (MRD) status prior to allogeneic stem cell transplantation is a powerful predictor for post-transplant outcome in children with ALL. Leukemia. 2002;16:1668–72.
Topp MS, Kufer P, Gokbuget N, Goebeler M, Klinger M, Neumann S, et al. Targeted therapy with the T-cell-engaging antibody blinatumomab of chemotherapy-refractory minimal residual disease in B-lineage acute lymphoblastic leukemia patients results in high response rate and prolonged leukemia-free survival. J Clin Oncol. 2011;29:2493–8.
Pongers-Willemse MJ, Seriu T, Stolz F, d’Aniello E, Gameiro P, Pisa P, et al. Primers and protocols for standardized detection of minimal residual disease in acute lymphoblastic leukemia using immunoglobulin and T cell receptor gene rearrangements and TAL1 deletions as PCR targets: report of the BIOMED-1 CONCERTED ACTION: investigation of minimal residual disease in acute leukemia. Leukemia. 1999;13:110–8.
van der Velden VH, Panzer-Grumayer ER, Cazzaniga G, Flohr T, Sutton R, Schrauder A, et al. Optimization of PCR-based minimal residual disease diagnostics for childhood acute lymphoblastic leukemia in a multi-center setting. Leukemia. 2007;21:706–13.
Bruggemann M, Schrauder A, Raff T, Pfeifer H, Dworzak M, Ottmann OG, et al. Standardized MRD quantification in European ALL trials: proceedings of the Second International Symposium on MRD assessment in Kiel, Germany, 18-20 September 2008. Leukemia. 2010;24:521–35.
Gabert J, Beillard E, van der Velden VH, Bi W, Grimwade D, Pallisgaard N, et al. Standardization and quality control studies of ‘real-time’ quantitative reverse transcriptase polymerase chain reaction of fusion gene transcripts for residual disease detection in leukemia - a Europe Against Cancer program. Leukemia. 2003;17:2318–57.
van der Velden VH, Cazzaniga G, Schrauder A, Hancock J, Bader P, Panzer-Grumayer ER, et al. Analysis of minimal residual disease by Ig/TCR gene rearrangements: guidelines for interpretation of real-time quantitative PCR data. Leukemia. 2007;21:604–11.
van der Velden VH, Hochhaus A, Cazzaniga G, Szczepanski T, Gabert J, van Dongen JJ. Detection of minimal residual disease in hematologic malignancies by real-time quantitative PCR: principles, approaches, and laboratory aspects. Leukemia. 2003;17:1013–34.
van Dongen JJ, van der Velden VH, Bruggemann M, Orfao A. Minimal residual disease diagnostics in acute lymphoblastic leukemia: need for sensitive, fast, and standardized technologies. Blood. 2015;125:3996–4009.
Chalandon Y, Thomas X, Hayette S, Cayuela JM, Abbal C, Huguet F, et al. Randomized study of reduced-intensity chemotherapy combined with imatinib in adults with Ph-positive acute lymphoblastic leukemia. Blood. 2015;125:3711–9.
Foa R, Vitale A, Vignetti M, Meloni G, Guarini A, De Propris MS, et al. Dasatinib as first-line treatment for adult patients with Philadelphia chromosome-positive acute lymphoblastic leukemia. Blood. 2011;118:6521–8.
Lee S, Kim DW, Cho BS, Yoon JH, Shin SH, Yahng SA, et al. Impact of minimal residual disease kinetics during imatinib-based treatment on transplantation outcome in Philadelphia chromosome-positive acute lymphoblastic leukemia. Leukemia. 2012;26:2367–74.
Ravandi F, Jorgensen JL, Thomas DA, O’Brien S, Garris R, Faderl S, et al. Detection of MRD may predict the outcome of patients with Philadelphia chromosome-positive ALL treated with tyrosine kinase inhibitors plus chemotherapy. Blood. 2013;122:1214–21.
Rousselot P, Coude MM, Gokbuget N, Gambacorti Passerini C, Hayette S, Cayuela JM, et al. Dasatinib and low-intensity chemotherapy in elderly patients with Philadelphia chromosome-positive ALL. Blood. 2016;128:774–82.
Thomas DA, Faderl S, Cortes J, O’Brien S, Giles FJ, Kornblau SM, et al. Treatment of Philadelphia chromosome-positive acute lymphocytic leukemia with hyper-CVAD and imatinib mesylate. Blood. 2004;103:4396–407.
Wassmann B, Pfeifer H, Goekbuget N, Beelen DW, Beck J, Stelljes M, et al. Alternating versus concurrent schedules of imatinib and chemotherapy as front-line therapy for Philadelphia-positive acute lymphoblastic leukemia (Ph+ALL). Blood. 2006;108:1469–77.
Schrappe M. Minimal residual disease: optimal methods, timing, and clinical relevance for an individual patient. Hematol Am Soc Hematol Educ Program. 2012;2012:137–42.
van der Velden VH, Boeckx N, van Wering ER, van Dongen JJ. Detection of minimal residual disease in acute leukemia. J Biol Regul Homeost Agents. 2004;18:146–54.
van Dongen JJ, Macintyre EA, Gabert JA, Delabesse E, Rossi V, Saglio G, et al. Standardized RT-PCR analysis of fusion gene transcripts from chromosome aberrations in acute leukemia for detection of minimal residual disease. Report of the BIOMED-1 Concerted Action: investigation of minimal residual disease in acute leukemia. Leukemia. 1999;13:1901–28.
Beillard E, Pallisgaard N, van der Velden VH, Bi W, Dee R, van der Schoot E, et al. Evaluation of candidate control genes for diagnosis and residual disease detection in leukemic patients using ‘real-time’ quantitative reverse-transcriptase polymerase chain reaction (RQ-PCR) - a Europe against cancer program. Leukemia. 2003;17:2474–86.
Branford S, Cross NC, Hochhaus A, Radich J, Saglio G, Kaeda J, et al. Rationale for the recommendations for harmonizing current methodology for detecting BCR-ABL transcripts in patients with chronic myeloid leukaemia. Leukemia. 2006;20:1925–30.
Hughes T, Deininger M, Hochhaus A, Branford S, Radich J, Kaeda J, et al. Monitoring CML patients responding to treatment with tyrosine kinase inhibitors: review and recommendations for harmonizing current methodology for detecting BCR-ABL transcripts and kinase domain mutations and for expressing results. Blood. 2006;108:28–37.
Ottmann OG, Pfeifer H. Management of Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph+ALL). Hematology Am Soc Hematol Educ Program 2009:371–81.
De Klein A, Hagemeijer A, Bartram CR, Houwen R, Hoefsloot L, Carbonell F, et al. bcr rearrangement and translocation of the c-abl oncogene in Philadelphia positive acute lymphoblastic leukemia. Blood. 1986;68:1369–75.
Biondi A, Schrappe M, De Lorenzo P, Castor A, Lucchini G, Gandemer V, et al. Imatinib after induction for treatment of children and adolescents with Philadelphia-chromosome-positive acute lymphoblastic leukaemia (EsPhALL): a randomised, open-label, intergroup study. Lancet Oncol. 2012;13:936–45.
Lee S, Kim YJ, Chung NG, Lim J, Lee DG, Kim HJ, et al. The extent of minimal residual disease reduction after the first 4-week imatinib therapy determines outcome of allogeneic stem cell transplantation in adults with Philadelphia chromosome-positive acute lymphoblastic leukemia. Cancer . 2009;115:561–70.
Pfeifer H, Wassmann B, Bethge W, Dengler J, Bornhauser M, Stadler M, et al. Randomized comparison of prophylactic and minimal residual disease-triggered imatinib after allogeneic stem cell transplantation for BCR-ABL1-positive acute lymphoblastic leukemia. Leukemia. 2013;27:1254–62.
Wassmann B, Pfeifer H, Stadler M, Bornhauser M, Bug G, Scheuring UJ, et al. Early molecular response to posttransplantation imatinib determines outcome in MRD+Philadelphia-positive acute lymphoblastic leukemia (Ph+ALL). Blood. 2005;106:458–63.
Yanada M, Sugiura I, Takeuchi J, Akiyama H, Maruta A, Ueda Y, et al. Prospective monitoring of BCR-ABL1 transcript levels in patients with Philadelphia chromosome-positive acute lymphoblastic leukaemia undergoing imatinib-combined chemotherapy. Br J Haematol. 2008;143:503–10.
Bassan R, Rossi G, Pogliani EM, Di Bona E, Angelucci E, Cavattoni I, et al. Chemotherapy-phased imatinib pulses improve long-term outcome of adult patients with Philadelphia chromosome-positive acute lymphoblastic leukemia: Northern Italy Leukemia Group protocol 09/00. J Clin Oncol. 2010;28:3644–52.
Radich J, Gehly G, Lee A, Avery R, Bryant E, Edmands S, et al. Detection of bcr-abl transcripts in Philadelphia chromosome-positive acute lymphoblastic leukemia after marrow transplantation. Blood. 1997;89:2602–9.
Zaliova M, Fronkova E, Krejcikova K, Muzikova K, Mejstrikova E, Stary J, et al. Quantification of fusion transcript reveals a subgroup with distinct biological properties and predicts relapse in BCR/ABL-positive ALL: implications for residual disease monitoring. Leukemia. 2009;23:944–51.
Cross NC, White HE, Ernst T, Welden L, Dietz C, Saglio G, et al. Development and evaluation of a secondary reference panel for BCR-ABL1 quantification on the International Scale. Leukemia. 2016;30:1844–52.
White H, Deprez L, Corbisier P, Hall V, Lin F, Mazoua S, et al. A certified plasmid reference material for the standardisation of BCR-ABL1 mRNA quantification by real-time quantitative PCR. Leukemia. 2015;29:369–76.
Branford S, Fletcher L, Cross NC, Muller MC, Hochhaus A, Kim DW, et al. Desirable performance characteristics for BCR-ABL measurement on an international reporting scale to allow consistent interpretation of individual patient response and comparison of response rates between clinical trials. Blood. 2008;112:3330–8.
Cross NC, Hochhaus A, Muller MC. Molecular monitoring of chronic myeloid leukemia: principles and interlaboratory standardization. Ann Hematol. 2015;94(Suppl 2):S219–25.
Bruggemann M, Kotrova M. Minimal residual disease in adult ALL: technical aspects and implications for correct clinical interpretation. Blood Adv. 2017;1:2456–66.
Scheuring UJ, Pfeifer H, Wassmann B, Bruck P, Gehrke B, Petershofen EK, et al. Serial minimal residual disease (MRD) analysis as a predictor of response duration in Philadelphia-positive acute lymphoblastic leukemia (Ph+ALL) during imatinib treatment. Leukemia. 2003;17:1700–6.
Lussana F, Intermesoli T, Gianni F, Boschini C, Masciulli A, Spinelli O, et al. Achieving Molecular Remission before Allogeneic Stem Cell Transplantation in Adult Patients with Philadelphia Chromosome-Positive Acute Lymphoblastic Leukemia: Impact on Relapse and Long-Term Outcome. Biol Blood Marrow Transplant. 2016;22:1983–7.
Nishiwaki S, Imai K, Mizuta S, Kanamori H, Ohashi K, Fukuda T, et al. Impact of MRD and TKI on allogeneic hematopoietic cell transplantation for Ph+ALL: a study from the adult ALL WG of the JSHCT. Bone Marrow Transplant. 2016;51:43–50.
van der Velden VH, Jacobs DC, Wijkhuijs AJ, Comans-Bitter WM, Willemse MJ, Hahlen K, et al. Minimal residual disease levels in bone marrow and peripheral blood are comparable in children with T cell acute lymphoblastic leukemia (ALL), but not in precursor-B-ALL. Leukemia. 2002;16:1432–6.
Cave H, Guidal C, Rohrlich P, Delfau MH, Broyart A, Lescoeur B, et al. Prospective monitoring and quantitation of residual blasts in childhood acute lymphoblastic leukemia by polymerase chain reaction study of delta and gamma T-cell receptor genes. Blood. 1994;83:1892–902.
Seriu T, Yokota S, Nakao M, Misawa S, Takaue Y, Koizumi S, et al. Prospective monitoring of minimal residual disease during the course of chemotherapy in patients with acute lymphoblastic leukemia, and detection of contaminating tumor cells in peripheral blood stem cells for autotransplantation. Leukemia. 1995;9:615–23.
van Dongen JJ, Seriu T, Panzer-Grumayer ER, Biondi A, Pongers-Willemse MJ, Corral L, et al. Prognostic value of minimal residual disease in acute lymphoblastic leukaemia in childhood. Lancet. 1998;352:1731–8.
Coustan-Smith E, Sancho J, Hancock ML, Razzouk BI, Ribeiro RC, Rivera GK, et al. Use of peripheral blood instead of bone marrow to monitor residual disease in children with acute lymphoblastic leukemia. Blood. 2002;100:2399–402.
van der Velden VH, Boeckx N, Gonzalez M, Malec M, Barbany G, Lion T, et al. Differential stability of control gene and fusion gene transcripts over time may hamper accurate quantification of minimal residual disease—a study within the Europe Against Cancer Program. Leukemia. 2004;18:884–6.
Acknowledgements
We are grateful to the teams of technicians, particularly Brigitte Gehrke, Doreen Badowski, Gabriele Lippok from the coordinating lab for their excellent work. This study was supported financially by the Deutsche José Carreras Leukämie-Stiftung e.V. (Förderkennzeichen: R09/11 and R12/01). OGO held a Stiftungsprofessur für Molekulare Therapieforschung der Deutschen José Carreras Leukämie Stiftung.
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HP, OGO, GC, VHJvdV, JMC, JJMvD, OO designed the study, analyzed data, wrote the manuscript; HP,GC, VHJ.vdV., JMC, BS, OS, SAk, SAv, IB, KB, HC, LE, JG-F, GG., SH, MH, AJ, TJ, MCC, CH, II, VK., TLa, TLi, MCM, FP, LR, CDW, TS, SS, TT, HV, P.VdB., JZ, HS, SM performed the molecular analyses, collected data, HP, EH performed the statistical analysis.
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VHJvdV: contract research for Amgen, Roche, Pfizer, Janssen and BD Biosciences; consultancy fees from Celgene. JJMvD: contract research for Roche, Amgen and BD Biosciences. HP: research support from Ipsogen. The other authors declare that they have no conflict of interest.
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Pfeifer, H., Cazzaniga, G., van der Velden, V.H.J. et al. Standardisation and consensus guidelines for minimal residual disease assessment in Philadelphia-positive acute lymphoblastic leukemia (Ph + ALL) by real-time quantitative reverse transcriptase PCR of e1a2 BCR-ABL1. Leukemia 33, 1910–1922 (2019). https://doi.org/10.1038/s41375-019-0413-0
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DOI: https://doi.org/10.1038/s41375-019-0413-0
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