Detection of minimal residual disease in acute lymphoblastic leukemia

Detection of minimal residual disease in acute lymphoblastic leukemia


Dario Campana

Correspondence: D. Campana, M.D. Ph.D., Department of Oncology, St. Jude Children's Research Hospital, 262 Danny Thomas Place, Memphis TN 38105, USA.

This work was supported by grants CA60419 and CA21765 from the National Cancer Institute, and by the American Lebanese Syrian Associated Charities (ALSAC).

July 2009



I. Introduction
In patients with acute lymphoblastic leukemia (ALL), the degree of treatment response guides clinical decisions, and information about this response is essential for selecting the optimal clinical management approach. Unfortunately, determining whether residual leukemia is present during treatment by traditional methods, i.e. the morphologic examination of cells in bone marrow smears, is typically a subjective and imprecise endeavor owing to the fact that the morphology of ALL cells is very similar to that of normal bone marrow cell subpopulations, such as immature B cells and activated mature lymphocytes. Hence, the remission status of patients with ALL often raises doubt in the mind of pathologists and clinicians; this uncertainty can lead to overtreatment (and excessive toxicities) or undertreatment (and increased risk of relapse). The advent of methods for detecting minimal residual disease (MRD) has revealed that many patients considered to be in "remission" by morphologic analysis still have substantial amounts of residual leukemia (Campana, 2008a). Because of the strong correlation between MRD levels and treatment outcome, MRD testing is increasingly being incorporated in clinical trials.

II. A brief review of methods for MRD detection

Polymerase chain reaction
Two main types of molecular targets can be used to identify leukemic cells. One is represented by clonally rearranged antigen-receptor genes, i.e, immunoglobulin (IG) and T-cell receptor (TCR) genes. The junctional regions of the rearranged genes are unique to the leukemic clone. Typically, the unique gene signature is identified at diagnosis in each case using PCR primers matched to the V and J regions of various IG and TCR genes. If a rearrangement is found, the PCR product is further analyzed to ensure its clonal origin by using heteroduplex analysis (van der Velden et al., 2007). The junctional regions of the IG/TCR gene rearrangements are then sequenced to design specific oligonucleotides which are then applied to monitor MRD (van der Velden et al., 2007). Investigators have developed methods to detect clonal IG/TCR gene rearrangements without the need for patient-specific oligonucleotides. These efforts have relied on high-resolution electrophoresis, such as radioactive fingerprinting or fluorescent gene scanning, but this approach has a considerably lower sensitivity, usually not better than 0.1%, and date interpretation may be difficult (Delabesse et al., 2000; Knechtli et al., 1998).
Because the majority of B-lineage ALL cases have IG (Beishuizen et al., 1993) and cross-lineage TCR gene rearrangements (Szczepanski et al., 1999a), MRD monitoring by using these genes as targets is feasible in > 90% of cases of B-lineage ALL. Likewise, TCR genes are rearranged in most cases of T-lineage ALL and cross-lineage IG gene rearrangements occur in approximately 20% of T-ALL (Szczepanski et al., 2000; Kneba et al., 1995). In sum, the method can be used to monitor MRD in most cases of childhood and adult ALL (van der Velden et al., 2003; van der Velden et al., 2007; Bruggemann et al., 2006; Flohr et al., 2008).
Detection of MRD by PCR using IG/TCR gene rearrangements is most frequently performed by using "real-time" quantitative PCR (RQ-PCR) (van der Velden et al., 2003) and less commonly by limiting dilution (Neale et al., 1999). Because rearranged IG and TCR genes are present in one copy per cells, very precise estimates of the MRD levels can be achieved. IG and TCR genes may be affected by continuing or secondary rearrangements (Szczepanski et al., 1999b), resulting in subclones with distinct clonal IG/TCR gene rearrangements, and minor clones at diagnosis may become predominant at relapse (Szczepanski et al., 2002; van der Velden et al., 2004). These possibilities have prompted the recommendation of targeting two or more different rearrangements during MRD studies (van der Velden et al., 2007). Multiple targets are identifiable in the majority of ALL cases although in approximately 30% of cases it is not possible to identify multiple targets that allow detection of MRD with a high sensitivity (e.g., 0.01%) (Pongers-Willemse et al., 1999; Flohr et al., 2008).
The second type of gene target for MRD monitoring by PCR is represented by gene fusions, such as BCR-ABL1, MLL-AFF1, TCF3-PBX1, and ETV6-RUNX1, and their resulting aberrant mRNA transcripts (van Dongen et al., 1999; Gabert et al., 2003). Recurrent fusions are identified in less than half of patients with newly diagnosed ALL (Gabert et al., 2003), thus limiting the applicability of this approach. However, with the systematic use of novel whole-genome screening technologies (Mullighan et al., 2007; Mullighan et al., 2009), it is very likely that additional genetic targets will enrich the available array of gene targets for MRD studies.
One potential advantage of using fusion transcripts to monitor MRD is that it might be possible to detect pre-leukemic cells (Hong et al., 2008). If so, the clinical significance of such finding needs to be investigated. A clear disadvantage of using fusion transcripts as targets is an accurate estimate of the number of leukemic cells present in the sample is difficult. This is because that ratio between amount of PCR product and target cell number is uncertain, there may be interpatient variability in the number of transcripts per leukemic cell within the same genetic subtype of ALL, and this number could be altered by chemotherapy (Gabert et al., 2003).

Flow cytometry
Immunophenotypes characteristic of leukemic cells can be used to distinguish ALL from normal cells by flow cytometry (Campana, 2008). There are three main categories of leukemia-associate immunophenotypes. One is characterized by the expression of fusion proteins derived from fusion transcripts, such as BCR-ABL1, ETV6-RUNX1, or TCF3-PBX1. However, suitable antibodies for reliable flow cytometric analysis of these proteins are lacking. A second group is represented by the immunophenotype of T-lineage ALL cells, which is normally expressed only by a subset of thymocytes and it is not expressed by cells outside the thymus. Immature T-cell phenotypes can be effectively used to monitor MRD in T-lineage ALL (Coustan-Smith et al., 2002a), and also to detect disease dissemination in T-cell lymphoblastic lymphoma (Coustan-Smith et al., 2009a). The third group of leukemia-associated immunophenotype is constituted by multiple marker combinations that are found in B-lineage ALL cells but are normally not expressed during lympho-hematopoiesis. The use of these immunophenotypes, named "asynchronous" or "aberrant" (Hurwitz et al., 1988; Lucio et al., 1999; Campana and Coustan-Smith, 1999; Ciudad et al., 1998), requires a particularly good knowledge of the immunophenotypes expressed by normal hematopoietic cells, in both normal and recovering bone marrow.
Leukemia-associated immunophenotypes that are suitable for MRD studies and afford a sensitivity of at least 0.01% can be identified in nearly all patients with ALL (Coustan-Smith et al., 2002b; Campana and Coustan-Smith, 1999). Results obtained by flow cytometry are very similar to those obtained by PCR amplification of IG/TCR genes, if MRD is present at a ≥ 0.01% level (Neale et al., 1999; Neale et al., 2004; Kerst et al., 2005).
Current methods for MRD testing by flow cytometry typically require the use of extensive antibody panels and considerable interpretative expertise. We developed a simplified flow cytometric MRD test that can detect residual B-lineage ALL cells (which express CD19 plus CD10 and/or CD34) on day 15-26 of treatment with a minimum panel of antibodies (Coustan-Smith et al., 2006). The rationale for this strategy is that normal immature CD19+ cells, or those expressing CD10 and/or CD34, are consistently undetectable in bone marrow samples collected from children with T-lineage ALL after 2 weeks of remission induction chemotherapy, because of their high sensitivity to glucocorticoids and other antileukemic drugs. We therefore reasoned that any cell with this immunophenotype detected in patients with B-lineage ALL on day 19 of induction treatment would likely be residual leukemic cells. Indeed, our findings indicate that the results of the simplified test correlate very well with those of more complex flow cytometric assays or PCR amplification of IGH/TCR genes. It should be stressed that this test cannot be used beyond this early treatment interval because of the high risk of false-positive results in recovering marrow samples.

III. Results of correlative studies with treatment outcome

Studies in pediatric ALL
The clinical significance of MRD testing during the initial phases of treatment was definitively demonstrated by 3 prospective studies published in 1998 by the EORTC (Cave et al., 1998), St Jude (Coustan-Smith et al., 1998) and BFM groups (van Dongen et al., 1998). The results these studies consolidated those of many other previous reports of smaller series, and have been confirmed by several subsequent studies (reviewed in Campana, 2009). MRD testing is also clinically informative for patients with specific ALL subtypes (Coustan-Smith et al., 2000; Biondi et al., 2000; Attarbaschi et al., 2008; van der Velden et al., 2009), patients with relapsed ALL who achieve a second remission (Eckert et al., 2001; Coustan-Smith et al., 2004; Paganin et al., 2008), patients with extramedullary relapse (Hagedorn et al., 2007) and patients undergoing allogeneic hematopoietic stem cell transplantation (Knechtli et al., 1998; van der Velden et al., 2001; Bader et al., 2002; Uzunel et al., 2001; Krejci et al., 2003; Goulden et al., 2003).
Levels of MRD are directly proportional to the risk of subsequent relapse. Thus, MRD ≥ 1% at the end of remission induction therapy predicted an extremely high rate of relapse in St Jude studies (Coustan-Smith et al., 2000), while MRD ≥ 0.1% on both day 33 and day 78 of treatment had a very high risk of relapse in the I-BFM Study Group studies (van Dongen et al., 1998; Flohr et al., 2008). The threshold level commonly used to define MRD positivity is 0.01% of bone marrow mononuclear cells. Patients with ≥ 0.01% MRD at any time point during treatment had a higher risk of relapse in earlier St Jude studies (Coustan-Smith et al., 1998; Coustan-Smith et al., 2000; Coustan-Smith et al., 2002b), as had those with ≥ 0.01% MRD on day 29 of treatment in studies of the Children's Oncology Group (Borowitz et al., 2008). In other studies, however, a threshold of 0.1% appeared to be more informative (Cave et al., 1998; Dworzak et al., 2002; Zhou et al., 2007).
In addition to providing a parameter to identify patients at a higher risk of relapse, MRD can also identify patients with excellent early treatment response and undetectable (< 0.01%) MRD after 2-3 weeks of therapy. We found that 183 of 402 (45.5%) B-lineage ALL patients were MRD < 0.01% on day 19 of treatment (Campana, 2008b), a feature that is associated with excellent prognosis overall (Panzer-Grumayer et al., 2000; Coustan-Smith et al., 2002b).
The prevalence of MRD differs among different genetic subtypes of childhood ALL (Pui et al., 2001; Borowitz et al., 2003). Thus, MRD is generally more prevalent among patients with BCR-ABL1 ALL and less prevalent among those with ETV6-RUNX1, hyperdiploid (> 50 chromosomes) and TCF3-PBX1 ALL (Campana, 2008c). More recently, it has been shown that patients with B-lineage ALL and mutations or deletions of the Ikaros (lIKZF1) gene had a higher prevalence of MRD during remission induction therapy than those without this abnormality (Mullighan et al., 2009). In addition, among patients with T-lineage ALL, MRD-positive findings were strikingly more frequent and levels higher in the subgroup of patients with early thymic precursor (ETP)-ALL (Coustan-Smith et al., 2009b).
MRD studies have now been included in clinical trials to guide therapy. Thus, the AIEOP-BFM group uses MRD to classify patients with newly diagnosed ALL into three risk groups: standard risk (MRD negative on days 33 and 78), intermediate risk (any MRD positivity on days 33 and 78 but < 0.1% on day 78) and high risk (MRD ≥ 0.1% on day 78) (Flohr et al., 2008). In the AIEOP-BFM ALL 2000 trial, of the 3341 diagnostic samples examined, 88 (3%) lacked suitable gene rearrangements targets for PCR analysis, and an additional 217 (7%) had a target but not sufficient to reach a sensitivity of 0.01% (Flohr et al., 2008). At least two sensitive gene rearrangement targets could be identified in 71% of patients. Adequate data for MRD-based stratification were obtained in 2594 (78%) of the 3341 patients (78%).
In the St Jude Total XV trial for children with newly diagnosed ALL, our laboratory monitored MRD by using flow cytometric detection of aberrant immunophenotypes and/or PCR amplification of antigen-receptor genes (Pui et al., 2009). Overall, 482 of 492 patients (98%) were monitored by flow cytometry and 403 of 492 (82%) by PCR (applied only to patients with B-lineage ALL). As previously shown (Neale et al., 1999; Neale et al., 2004; Kerst et al., 2005), both methods yielded virtually identical results above the threshold level of 0.01%. The two methods in combination could be applied to study 491 of 492 patients (99.8%) (Pui et al., 2009). The single patient with no available immunophenotypic or antigen-receptor gene rearrangements had a MLL-AF9 fusion transcript and was monitored by RQ-PCR using that marker. In our current Total XVI trial, patients with MRD ≥ 1% on day 15 receive intensified remission induction therapy; further intensification is reserved for patients with ≥ 5% leukemic cells. By contrast, patients with MRD < 0.01% on day 15 receive less intensive reinduction therapy and lower cumulative doses of anthracyclin. Patients with standard-risk ALL who have MRD of ≥ 0.01% on day 42 are reclassified as high-risk; patients with MRD ≥ 1% are eligible for transplant in first remission. Because in patients with T-lineage ALL MRD levels in peripheral blood are similar to those in bone marrow (Coustan-Smith et al., 2002a; van der Velden et al., 2002), it is our current practice to use blood instead of marrow to monitor MRD after day 42 in these patients.

Studies in adult ALL
Several studies have also demonstrated the prognostic importance of MRD in adult ALL patients (Mortuza et al., 2002; Bruggemann et al., 2006; Raff et al., 2007; Holowiecki et al., 2008; Bassan et al., 2009). Bruggeman et al. (Bruggemann et al., 2006) studied MRD in 196 standard-risk patients using PCR amplification of antigen-receptor genes and segregated three groups: 10% of patients had < 0.01% MRD on days 11 and 24 of treatment and 23% had persistent MRD ≥ 0.01% until week 16. The 3-year relapse rates were 0% and 94%; for the remaining patients, the relapse rate was 47%. The same group subsequently studied post-consolidation samples from 105 patients who were in hematologic remission, had completed the first-year chemotherapy, and were MRD-negative before enrolling in the study. MRD was detected in 28 patients, 17 of whom relapsed. By contrast, 77 patients remained MRD-negative and only 5 relapsed (Raff et al., 2007). Using IG/TCR gene rearrangements or fusion transcripts as targets, Bassan et al. (Bassan et al., 2009) measured MRD at the end of consolidation. Five-year overall disease-free survival estimates were 72% among 58 MRD negative patients and 14% among the 54 patients with positive MRD. In a study using flow cytometry, Holowiecki et al. (Holowiecki et al., 2008) measured MRD in 116 patients with Philadelphia-negative ALL and found that MRD ≥ 0.1% after remission induction therapy was an independent predictor for relapse. Together, the results of these studies provide convincing evidence of the clinical significance of MRD in adult ALL, although the strengths of the correlations with outcome depend on the subgroup of patients studied and the type of treatment.
Monitoring of MRD in adult patients with Philadelphia-positive ALL receiving transplant and/or imatinib therapy has been shown to predict treatment outcome (Radich et al., 1997; Wassmann et al., 2005; Pane et al., 2005). It has been shown that MRD detected before initiation of conditioning is a significant predictor of failure post-transplant (Sanchez et al., 2002; Spinelli et al., 2007).

Areas for further research
Measuring MRD provides unprecedented insights into the kinetics of treatment response in patients with acute leukemia which not only have prognostic ramifications but can also provide novel endpoints for correlative studies with cellular and biologic features. For example, the correlation between MRD and gene expression of leukemic lymphoblasts at diagnosis revealed genes associated with treatment response (Cario et al., 2005; Flotho et al., 2006; Flotho et al., 2007), while correlations with gene polymorphisms has pointed to drug-metabolizing molecules which may have a direct impact on leukemia response to treatment (Rocha et al., 2005; Yang et al., 2009). These areas are clearly worthy of further research, which may lead to the identification of new prognostic factors and provide clues about targets for molecular therapies.
Although MRD can be studied in virtually all patients with ALL using molecular and/or flow cytometric methods, MRD assays require considerable expertise and can be performed well only in specialized centers. Simplification of the methodologies to widen the applicability of MRD testing should be an objective for future research. At the same time, increasingly sophisticated methodologies provide new opportunities for investigation. To this end, the availability of reliable flow cytometers that can detect 6 or more fluorochromes together with the a wide array of commercial antibodies open the possibility to investigate the biologic features of the leukemic cells that contribute to MRD in extraordinary detail. In turn, such studies should help unearthing some of the biologic roots of drug resistance in ALL and ultimately lead to more effective and less toxic treatment.


Asynchronous antigen expression in B lineage acute lymphoblastic leukemia.
Hurwitz CA, Loken MR, Graham ML, Karp JE, Borowitz MJ, Pullen DJ, Civin CI.
Blood. 1988 Jul;72(1):299-307.
PMID 3291983
Detection of immunoglobulin heavy-chain gene rearrangements by Southern blot analysis: recommendations for optimal results.
Beishuizen A, Verhoeven MA, Mol EJ, Breit TM, Wolvers-Tettero IL, van Dongen JJ.
Leukemia. 1993 Dec;7(12):2045-53.
PMID 7902888
Analysis of rearranged T-cell receptor beta-chain genes by polymerase chain reaction (PCR) DNA sequencing and automated high resolution PCR fragment analysis.
Kneba M, Bolz I, Linke B, Hiddemann W.
Blood. 1995 Nov 15;86(10):3930-7.
PMID 7579363
Detection of bcr-abl transcripts in Philadelphia chromosome-positive acute lymphoblastic leukemia after marrow transplantation.
Radich J, Gehly G, Lee A, Avery R, Bryant E, Edmands S, Gooley T, Kessler P, Kirk J, Ladne P, Thomas ED, Appelbaum FR.
Blood. 1997 Apr 1;89(7):2602-9.
PMID 9116308
Clinical significance of minimal residual disease in childhood acute lymphoblastic leukemia. European Organization for Research and Treatment of Cancer--Childhood Leukemia Cooperative Group.
Cave H, van der Werff ten Bosch J, Suciu S, Guidal C, Waterkeyn C, Otten J, Bakkus M, Thielemans K, Grandchamp B, Vilmer E.
N Engl J Med. 1998 Aug 27;339(9):591-8.
PMID 9718378
Prognostic value of immunophenotypic detection of minimal residual disease in acute lymphoblastic leukemia.
Ciudad J, San Miguel JF, Lopez-Berges MC, Vidriales B, Valverde B, Ocqueteau M, Mateos G, Caballero MD, Hernandez J, Moro MJ, Mateos MV, Orfao A.
J Clin Oncol. 1998 Dec;16(12):3774-81.
PMID 9850021
Immunological detection of minimal residual disease in children with acute lymphoblastic leukaemia.
Coustan-Smith E, Behm FG, Sanchez J, Boyett JM, Hancock ML, Raimondi SC, Rubnitz JE, Rivera GK, Sandlund JT, Pui CH, Campana D.
Lancet. 1998 Feb 21;351(9102):550-4.
PMID 9492773
Minimal residual disease status before allogeneic bone marrow transplantation is an important determinant of successful outcome for children and adolescents with acute lymphoblastic leukemia.
Knechtli CJ, Goulden NJ, Hancock JP, Grandage VL, Harris EL, Garland RJ, Jones CG, Rowbottom AW, Hunt LP, Green AF, Clarke E, Lankester AW, Cornish JM, Pamphilon DH, Steward CG, Oakhill A.
Blood. 1998 Dec 1;92(11):4072-9.
PMID 9834212
Prognostic value of minimal residual disease in acute lymphoblastic leukaemia in childhood.
van Dongen JJ, Seriu T, Panzer-Grumayer ER, Biondi A, Pongers-Willemse MJ, Corral L, Stolz F, Schrappe M, Masera G, Kamps WA, Gadner H, van Wering ER, Ludwig WD, Basso G, de Bruijn MA, Cazzaniga G, Hettinger K, van der Does-van den Berg A, Hop WC, Riehm H, Bartram CR.
Lancet. 1998 Nov 28;352(9142):1731-8.
PMID 9848348
Detection of minimal residual disease in acute leukemia by flow cytometry.
Campana D, Coustan-Smith E.
Cytometry. 1999 Aug 15;38(4):139-52.
PMID 10440852
Flow cytometric analysis of normal B cell differentiation: a frame of reference for the detection of minimal residual disease in precursor-B-ALL.
Lucio P, Parreira A, van den Beemd MW, van Lochem EG, van Wering ER, Baars E, Porwit-MacDonald A, Bjorklund E, Gaipa G, Biondi A, Orfao A, Janossy G, van Dongen JJ, San Miguel JF.
Leukemia. 1999 Mar;13(3):419-27.
PMID 10086733
Tandem application of flow cytometry and polymerase chain reaction for comprehensive detection of minimal residual disease in childhood acute lymphoblastic leukemia.
Neale GA, Coustan-Smith E, Pan Q, Chen X, Gruhn B, Stow P, Behm FG, Pui CH, Campana D.
Leukemia. 1999 Aug;13(8):1221-6.
PMID 10450750
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.
Pongers-Willemse MJ, Seriu T, Stolz F, d'Aniello E, Gameiro P, Pisa P, Gonzalez M, Bartram CR, Panzer-Grumayer ER, Biondi A, San Miguel JF, van Dongen JJ.
Leukemia. 1999 Jan;13(1):110-8.
PMID 10049045
Cross-lineage T cell receptor gene rearrangements occur in more than ninety percent of childhood precursor-B acute lymphoblastic leukemias: alternative PCR targets for detection of minimal residual disease.
Szczepanski T, Beishuizen A, Pongers-Willemse MJ, Hahlen K, Van Wering ER, Wijkhuijs AJ, Tibbe GJ, De Bruijn MA, Van Dongen JJ.
Leukemia. 1999a Feb;13(2):196-205.
PMID 10025893
Unusual immunoglobulin and T-cell receptor gene rearrangement patterns in acute lymphoblastic leukemias.
Szczepanski T, Pongers-Willemse MJ, Langerak AW, van Dongen JJ.
Curr Top Microbiol Immunol. 1999b;246:205-13
PMID 10396058
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.
van Dongen JJ, Macintyre EA, Gabert JA, Delabesse E, Rossi V, Saglio G, Gottardi E, Rambaldi A, Dotti G, Griesinger F, Parreira A, Gameiro P, Diaz MG, Malec M, Langerak AW, San Miguel JF, Biondi A.
Leukemia. 1999 Dec;13(12):1901-28.
PMID 10602411
Molecular detection of minimal residual disease is a strong predictive factor of relapse in childhood B-lineage acute lymphoblastic leukemia with medium risk features. A case control study of the International BFM study group.
Biondi A, Valsecchi MG, Seriu T, D'Aniello E, Willemse MJ, Fasching K, Pannunzio A, Gadner H, Schrappe M, Kamps WA, Bartram CR, van Dongen JJ, Panzer-Grumayer ER.
Leukemia. 2000 Nov;14(11):1939-43.
PMID 11069029
Rapid, multifluorescent TCRG Vgamma and Jgamma typing: application to T cell acute lymphoblastic leukemia and to the detection of minor clonal populations.
Delabesse E, Burtin ML, Millien C, Madonik A, Arnulf B, Beldjord K, Valensi F, Macintyre EA.
Leukemia. 2000 Jun;14(6):1143-52.
PMID 10865981
Rapid molecular response during early induction chemotherapy predicts a good outcome in childhood acute lymphoblastic leukemia.
Panzer-Grumayer ER, Schneider M, Panzer S, Fasching K, Gadner H.
Blood. 2000 Feb 1;95(3):790-4.
PMID 10648387
Clinical importance of minimal residual disease in childhood acute lymphoblastic leukemia.
Coustan-Smith E, Sancho J, Hancock ML, Boyett JM, Behm FG, Raimondi SC, Sandlund JT, Rivera GK, Rubnitz JE, Ribeiro RC, Pui CH, Campana D.
Blood. 2000 Oct 15;96(8):2691-6.
PMID 11023499
T cell receptor gamma (TCRG) gene rearrangements in T cell acute lymphoblastic leukemia refelct 'end-stage' recombinations: implications for minimal residual disease monitoring.
Szczepanski T, Langerak AW, Willemse MJ, Wolvers-Tettero IL, van Wering ER, van Dongen JJ.
Leukemia. 2000 Jul;14(7):1208-14.
PMID 10914544
Prognostic value of minimal residual disease in relapsed childhood acute lymphoblastic leukaemia.
Eckert C, Biondi A, Seeger K, Cazzaniga G, Hartmann R, Beyermann B, Pogodda M, Proba J, Henze G.
Lancet. 2001 Oct 13;358(9289):1239-41.
PMID 11675066
Childhood acute lymphoblastic leukaemia--current status and future perspectives.
Pui CH, Campana D, Evans WE.
Lancet Oncol. 2001 Oct;2(10):597-607.
PMID 11902549
The significance of graft-versus-host disease and pretransplantation minimal residual disease status to outcome after allogeneic stem cell transplantation in patients with acute lymphoblastic leukemia.
Uzunel M, Mattsson J, Jaksch M, Remberger M, Ringden O.
Blood. 2001 Sep 15;98(6):1982-4.
PMID 11535539
Real-time quantitative PCR for detection of minimal residual disease before allogeneic stem cell transplantation predicts outcome in children with acute lymphoblastic leukemia.
van der Velden VH, Joosten SA, Willemse MJ, van Wering ER, Lankester AW, van Dongen JJ, Hoogerbrugge PM.
Leukemia. 2001 Sep;15(9):1485-7.
PMID 11516112
Use of peripheral blood instead of bone marrow to monitor residual disease in children with acute lymphoblastic leukemia.
Coustan-Smith E, Sancho J, Hancock ML, Razzouk BI, Ribeiro RC, Rivera GK, Rubnitz JE, Sandlund JT, Pui CH, Campana D.
Blood. 2002a Oct 1;100(7):2399-402.
PMID 12239148
Prognostic importance of measuring early clearance of leukemic cells by flow cytometry in childhood acute lymphoblastic leukemia.
Coustan-Smith E, Sancho J, Behm FG, Hancock ML, Razzouk BI, Ribeiro RC, Rivera GK, Rubnitz JE, Sandlund JT, Pui CH, Campana D.
Blood. 2002b Jul 1;100(1):52-8.
PMID 12070008
Minimal residual disease (MRD) status prior to allogeneic stem cell transplantation is a powerful predictor for post-transplant outcome in children with ALL.
Bader P, Hancock J, Kreyenberg H, Goulden NJ, Niethammer D, Oakhill A, Steward CG, Handgretinger R, Beck JF, Klingebiel T.
Leukemia. 2002 Sep;16(9):1668-72.
PMID 12200679
Prognostic significance and modalities of flow cytometric minimal residual disease detection in childhood acute lymphoblastic leukemia.
Dworzak MN, Froschl G, Printz D, Mann G, Potschger U, Muhlegger N, Fritsch G, Gadner H; Austrian Berlin-Frankfurt-Munster Study Group.
Blood. 2002 Mar 15;99(6):1952-8.
PMID 11877265
Minimal residual disease tests provide an independent predictor of clinical outcome in adult acute lymphoblastic leukemia.
Mortuza FY, Papaioannou M, Moreira IM, Coyle LA, Gameiro P, Gandini D, Prentice HG, Goldstone A, Hoffbrand AV, Foroni L.
J Clin Oncol. 2002 Feb 15;20(4):1094-104.
PMID 11844835
Clinical value of immunological monitoring of minimal residual disease in acute lymphoblastic leukaemia after allogeneic transplantation.
Sanchez J, Serrano J, Gomez P, Martinez F, Martin C, Madero L, Herrera C, Garcia JM, Casano J, Torres A.
Br J Haematol. 2002 Mar;116(3):686-94.
PMID 11849234
Comparative analysis of Ig and TCR gene rearrangements at diagnosis and at relapse of childhood precursor-B-ALL provides improved strategies for selection of stable PCR targets for monitoring of minimal residual disease.
Szczepanski T, Willemse MJ, Brinkhof B, van Wering ER, van der Burg M, van Dongen JJ.
Blood. 2002 Apr 1;99(7):2315-23.
PMID 11895762
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.
van der Velden VH, Jacobs DC, Wijkhuijs AJ, Comans-Bitter WM, Willemse MJ, Hahlen K, Kamps WA, van Wering ER, van Dongen JJ.
Leukemia. 2002 Aug;16(8):1432-6.
PMID 12145681
Minimal residual disease detection in childhood precursor-B-cell acute lymphoblastic leukemia: relation to other risk factors. A Children's Oncology Group study.
Borowitz MJ, Pullen DJ, Shuster JJ, Viswanatha D, Montgomery K, Willman CL, Camitta B.
Leukemia. 2003 Aug;17(8):1566-72.
PMID 12886244
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.
Gabert J, Beillard E, van der Velden VH, Bi W, Grimwade D, Pallisgaard N, Barbany G, Cazzaniga G, Cayuela JM, Cave H, Pane F, Aerts JL, De Micheli D, Thirion X, Pradel V, Gonzalez M, Viehmann S, Malec M, Saglio G, van Dongen JJ.
Leukemia. 2003 Dec;17(12):2318-57.
PMID 14562125
Minimal residual disease prior to stem cell transplant for childhood acute lymphoblastic leukaemia.
Goulden N, Bader P, Van Der Velden V, Moppett J, Schilham M, Masden HO, Krejci O, Kreyenberg H, Lankester A, Revesz T, Klingebiel T, Van Dongen J.
Br J Haematol. 2003 Jul;122(1):24-9.
PMID 12823342
Level of minimal residual disease prior to haematopoietic stem cell transplantation predicts prognosis in paediatric patients with acute lymphoblastic leukaemia: a report of the Pre-BMT MRD Study Group.
Krejci O, van der Velden VH, Bader P, Kreyenberg H, Goulden N, Hancock J, Schilham MW, Lankester A, Revesz T, Klingebiel T, van Dongen JJ.
Bone Marrow Transplant. 2003 Oct;32(8):849-51.
PMID 14520434
Detection of minimal residual disease in hematologic malignancies by real-time quantitative PCR: principles, approaches, and laboratory aspects.
van der Velden VH, Hochhaus A, Cazzaniga G, Szczepanski T, Gabert J, van Dongen JJ.
Leukemia. 2003 Jun;17(6):1013-34.
PMID 12764363
Clinical significance of minimal residual disease in childhood acute lymphoblastic leukemia after first relapse.
Coustan-Smith E, Gajjar A, Hijiya N, Razzouk BI, Ribeiro RC, Rivera GK, Rubnitz JE, Sandlund JT, Andreansky M, Hancock ML, Pui CH, Campana D.
Leukemia. 2004 Mar;18(3):499-504.
PMID 14981525
Comparative analysis of flow cytometry and polymerase chain reaction for the detection of minimal residual disease in childhood acute lymphoblastic leukemia.
Neale GA, Coustan-Smith E, Stow P, Pan Q, Chen X, Pui CH, Campana D.
Leukemia. 2004 May;18(5):934-8.
PMID 15029212
TCRB gene rearrangements in childhood and adult precursor-B-ALL: frequency, applicability as MRD-PCR target, and stability between diagnosis and relapse.
van der Velden VH, Bruggemann M, Hoogeveen PG, de Bie M, Hart PG, Raff T, Pfeifer H, Luschen S, Szczepanski T, van Wering ER, Kneba M, van Dongen JJ.
Leukemia. 2004 Dec;18(12):1971-80.
PMID 15470492
Distinct gene expression profiles determine molecular treatment response in childhood acute lymphoblastic leukemia.
Cario G, Stanulla M, Fine BM, Teuffel O, Neuhoff NV, Schrauder A, Flohr T, Schafer BW, Bartram CR, Welte K, Schlegelberger B, Schrappe M.
Blood. 2005 Jan 15;105(2):821-6. Epub 2004 Sep 23.
PMID 15388585
Concurrent detection of minimal residual disease (MRD) in childhood acute lymphoblastic leukaemia by flow cytometry and real-time PCR.
Kerst G, Kreyenberg H, Roth C, Well C, Dietz K, Coustan-Smith E, Campana D, Koscielniak E, Niemeyer C, Schlegel PG, Muller I, Niethammer D, Bader P.
Br J Haematol. 2005 Mar;128(6):774-82.
PMID 15755280
Significant reduction of the hybrid BCR/ABL transcripts after induction and consolidation therapy is a powerful predictor of treatment response in adult Philadelphia-positive acute lymphoblastic leukemia.
Pane F, Cimino G, Izzo B, Camera A, Vitale A, Quintarelli C, Picardi M, Specchia G, Mancini M, Cuneo A, Mecucci C, Martinelli G, Saglio G, Rotoli B, Mandelli F, Salvatore F, Foa R.
Leukemia. 2005 Apr;19(4):628-35.
PMID 15744351
Pharmacogenetics of outcome in children with acute lymphoblastic leukemia.
Rocha JC, Cheng C, Liu W, Kishi S, Das S, Cook EH, Sandlund JT, Rubnitz J, Ribeiro R, Campana D, Pui CH, Evans WE, Relling MV.
Blood. 2005 Jun 15;105(12):4752-8. Epub 2005 Feb 15.
PMID 15713801
Early molecular response to posttransplantation imatinib determines outcome in MRD+ Philadelphia-positive acute lymphoblastic leukemia (Ph+ ALL).
Wassmann B, Pfeifer H, Stadler M, Bornhauser M, Bug G, Scheuring UJ, Bruck P, Stelljes M, Schwerdtfeger R, Basara N, Perz J, Bunjes D, Ledderose G, Mahlberg R, Binckebanck A, Gschaidmeier H, Hoelzer D, Ottmann OG.
Blood. 2005 Jul 15;106(2):458-63. Epub 2005 Apr 7.
PMID 15817679
A simplified flow cytometric assay identifies children with acute lymphoblastic leukemia who have a superior clinical outcome.
Coustan-Smith E, Ribeiro RC, Stow P, Zhou Y, Pui CH, Rivera GK, Pedrosa F, Campana D.
Blood. 2006 Jul 1;108(1):97-102. Epub 2006 Mar 14.
PMID 16537802
Clinical significance of minimal residual disease quantification in adult patients with standard-risk acute lymphoblastic leukemia.
Bruggemann M, Raff T, Flohr T, Gokbuget N, Nakao M, Droese J, Luschen S, Pott C, Ritgen M, Scheuring U, Horst HA, Thiel E, Hoelzer D, Bartram CR, Kneba M.
Blood. 2006 Feb 1;107(3):1116-23. Epub 2005 Sep 29.
PMID 16195338
Genes contributing to minimal residual disease in childhood acute lymphoblastic leukemia: prognostic significance of CASP8AP2.
Flotho C, Coustan-Smith E, Pei D, Iwamoto S, Song G, Cheng C, Pui CH, Downing JR, Campana D.
Blood. 2006 Aug 1;108(3):1050-7. Epub 2006 Apr 20.
PMID 16627760
A set of genes that regulate cell proliferation predicts treatment outcome in childhood acute lymphoblastic leukemia.
Flotho C, Coustan-Smith E, Pei D, Cheng C, Song G, Pui CH, Downing JR, Campana D.
Blood. 2007 Aug 15;110(4):1271-7. Epub 2007 Apr 24.
PMID 17456722
Submicroscopic bone marrow involvement in isolated extramedullary relapses in childhood acute lymphoblastic leukemia: a more precise definition of "isolated" and its possible clinical implications, a collaborative study of the Resistant Disease Committee of the International BFM study group.
Hagedorn N, Acquaviva C, Fronkova E, von SA, Barth A, zur SU, Schrauder A, Trka J, Gaspar N, Seeger K, Henze G, Cave H, Eckert C.
Blood. 2007 Dec 1;110(12):4022-9. Epub 2007 Aug 24.
PMID 17720883
Genome-wide analysis of genetic alterations in acute lymphoblastic leukaemia.
Mullighan CG, Goorha S, Radtke I, Miller CB, Coustan-Smith E, Dalton JD, Girtman K, Mathew S, Ma J, Pounds SB, Su X, Pui CH, Relling MV, Evans WE, Shurtleff SA, Downing JR.
Nature. 2007 Apr 12;446(7137):758-64.
PMID 17344859
Clearance of minimal residual disease after allogeneic stem cell transplantation and the prediction of the clinical outcome of adult patients with high-risk acute lymphoblastic leukemia.
Spinelli O, Peruta B, Tosi M, Guerini V, Salvi A, Zanotti MC, Oldani E, Grassi A, Intermesoli T, Mico C, Rossi G, Fabris P, Lambertenghi-Deliliers G, Angelucci E, Barbui T, Bassan R, Rambaldi A.
Haematologica. 2007 May;92(5):612-8.
PMID 17488684
Molecular relapse in adult standard-risk ALL patients detected by prospective MRD monitoring during and after maintenance treatment: data from the GMALL 06/99 and 07/03 trials.
Raff T, Gokbuget N, Luschen S, Reutzel R, Ritgen M, Irmer S, Bottcher S, Horst HA, Kneba M, Hoelzer D, Bruggemann M.
lood. 2007 Feb 1;109(3):910-5. Epub 2006 Oct 5.
PMID 17023577
Analysis of minimal residual disease by Ig/TCR gene rearrangements: guidelines for interpretation of real-time quantitative PCR data.
van der Velden VH, Cazzaniga G, Schrauder A, Hancock J, Bader P, Panzer-Grumayer ER, Flohr T, Sutton R, Cave H, Madsen HO, Cayuela JM, Trka J, Eckert C, Foroni L, Zur Stadt U, Beldjord K, Raff T, van der Schoot CE, van Dongen JJ.
Leukemia. 2007 Apr;21(4):604-11. Epub 2007 Feb 8.
PMID 17287850
Quantitative analysis of minimal residual disease predicts relapse in children with B-lineage acute lymphoblastic leukemia in DFCI ALL Consortium Protocol 95-01.
Zhou J, Goldwasser MA, Li A, Dahlberg SE, Neuberg D, Wang H, Dalton V, McBride KD, Sallan SE, Silverman LB, Gribben JG.
Blood. 2007 Sep 1;110(5):1607-11. Epub 2007 May 7.
PMID 17485550
Minimal residual disease values discriminate between low and high relapse risk in children with B-cell precursor acute lymphoblastic leukemia and an intrachromosomal amplification of chromosome 21: the Austrian and German acute lymphoblastic leukemia Berlin-Frankfurt-Munster (ALL-BFM) trials.
Attarbaschi A, Mann G, Panzer-Grumayer R, Rottgers S, Steiner M, Konig M, Csinady E, Dworzak MN, Seidel M, Janousek D, Moricke A, Reichelt C, Harbott J, Schrappe M, Gadner H, Haas OA.
J Clin Oncol. 2008 Jun 20;26(18):3046-50.
PMID 18565891
Clinical significance of minimal residual disease in childhood acute lymphoblastic leukemia and its relationship to other prognostic factors: a Children's Oncology Group study.
Borowitz MJ, Devidas M, Hunger SP, Bowman WP, Carroll AJ, Carroll WL, Linda S, Martin PL, Pullen DJ, Viswanatha DS, Willman CL, Winick N, Camitta BM.
Blood. 2008 Jun 15;111(12):5477-85. Epub 2008 Apr 3.
PMID 18388178
Status of minimal residual disease testing in childhood haematological malignancies.
Campana D.
Br J Haematol. 2008a Nov;143(4):481-9. Epub 2008 Aug 15.
PMID 18710378
Molecular determinants of treatment response in acute lymphoblastic leukemia.
Campana D.
Hematology Am Soc Hematol Educ Program. 2008b:366-73.
PMID 19074112
Minimal residual disease-directed risk stratification using real-time quantitative PCR analysis of immunoglobulin and T-cell receptor gene rearrangements in the international multicenter trial AIEOP-BFM ALL 2000 for childhood acute lymphoblastic leukemia.
Flohr T, Schrauder A, Cazzaniga G, Panzer-Grumayer R, van der Velden V, Fischer S, Stanulla M, Basso G, Niggli FK, Schafer BW, Sutton R, Koehler R, Zimmermann M, Valsecchi MG, Gadner H, Masera G, Schrappe M, van Dongen JJ, Biondi A, Bartram CR.
Leukemia. 2008 Apr;22(4):771-82. Epub 2008 Jan 31.
PMID 18239620
Status of minimal residual disease after induction predicts outcome in both standard and high-risk Ph-negative adult acute lymphoblastic leukaemia. The Polish Adult Leukemia Group ALL 4-2002 MRD Study.
Holowiecki J, Krawczyk-Kulis M, Giebel S, Jagoda K, Stella-Holowiecka B, Piatkowska-Jakubas B, Paluszewska M, Seferynska I, Lewandowski K, Kielbinski M, Czyz A, Balana-Nowak A, Krol M, Skotnicki AB, Jedrzejczak WW, Warzocha K, Lange A, Hellmann A.
Br J Haematol. 2008 May 19.
PMID 18492099
Initiating and cancer-propagating cells in TEL-AML1-associated childhood leukemia.
Hong D, Gupta R, Ancliff P, Atzberger A, Brown J, Soneji S, Green J, Colman S, Piacibello W, Buckle V, Tsuzuki S, Greaves M, Enver T.
Science. 2008 Jan 18;319(5861):336-9.
PMID 18202291
Minimal residual disease is an important predictive factor of outcome in children with relapsed 'high-risk' acute lymphoblastic leukemia.
Paganin M, Zecca M, Fabbri G, Polato K, Biondi A, Rizzari C, Locatelli F, Basso G.
Leukemia. 2008 Dec;22(12):2193-200. Epub 2008 Aug 28.
PMID 18754029
Minimal residual disease in acute lymphoblastic leukemia.
Campana D.
Semin Hematol. 2009 Jan;46(1):100-6.
PMID 19100372
Minimal disseminated disease in childhood T-cell lymphoblastic lymphoma: a report from the children's oncology group.
J Clin Oncol. 2009a Jul 20;27(21):3533-9. Epub 2009 Jun 22.
Coustan-Smith E, Sandlund JT, Perkins SL, Chen H, Chang M, Abromowitch M, Campana D.
PMID 19546402
Early T-cell precursor leukaemia: a subtype of very high-risk acute lymphoblastic leukaemia.
Coustan-Smith E, Mullighan CG, Onciu M, Behm FG, Raimondi SC, Pei D, Cheng C, Su X, Rubnitz JE, Basso G, Biondi A, Pui CH, Downing JR, Campana D.
Lancet Oncol. 2009b Feb;10(2):147-56. Epub 2009 Jan 13.
PMID 19147408
Improved risk classification for risk-specific therapy based on the molecular study of minimal residual disease (MRD) in adult acute lymphoblastic leukemia (ALL).
Bassan R, Spinelli O, Oldani E, Intermesoli T, Tosi M, Peruta B, Rossi G, Borlenghi E, Pogliani EM, Terruzzi E, Fabris P, Cassibba V, Lambertenghi-Deliliers G, Cortelezzi A, Bosi A, Gianfaldoni G, Ciceri F, Bernardi M, Gallamini A, Mattei D, Di BE, Romani C, Scattolin AM, Barbui T, Rambaldi A.
Blood. 2009 Apr 30;113(18):4153-62. Epub 2009 Jan 13.
PMID 19141862
Deletion of IKZF1 and prognosis in acute lymphoblastic leukemia.
Mullighan CG, Su X, Zhang J, Radtke I, Phillips LA, Miller CB, Ma J, Liu W, Cheng C, Schulman BA, Harvey RC, Chen IM, Clifford RJ, Carroll WL, Reaman G, Bowman WP, Devidas M, Gerhard DS, Yang W, Relling MV, Shurtleff SA, Campana D, Borowitz MJ, Pui CH, Smith M, Hunger SP, Willman CL, Downing JR.
N Engl J Med. 2009 Jan 29;360(5):470-80. Epub 2009 Jan 7.
PMID 19129520
Treating childhood acute lymphoblastic leukemia without cranial irradiation.
Pui CH, Campana D, Pei D, Bowman WP, Sandlund JT, Kaste SC, Ribeiro RC, Rubnitz JE, Raimondi SC, Onciu M, Coustan-Smith E, Kun LE, Jeha S, Cheng C, Howard SC, Simmons V, Bayles A, Metzger ML, Boyett JM, Leung W, Handgretinger R, Downing JR, Evans WE, Relling MV.
N Engl J Med. 2009 Jun 25;360(26):2730-41.
PMID 19553647
Prognostic significance of minimal residual disease in infants with acute lymphoblastic leukemia treated within the Interfant-99 protocol.
van der Velden V, Corral L, Valsecchi MG, Jansen MW, De LP, Cazzaniga G, Panzer-Grumayer ER, Schrappe M, Schrauder A, Meyer C, Marschalek R, Nigro LL, Metzler M, Basso G, Mann G, den Boer ML, Biondi A, Pieters R, van Dongen JJ.
Leukemia. 2009 Jun;23(6):1073-9. Epub 2009 Feb 12.
PMID 19212338
Genome-wide interrogation of germline genetic variation associated with treatment response in childhood acute lymphoblastic leukemia.
Yang JJ, Cheng C, Yang W, Pei D, Cao X, Fan Y, Pounds SB, Neale G, Trevino LR, French D, Campana D, Downing JR, Evans WE, Pui CH, Devidas M, Bowman WP, Camitta BM, Willman CL, Davies SM, Borowitz MJ, Carroll WL, Hunger SP, Relling MV.
JAMA. 2009 Jan 28;301(4):393-403.
PMID 19176441
Written2009-07Dario Campana
of Oncology, St. Jude Children's Research Hospital, 262 Danny Thomas Place, Memphis TN 38105, USA


This paper should be referenced as such :
Campana, D
Detection of minimal residual disease in acute lymphoblastic leukemia
Atlas Genet Cytogenet Oncol Haematol. 2010;14(6):602-608.
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