MINIMAL RESIDUAL DISEASE ASSESSMENT IN ACUTE LYMPHOBLASTIC LEUKEMIA: METHODS FOR DETECTION AND QUANTIFICATION

Authors

  • Sara Addakiri Hassan II University of Medicine and Pharmacy, Casablanca, Morocco
  • Hanaa Bencharef Hassan II University of Medicine and Pharmacy, Casablanca, Morocco
  • Asmaa Harrach Hassan II University of Medicine and Pharmacy, Casablanca, Morocco
  • Samiha Jaddaoui Hassan II University of Medicine and Pharmacy, Casablanca, Morocco
  • Khadija Ait-Ichou IBN ROCHD Hospital Centre, Casablanca, Morocco
  • Bouchra Oukkache Hassan II University of Medicine and Pharmacy, Casablanca, Morocco

DOI:

https://doi.org/10.15407/exp-oncology.2026.02.098

Keywords:

MRD, flow cytometry, molecular technique

Abstract

Recently, the identification of minimal residual disease (MRD), also known as measurable residual disease, has seen significant advances driven by the evolving technological landscape in this field. Initially based on raw morphology, the field has since evolved to incorporate karyotyping, cytogenetics, flow cytometry, and other sensitive methods. This updated review discusses key technical considerations for successful MRD detection. We explore the correlation between the results obtained by flow cytometry and molecular genetic techniques.

References

Swerdlow SH, Campo E, Pileri SA, et al. The 2016 revisionof the World Health Organization classification of lymphoidneoplasms. Blood. 2016;127(20):2375-2390. https://doi.org/10.1182/blood-2016-01-643569

Coustan-Smith E, Mullighan CG, Onciu M, et al. Early T-cell precursorleukaemia: a subtype of very high-risk acute lymphoblastic leukaemia. Lancet Oncol. 2009;10(2):147-156. https://doi.org/10.1016/S1470-2045(08)70314-0

Gaynon PS, Trigg ME, Heerema NA, et al. Children’s Cancer Group trials in childhoodacute lymphoblastic leukemia: 1983–1995. Leukemia. 2000;14(12):2223-2233. https://doi.org/10.1038/sj.leu.2401939

Schultz KR, PullenDJ, Sather HN, et al. Risk- and response-based classification of childhoodB-precursor acute lymphoblastic leukemia: a combinedanalysis of prognostic markers from the PediatricOncology Group (POG) and Children’sCancer Group (CCG). Blood. 2006;109(3):926-935. https://doi.org/10.1182/blood-2006-01-024729

Hunger SP, Loh ML, WhitlockJA, et al. COG Acute Lymphoblastic Leukemia Committee. Children’s OncologyGroup’s 2013 blueprintfor research: Acute lymphoblastic leukemia. Pediatr Blood Cancer. 2013;60:957-963. https://doi.org/10.1002/pbc.24420

Goldberg JM, Silverman LB, Levy DE, et al. Childhood T-cell acute lymphoblastic leukemia: the Dana-Farber Cancer Institute acute lymphoblastic leukemia consortium experience. J Clin Oncol. 2003;21(19):3616-3622. https://doi.org/10.1200/JCO.2003.10.116

Wood BL, Winter SS, DunsmoreKP, et al. T-lymphoblastic Leukemia (T-ALL) shows excellent outcome, lack of significance of the early thymic precursor(ETP) immunophenotype, and validation of the prognostic value of end-induction minimalresidual disease(MRD) in Children’s OncologyGroup (COG) study AALL0434. Blood.2014;124(21):1.https://doi.org/10.1182/blood.V124.21.1.1

Smith M, Arthur D, Camitta B, et al. Uniformapproach to risk classification and treatment assignment for children withacute lymphoblastic leukemia.J ClinOncol. 1996;14(1):18-24. https://doi.org/10.1200/JCO.1996.14.1.18

Pullen J, Shuster JJ, Link M, et al. Significance of commonly used prognostic factors differs for childrenwith T cell acute lymphocyticleukemia (ALL), as comparedto those with B-precursor ALL. A Pediatric Oncology Group (POG)study. Leukemia. 1999;13(11):1696-1707. https://doi.org/10.1038/sj.leu.2401555

Rambaldi A, Borleri G, Dotti G, et al. Innovativetwo-step negative selectionof granulocyte colony-stimulating factor–mobilized circulating progenitor cells: Adequacy for autologous and allogeneic transplantation. Blood.1998;91(6):2189-2196.https://doi.org/10.1182/blood.V91.6.2189

Gaipa G, BassoG, Biondi A, et al. Detectionof minimal residualdisease in pediatric acute lymphoblastic leukemia. Cytometry B Clin Cytom. 2013;84(6):359-369. https://doi.org/10.1002/cyto.b.21101

Mancini M, Cedrone M, Diverio D, et al. Use of dual-color interphase FISH for the detectionof inv(16) in acute myeloid leukemiaat diagnosis, relapse and during follow-up: a study of 23 patients. Leukemia.2000;14(3):364-368. https://doi.org/10.1038/sj.leu.2401678

Bielorai B, Golan H, Trakhtenbrot L, et al. Combined analysis of morphology and fluorescence in situ hybridiza- tion in follow-upof minimal residualdisease in a child with Philadelphia-positive acute lymphoblastic leukemia. Cancer GenetCytogenet. 2002;138(1):64-68. https://doi.org/10.1016/S0165-4608(02)00578-2

Coustan-Smith E, Behm FG, Sanchez J, et al. Immunological detection of minimalresidual diseasein children with acute lymphoblastic leukaemia. Lancet. 1998;351(9102):550-554.https://doi.org/10.1016/S0140-6736(97)10295-1

Farahat N, Morilla A, Owusu-Ankomah K, et al. Detection of minimalresidual diseasein B-lineage acute lym- phoblastic leukaemia by quantitative flow cytometry. Br J Haematol. 1998;101(1):158-164. https://doi.org/10.1046/j.1365-2141.1998.00675.x

Coustan-Smith E, Sancho J, Hancock ML, et al. Clinicalimportance of minimalresidual diseasein childhood acute lymphoblasticleukemia. Blood. 2000;96(8):2691-2696. https://doi.org/10.1182/blood.V96.8.2691

San Miguel JF, Vidriales MB, López-Berges C, et al. Early immunophenotypical evaluation of minimalresidual diseasein acute myeloidleukemia identifies different patientrisk groups and may contribute to postinduction treatment stratification. Blood. 2001;98(6):1746-1751. https://doi.org/10.1182/blood.V98.6.1746.

Dworzak MN, Fröschl G, PrintzD, et al.; AustrianBerlin-Frankfurt-Münster Study Group. Prognostic significance and modalities of flow cytometric minimal residualdisease detection in childhoodacute lymphoblastic leukemia. Blood. 2002;99(6):1952-1958. https://doi.org/10.1182/blood.V99.6.1952

Coustan-Smith E, Sancho J, Behm FG, et al. Prognostic importance of measuring early clearance of leukemiccells by flow cytometryin childhood acute lymphoblastic leukemia. Blood. 2002;100(1):52-58. https://do.org/10.1182/blood-2002-01-0006

Coustan-Smith E, Sancho J, Hancock ML, et al. Use of peripheral bloodinstead of bone marrow to monitor residual diseasein children with acute lymphoblastic leukemia. Blood.2002;100(7):2399-2402. https://doi.org/10.1182/blood-2002-04-1130

Neale GA, Coustan-Smith E, Pan Q, et al. Tandem application of flow cytometryand polymerase chain reaction for comprehensive detection of minimalresidual diseasein childhood acute lymphoblastic leukemia. Leukemia.1999;13(8):1221-1226. https://doi.org/10.1038/sj.leu.2401459

Oelschlägel U, Nowak R, Schaub A, et al. Shift ofaberrant antigen expression atrelapse orattreatment failure in acute leuke- mia. Cytometry.2000;42(4):247-253. https://doi.org/10.1002/1097-0320(20000815)42:4<247::AID-CYTO5>3.0.CO;2-V

Baer MR, Stewart CC, Dodge RK, et al. High frequencyof immunophenotype changes in acute myeloid leuke-mia at relapse:implications for residual diseasedetection (Cancer and LeukemiaGroup B Study 8361). Blood. 2001;97(11):3574-3580. https://doi.org/10.1182/blood.V97.11.3574

Porwit-MacDonald A, Björklund E, Lucio P, et al. BIOMED-1 concerted action report: Flow cytometric charac- terization of CD7+ cell subsets in normal bone marrow as a basis for the diagnosisand follow-up of T cell acute lymphoblastic leukemia (T-ALL). Leukemia.2000;14(5):816-825.https://doi.org/10.1038/sj.leu.2401741

Campana D, Coustan-Smith E. Advances in the immunological monitoring of childhood acute lymphoblastic leu- kaemia. BestPract Res Clin Haematol.2002;15(1):1-19. https://doi.org/10.1053/beha.2002.0182

Dworzak MN, Fritsch G, Fleischer C, et al. Comparative phenotype mapping of normal vs. malignant pediatric B-lymphopoiesis unveils leukemia-associated aberrations. Exp Hematol.1998;26(4):305-313. PMID: 9546313

Ciudad J, San Miguel JF, López-Berges MC, et al. Detectionof abnormalities in B-cell differentiation pattern is a useful tool to predict relapse in precursor-B-ALL. Br J Haematol. 1999;104(4):695-705. https://doi.org/10.1046/j.1365-2141.1999.01236.x

Fuda F, Chen W. Minimal/measurable residual disease detectionin acute leukemiasby multiparameter flow cyto- metry. Curr HematolMalig Rep. 2018;13(6):455-466. https://doi.org/10.1007/s11899-018-0479-1

Lavabre-Bertrand T, Janossy G, Ivory K, et al. Leukemia-associated changes identified by quantitative flow cyto- metry: I. CD10 expression. Cytometry. 1994;18(4):209-217. https://doi.org/10.1002/cyto.990180404

Flores-Montero J, Sanoja-Flores L, Paiva B, et al. Next Generation Flow for highly sensitive and standardized detection of minimal residual disease in multiple myeloma. Leukemia. 2017;31(10):2094-2103. https://doi.org/10.1038/leu.2017.29

Theunissen P, Mejstrikova E, Sedek L, et al. Standardized flow cytometryfor highly sensitiveMRD measurements in B-cell acutelymphoblastic leukemia. Blood. 2017;129(3):347-357. https://doi.org/10.1182/blood-2016-07-726307

Della Starza I, ChiarettiS, De Propris MS, et al. Minimal residual diseasein acute lymphoblastic leukemia: Techni- cal andclinical advances. Front Oncol.2019;9:726. https://doi.org/10.3389/fonc.2019.00726

Theunissen P, Mejstrikova E, Sedek L, et al. Standardized flow cytometryfor highly sensitiveMRD measurements in B-cell acutelymphoblastic leukemia. Blood. 2017;129(3):347-357. https://doi.org/10.1182/blood-2016-07-726307

Liu Z, Li Y, Shi C. Monitoring minimal/measurable residual diseasein B-cell acute lymphoblastic leukemia by flow cytometryduring targeted therapy. Int J Hematol. 2021;113(3):337-343. https://doi.org/10.1007/s12185-021-03085-y

Kruse A, Abdel-Azim N, Kim HN, et al. Minimalresidual disease detectionin acute lymphoblastic leukemia. Int JMol Sci. 2020;21(3):1054. https://doi.org/10.3390/ijms21031054

Szczepański T, Beishuizen A, Pongers-Willemse MJ, etal. Cross-lineage T cell receptor gene rearrangements occurin more than ninetypercent of childhood precursor-Bacute lymphoblastic leukemias:Alternative PCR targets for detectionof minimal residual disease. Leukemia.1999;13(2):196-205. https://doi.org/10.1038/sj.leu.2401277

Langerak AW, Groenen PJTA, Brüggemann M, et al. EuroClonality/BIOMED-2 guidelines for interpretation and reporting of Ig/TCR clonality testing in suspectedlymphoproliferations. Leukemia.2012;26(10):2159-2171. https://doi.org/10.1038/leu.2012.246

Langerak AW, Szczepański T, van der Burg M, et al. Heteroduplex PCR analysisof rearranged T cell receptor genes for clonality assessment in suspect T cell proliferations. Leukemia.1997;11(12):2192-2199. https://doi.org/10.1038/sj.leu.2400887

Germano G, del Giudice L, PalatronS, et al. Clonality profile in relapsedprecursor-B-ALL children by GeneScanand sequencing analyses. Consequences on minimalresidual diseasemonitoring. Leukemia.2003;17(8):1573-1582. https://doi.org/10.1038/sj.leu.2403008

Verhagen O, Willemse MJ, Breunis WB, et al. Application of germline IGH probes in real-timequantitative PCR for the detection of minimal residualdisease in acute lymphoblastic leukemia. Leukemia.2000;14(8):1426-1435. https://doi.org/10.1038/sj.leu.2401801

van der Velden VH, CazzanigaG, Schrauder A, et al.; EuropeanStudy Group on MRD detectionin ALL (ESG-MRD-ALL). Analysis of minimal residual disease by Ig/TCR gene rearrangements: guidelines for interpretation of real-timequantitative PCR data. Leukemia. 2007;21(4):604-611. https://doi.org/10.1038/sj.leu.2404586

van der Velden VH, NoordijkR, Brussee M, et al. Minimalresidual diseasediagnostics in acute lymphoblastic leukaemia: Impact of primer characteristics and size of junctional regions. Br J Haematol. 2014;164(3):451-453. https://doi.org/10.1111/bjh.12621

Szczepański T, Velden VHJ van der, Raff T, et al. Comparative analysis of T-cell receptor gene rearrangements at diagnosis and relapse of T-cell acute lymphoblastic leukemia (T-ALL) shows high stabilityof clonal markers for monitoring of minimalresidual diseaseand reveals the occurrence of second T-ALL. Leukemia.2003;17(11):2149-2156. https://doi.org/10.1038/sj.leu.2403081

Szczepanski T, Willemse MJ, Brinkhof B, et al. Comparative analysis of Ig and TCR gene rearrangements at diagno- sis and at relapse of childhoodprecursor-B–ALL provides improved strategies for selection of stable PCR targetsfor monitoring of minimal residual disease. Blood.2002;99(7):2315-2323. https://doi.org/10.1182/blood.v99.7.2315

Pui CH, RellingMV, Downing JR. Acute lymphoblastic leukemia. N Engl J Med. 2004;350(15):1535-1548. https://doi.org/10.1056/NEJMra023001

Downing JR, Shannon KM. Acute leukemia:A pediatric perspective. Cancer Cell. 2002;2(6):437-445. https://doi.org/10.1016/S1535-6108(02)00211-8

Ajuba IC, Madu AJ, Okocha C, et al. Frequencyand clinical impact of ETV6/RUNX1, AF4-MLL,and BCR/ABL fusion genes on features of acute lymphoblastic leukemia at presentation. Niger J Clin Pract. 2016;19(2):237-241. https://doi.org/10.4103/1119-3077.164351

Wu D, Sherwood A, Fromm JR, et al. High-throughput sequencing detects minimal residual disease in acute T lymphoblastic leukemia. Sci Transl Med. 2012;4(134):134ra63. https://doi.org/10.1126/scitranslmed.3003656

Wu D, Emerson RO, Sherwood A, et al. Detection of minimalresidual diseasein B lymphoblastic leukemia by high-throughput sequencing of IGH. Clin Cancer Res. 2014;20(17):4540-4548. https://doi.org/10.1158/1078-0432.CCR-13-3231

Wood B, Wu D, Crossley B, et al. Measurable residual diseasedetection by high-throughput sequencing improves risk stratificationfor pediatric B-ALL. Blood. 2018;131(12):1350-1359. https://doi.org/10.1182/blood-2017-09-806521

Wood BL, Wu D, Kirsch IM, et al. Residualdisease monitoringby high throughputsequencing provides risk stratifi- cation in childhood B-ALL and identifies a novel subset of patientshaving poor outcome. Blood.2016;128(22):1086. https://doi.org/10.1182/blood.V128.22.1086.1086

Pulsipher MA, Carlson C, LangholzB, et al. IgH-V(D)JNGS-MRD measurement pre-and early post-allotrans- plant defines very low-and very high-riskALL patients. Blood. 2015;125(22):3501-3508. https://doi.org/10.1182/blood-2014-12-615757

Kotrova M, Muzikova K, Mejstrikova E, et al. The predictive strength of next-generation sequencing MRD detec-tion for relapse compared with current methods in childhood ALL. Blood. 2015;126(8):1045-1047. https://doi.org/10.1182/blood-2015-07-655159

Huggett JF, Whale A. Digital PCR as a novel technology and its potential implications for molecular diagnostics. Clin Chem. 2013;59(12):1691-1693. https://doi.org/10.1373/clinchem.2013.214742

Huggett JF, Cowen S, Foy CA.Considerations for digital PCR as an accuratemolecular diagnostic tool. Clin Chem. 2015;61(1):79-88. https://doi.org/10.1373/clinchem.2013.214742

Reinert T, Schøler LV, Thomsen R, et al. Analysis of circulating tumour DNA tomonitor disease burden following colorectalcancer surgery. Gut. 2016;65(4):625-634. https://doi.org/10.1136/gutjnl-2014-308859

Waterhouse M, Follo M, PfeiferD, et al. Sensitive and accuratequantification of JAK2 V617F mutation in chronic myeloproliferative neoplasms by droplet digitalPCR. Ann Hematol. 2016;95(5):739-744. https://doi.org/10.1007/s00277-016-2623-0

Della Starza I, De Novi LA, Elia L, et al. Optimizing molecular minimalresidual diseaseanalysis in adult acute lymphoblastic leukemia. Cancers.2023;15(2):374. https://doi.org/10.3390/cancers15020374

Pecoraro S, BerbenG, Burns M, et al. Overview and recommendations for the application of digital PCR.Lux- embourg: Publications Office of the EuropeanUnion, ISBN 978-92-76-00180-5, Article115736. https://doi.org/10.2760/192883

van der Velden VH, CazzanigaG, Schrauder A, et al.; EuropeanStudy Group on MRD detectionin ALL (ESG-MRD-ALL). Analysis of minimal residual disease by Ig/TCR gene rearrangements: guidelines for interpretation of real-timequantitative PCR data. Leukemia. 2007;21:604-611. https://doi.org/10.1038/sj.leu.2404586

Fronkova E, Muzikova K, Mejstrikova E, et al. B-cell reconstitution after allogeneic SCT impairs minimalresidual diseasemonitoring in childrenwith ALL. Bone Marrow Transplant. 2008;42(3):187-196. https://doi.org/10.1038/bmt.2008

van Dongen JJ, van der Velden VH, Brüggemann M, et al. Minimal residual disease diagnostics in acute lympho-blastic leukemia: need for sensitive, fast, and standardized technologies. Blood. 2015;125(26):3996-4009. https://doi.org/10.1182/blood-2015-03-580027

Kotrova M, van der Velden VHJ, van DongenJJM, et al. Next-generation sequencing indicates false-positive MRD results and better predictsprognosis after SCT in patients with childhoodALL. Bone Marrow Transplant. 2017;52(7):962-968. https://doi.org/10.1038/bmt.2017.16

Kotrova M, Muzikova K, Mejstrikova E, et al. The predictive strength of next-generation sequencing MRD detec-tion for relapse compared with current methods in childhood ALL. Blood. 2015;126(8):1045-1047. https://doi.org/10.1182/blood-2015-07-655159

Eckert C, Flohr T, Köhler R, et al. Very early/early relapses of acute lymphoblastic leukemia show unexpected changes of clonal markers and high heterogeneity in responseto initial and relapse treatment. Leukemia.2011;25(8):1305-1313.https://doi.org/10.1038/leu.2011.89

Ladetto M, Brüggemann M, MonitilloL, et al. Next-generation sequencing and real-time quantitative PCR for minimal residual diseasedetection in B-cell disorders. Leukemia.2014;28(6):1299-1307. https://doi.org/10.1038/leu.2013.375

Submitted: September 10, 2025

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21.08.2026

How to Cite

Addakiri, S., Bencharef, H., Harrach, A., Jaddaoui, S., Ait-Ichou, K., & Oukkache, B. (2026). MINIMAL RESIDUAL DISEASE ASSESSMENT IN ACUTE LYMPHOBLASTIC LEUKEMIA: METHODS FOR DETECTION AND QUANTIFICATION. Experimental Oncology, 48(2), 98–107. https://doi.org/10.15407/exp-oncology.2026.02.098