Technical Aspects of Minimal Residual Disease Detection by Multicolor Flow Cytometry in Acute Myeloid Leukemia Patients

IV Galtseva, YuO Davydova, NM Kapranov, KA Nikiforova, EN Parovichnikova

National Research Center for Hematology, 4 Novyi Zykovskii pr-d, Moscow, Russian Federation, 125167

For correspondence: Yuliya Olegovna Davydova, MD, PhD, 4 Novyi Zykovskii pr-d, Moscow, Russian Federation, 125167; Tel.: 8(495)612-62-21; e-mail: juliya89mur@yandex.ru

For citation: Galtseva IV, Davydova YuO, Kapranov NM, et al. Technical Aspects of Minimal Residual Disease Detection by Multicolor Flow Cytometry in Acute Myeloid Leukemia Patients. Clinical oncohematology. 2021;14(3):503–12. (In Russ).

DOI: 10.21320/2500-2139-2021-14-4-503-512


ABSTRACT

Detection and monitoring of minimal residual disease (MRD) are essential components of programmed therapy.They are crucial for the choice of treatment strategy and for prognostic purposes practically in all hematologic diseases. MRD is often detected by multicolor flow cytometry, the method with fairly high specificity and sensitivity. However, to identify MRD in acute myeloid leukemia patients is one of the most challenging tasks flow cytometry specialists are faced with. Cytometric data analysis requires the expert knowledge of immunophenotype of all maturing bone marrow cells. Besides, MRD analysis in acute myeloid leukemia has not been standardized while approaches suggested by different studies vary considerably. The present paper reports the experience of MRD analysis, demonstrates the gating strategy, immunophenotype description of normal non-tumor hematopoietic cells, and presents some examples of MRD assessment. Additionally, panels of monoclonal antibodies are provided, along with an evaluation of their advantages and disadvantages.

Keywords: minimal residual disease, acute myeloid leukemias, flow cytometry, gating, immunophenotyping.

Received: June 9, 2021

Accepted: September 5, 2021

Read in PDF

Статистика Plumx английский

REFERENCES

  1. Cheson BD, Bennett JM, Kopecky KJ, et al. Revised recommendations of the international working group for diagnosis, standardization of response criteria, treatment outcomes, and reporting standards for therapeutic trials in acute myeloid leukemia. J Clin Oncol. 2003;21(24):4642–9. doi: 10.1200/JCO.2003.04.036.
  2. Pui CH, Campana D. New definition of remission in childhood acute lymphoblastic leukemia. Leukemia. 2000;14(5):783–5. doi: 10.1038/sj.leu.2401780.
  3. Schuurhuis GJ, Heuser M, Freeman S, et al. Minimal/measurable residual disease in AML: a consensus document from the European LeukemiaNet MRD Working Party. Blood. 2018;131(12):1275–91. doi: 10.1182/blood-2017-09-801498.
  4. Гальцева И.В., Давыдова Ю.О., Паровичникова Е.Н. Определение минимальной измеримой остаточной болезни у взрослых больных острыми лейкозами. Гематология и трансфузиология. 2020;65(4):460–72. doi: 10.35754/0234-5730-2020-65-4-460-472.
    [Galtseva IV, Davydova YO, Parovichnikova EN. Detection of measurable residual disease in adults with acute leukaemia. Russian journal of hematology and transfusiology. 2020;65(4):460–72. doi: 10.35754/0234-5730-2020-65-4-460-472. (In Russ)]
  5. Shen Z, Gu X, Mao W, et al. Influence of pre-transplant minimal residual disease on prognosis after Allo-SCT for patients with acute lymphoblastic leukemia: Systematic review and meta-analysis. BMC Cancer. 2018;18(1):755. doi: 10.1186/s12885-018-4670-5.
  6. Leung W, Pui C-H, Coustan-Smith E, et al. Detectable minimal residual disease before hematopoietic cell transplantation is prognostic but does not preclude cure for children with very-high-risk leukemia. Blood. 2012;120(2):468–72. doi: 10.1182/blood-2012-02-409813.
  7. Norkin M, Katragadda L, Zou F, et al. Minimal residual disease by either flow cytometry or cytogenetics prior to an allogeneic hematopoietic stem cell transplant is associated with poor outcome in acute myeloid leukemia. Blood Cancer J. 2017;7(12):634. doi: 10.1038/s41408-017-0007-x.
  8. Anthias C, Dignan FL, Morilla R, et al. Pre-transplant MRD predicts outcome following reduced-intensity and myeloablative allogeneic hemopoietic SCT in AML. Bone Marrow Transplant. 2014;49(5):679–83. doi: 10.1038/bmt.2014.9.
  9. Buckley SA, Wood BL, Othus M, et al. Minimal residual disease prior to allogeneic hematopoietic cell transplantation in acute myeloid leukemia: a meta-analysis. Haematologica. 2017;102(5):865–73. doi: 10.3324/haematol.2016.159343.
  10. Wood BL. Principles of minimal residual disease detection for hematopoietic neoplasms by flow cytometry. Cytometry B Clin Cytom. 2016;90(1):47–53. doi: 10.1002/cyto.b.21239.
  11. Wood BL. Multicolor immunophenotyping: human immune system hematopoiesis. Methods Cell Biol. 2004;75:559–76. doi: 10.1016/s0091-679x(04)75023-2.
  12. Wood BL. Flow cytometric monitoring of residual disease in acute leukemia. In: Czader M, ed. Hematological Malignancies. Methods in Molecular Biology (Methods and Protocols). Vol. 999. Totowa: Humana Press; 2013. pp. 123–36. doi: 10.1007/978-1-62703-357-2_8.
  13. Лобанова Т.И., Гальцева И.В., Паровичникова Е.Н. Исследование минимальной остаточной болезни у пациентов с острыми миелоидными лейкозами методом многоцветной проточной цитофлуориметрии (обзор литературы). Онкогематология. 2018;13(1):83–102. doi: 10.17650/1818-8346-2018-13-1-83-102.
    [Lobanova TI, Galtseva IV, Parovichnikova EN. Minimal residual disease assesment in patients with acute myeloid leukemia by multicolour flow cytometry (literature review). Oncohematology. 2018;13(1):83–102. doi: 10.17650/1818-8346-2018-13-1-83-102. (In Russ)]
  14. Tien HF, Wang CH. CD7 positive hematopoietic progenitors and acute myeloid leukemia and other minimally differentiated leukemia. Leuk Lymphoma. 1998;31(1–2):93–8. doi: 10.3109/10428199809057588.
  15. Jorgensen JL, Chen SS. Monitoring of minimal residual disease in acute myeloid leukemia: methods and best applications. Clin Lymphoma Myeloma Leuk. 2011;11(Suppl 1):S49–53. doi: 10.1016/j.clml.2011.03.023.
  16. Jaso JM, Wang SA, Jorgensen JL, Lin P. Multi-color flow cytometric immunophenotyping for detection of minimal residual disease in AML: past, present and future. Bone Marrow Transplant. 2014;49(9):1129–38. doi: 10.1038/bmt.2014.99.
  17. Buldini B, Maurer-Granofszky M, Varotto E, Dworzak MN. Flow-cytometric monitoring of minimal residual disease in pediatric patients with acute myeloid leukemia: recent advances and future strategies. Front Pediatr. 2019;7:412. doi: 10.3389/fped.2019.00412.
  18. Wood BL. Acute myeloid leukemia minimal residual disease detection: the difference from normal approach. Curr Protoc Cytom. 2020;93(1):e73. doi: 10.1002/cpcy.73.
  19. Ostendorf BN, Flenner E, Florcken A, Westermann J. Phenotypic characterization of aberrant stem and progenitor cell populations in myelodysplastic syndromes. PLoS One. 2018;13(5):e0197823. doi: 10.1371/journal.pone.0197823.
  20. Goardon N, Marchi E, Atzberger A, et al. Coexistence of LMPP-like and GMP-like leukemia stem cells in acute myeloid leukemia. Cancer Cell. 2011;19(1):138–52. doi: 10.1016/j.ccr.2010.12.012.
  21. Shameli A, Dharmani-Khan P, Luider J, et al. Exploring blast composition in myelodysplastic syndromes and myelodysplastic/myeloproliferative neoplasms: CD45RA and CD371 improve diagnostic value of flow cytometry through assessment of myeloblast heterogeneity and stem cell aberrancy. Cytom Part B: Clin Cytom. 2020:1–16. doi: 10.1002/cyto.b.21983. Epub ahead of print.
  22. Bill M, van Kooten Niekerk BP, Woll SP, et al. Mapping the CLEC12A expression on myeloid progenitors in normal bone marrow; implications for understanding CLEC12A-related cancer stem cell biology. J Cell Mol Med. 2018;22(4):2311–8. doi: 10.1111/jcmm.13519.
  23. Eissens DN, Spanholtz J, van der Meer A, et al. Defining early human NK cell developmental stages in primary and secondary lymphoid tissues. PLoS One. 2012;7(2):e30930. doi: 10.1371/journal.pone.0030930.
  24. Stetler-Stevenson M, Paiva B, Stoolman L, et al. Consensus guidelines for myeloma minimal residual disease sample staining and data acquisition. Cytom Part B: Clin Cytom. 2016;90(1):26–30. doi: 10.1002/cyto.b.21249.
  25. Palmieri R, Piciocchi A, Arena V, et al. Clinical relevance of- limit of detection (LOD) – limit of quantification (LOQ) – based flow cytometry approach for measurable residual disease (MRD) assessment in acute myeloid leukemia (AML). Blood. 2020;136(Suppl 1):37–8. doi: 10.1182/blood-2020-139557.

Role of Superficial CD200 Marker in Differential Diagnosis of Malignant B-Cell Lymphoproliferative Diseases

YuV Mirolyubova, EA Stadnik, TS Nikulina, VV Strugov, TO Andreeva, YuV Virts, RV Grozov, AYu Zaritskey

Federal Almazov North-West Medical Research Centre, 2 Akkuratova str., Saint Petersburg, Russian Federation, 197341

For correspondence: Yuliya Vladimirovna Mirolyubova, 2 Akkuratova str., Saint Petersburg, Russian Federation, 197341; e-mail: juli9702@yandex.ru

For citation: Mirolyubova YuV, Stadnik EA, Nikulina TS, et al. Role of Superficial CD200 Marker in Differential Diagnosis of Malignant B-Cell Lymphoproliferative Diseases. Clinical oncohematology. 2017;10(2):169–75 (In Russ).

DOI: 10.21320/2500-2139-2017-10-2-169-175


ABSTRACT

Background & Aims. Flow cytometry is successfully used for diagnosis of malignant lymphoproliferative disorders. However, there are atypical cases that are difficult to interpret; thus, new markers relevant for the differential diagnosis are to be searched for. The aim is to analyze CD200 expression in patients with B-cell lymphoproliferative disorders.

Materials & Methods. 187 patients with chronic lymphocytic leukemia (CLL), 14 patients with mantle cell lymphoma (MCL), 9 patients with marginal zone lymphoma (MZL), and 5 patients with hairy cell leukemia (HCL) were enrolled in the study. Neoplasm was not confirmed in 12 subjects. The patients underwent the following tests: CBC, immunophenotyping of peripheral blood or bone marrow lymphocytes, and a cytogenetic test. In some cases, an additional immunohistochemical test of bone marrow trepanobiopsy or lymph node biopsy samples was required.

Results. In all cases of CLL and HCL, the CD200 expression was positive; mean fluorescence intensity was higher in these cases as compared to other groups. Negative expression of CD200 prevailed in MCL patients; however, at the same time 2 cases of intermediate and positive expression were reported, both showing moderate fluorescence intensity values. CD200 expression was heterogeneous in MZL patients.

Conclusion. The CD200 negative expression excludes typical HCL and CLL. Additional cytogenetic and immunnohistoсhemical tests should be performed in such cases to verify the diagnosis, first of all, MCL or MZL.

Keywords: CD200, flow cytometry, diagnosis, chronic lymphocytic leukemia, mantle cell lymphoma, marginal zone lymphoma, hairy cell leukemia.

Received: September 7, 2016

Accepted: January 3, 2017

Read in PDF (RUS)pdficon


REFERENCES

  1. Купрышина Н.А., Тупицын Н.Н. Проточная цитометрия в онкогематологии. Часть II. Основы и нововведения в диагностике хронического лимфолейкоза. Клиническая онкогематология. 2012;5(4):349–54.
    [Kupryshina NA, Tupitsyn NN. Flow cytometry in hematology malignancies. Part II: ABC and news in diagnostics of chronic lymphocytic leukaemia. Klinicheskaya onkogematologiya. 2012;5(4):349–54. (In Russ)]
  2. Стадник Е.А., Стругов В.В., Вирц Ю.В., Зарицкий А.Ю. Хронический лимфолейкоз. Рекомендации по диагностике и лечению. Трансляционная медицина. 2012;17:104–15.
    [Stadnik EA, Strugov VV, Virts YuV, Zaritskey AYu. Chronic lymphocytic leukemia. Guidelines for diagnosis and treatment. Translyatsionnaya meditsina. 2012;17:104–15. (In Russ)]
  3. Swerdlow SH, Campo E, Harris NL, et al, eds. WHO Classification of Tumours of Haematopoietic and Lymphoid Tissues. 4th edition. Lyon: IARC Press; 2008.
  4. Kohnke T, Wittmann VK, Sauter D, et al. Proposal For a Novel Scoring System For The Diagnosis оf CLL. Blood. 2013;122(21):47–5599 (Plenary Abstracts).
  5. Morice WG, Kurtin PJ, Hodnefield JM, et al. Predictive Value of Blood and Bone Marrow Flow Cytometry in B-Cell Lymphoma Classification: Comparative Analysis of Flow Cytometry and Tissue Biopsy in 252 Patients. Mayo Clin Proc. 2008;83(7):776–85. doi: 10.4065/83.7.776.
  6. Луговская С.А., Кисиличина Д.Г., Почтарь М.Е. и др. Новые маркеры (CD160, CD200, LAIR-1) в диагностике В-клеточных лимфопролиферативных заболеваний. Клиническая онкогематология. 2013;6(1):45–52.
    [Lugovskaya SA, Kisilichina DG, Pochtar’ ME, et al. New markers (CD160, CD200, and LAIR-1) in diagnosis of B-cell lymphoproliferative disorders. Klinicheskaya onkogematologiya. 2013;6(1):45–52. (In Russ)]
  7. Brunetti L, Di Noto R, Abate G, et al. CD200/OX2, a cell surface molecule with immuno-regulatory function is consistently expressed on hairy cell leukaemia neoplastic cells. Br J Haematol. 2009;145(5):665–78. doi: 10.1111/j.1365-2141.2009.07644.x.
  8. Palumbo GA, Parrinello N, Fargione G, et al. CD200 expression may help in differential diagnosis between mantle cell lymphoma and B-cell chronic lymphocytic leukemia. Leuk Res. 2009;33(9):1212–6. doi: 10.1016/j.leukres.2009.01.017.
  9. Dorfman DM, Shahsafaei A. CD200 (OX-2 Membrane Glycoprotein) Expression in B Cell–Derived Neoplasms. Am J Clin Pathol. 2010;134(5):726–33. doi: 10.1309/ajcp38xrrugsqovc.
  10. Sander B. Mantle cell lymphoma: recent insights into pathogenesis, clinical variability, and new diagnostic markers. Semin Diagn Pathol. 2011;28(3):245–55. doi: 10.1053/j.semdp.2011.02.010.
  11. Alapat D, Coviello-Malle J, Owens R, et al. Diagnostic Usefulness and Prognostic Impact of CD200 Expression in Lymphoid Malignancies and Plasma Cell Myeloma. Am J Clin Pathol. 2012;137(1):93–100. doi: 10.1309/ajcp59uorcyzevqo.
  12. El Desoukey NA, Afify RA, Amin DG, et al. CD200 expression in B-cell chronic lymphoproliferative disorders. J Investig Med. 2012;60(1):56–61. doi: 10.2310/jim.0b013e31823908f9.
  13. Pillai V, Pozdnyakova O, Charest K, et al. CD200 flow cytometric assessment and semiquantitative immunohistochemical staining distinguishes hairy cell leukemia from hairy cell leukemia-variant and other B-cell lymphoproliferative disorders. Am J Clin Pathol. 2013;140(4):536–43. doi: 10.1309/ajcpebk31vqqnddr.
  14. Challagundla P, Medeiros LJ, Kanagal-Shamanna R, et al. Differential Expression of CD200 in B-Cell Neoplasms by Flow Cytometry Can Assist in Diagnosis, Subclassification, and Bone Marrow Staging. Am J Clin Pathol. 2014;142(6):837–44. doi: 10.1309/ajcpbv9elxc0ecvl.
  15. Sandes AF, de Lourdes Chauffaille M, Regina C, et al. CD200 Has an Important Role in the Differential Diagnosis of Mature B-Cell Neoplasms by Multiparameter Flow. Cytometry. 2013;86(2):98–105. doi: 10.1002/cyto.b.21128.
  16. McCaughan GW, Clark MJ, Barclay AN. Characterization of the human homolog of the rat MRC OX-2 membrane glycoprotein. Immunogenetics. 1987;25(5):329–35. doi: 10.1007/bf00404426.
  17. Wright GJ, Jones M, Puklavec MJ, et al. The unusual distribution of the neuronal/lymphoid cell surface CD200 (OX2) glycoprotein is conserved in humans. Immunology. 2001;102(2):173–9. doi: 10.1046/j.1365-2567.2001.01163.x.
  18. Kretz-Rommel A, Qin F, Dakappagari N, et al. CD200 expression on tumor cells suppresses antitumor immunity: new approaches to cancer immunotherapy. J Immunol. 2007;178(9):5595–605. doi: 10.4049/jimmunol.178.9.5595.
  19. Moreaux J, Hose D, Reme T, et al. CD200 is a new prognostic factor in multiple myeloma. Blood. 2006;108(13):4194–7. doi: 10.1182/blood-2006-06-029355.
  20. Tonks A, Hills R, White P, et al. CD200 as a prognostic factor in acute myeloid leukemia. Leukemia. 2007;21(3):566–8. doi: 10.1038/sj.leu.2404559.
  21. Moreaux J, Veyrune JL, Reme T, et al. CD200: a putative therapeutic target in cancer. Biochem Biophys Res Commun. 2008;366(1):117–22. doi: 10.1016/j.bbrc.2007.11.103.
  22. Kretz-Rommel A, Bowdish KS. Rationale for anti-CD200 immunotherapy in B-CLL and other hematologic malignancies: new concepts in blocking immune suppression. Expert Opin Biol Ther. 2008;8(1):5–15. doi: 10.1517/14712598.8.1.5.

Acute Leukemias: Immunophenotypic Differences between Blast Cells and Their Nonneoplastic Analogues in Bone Marrow

АM Popov1, ТYu Verzhbitskaya2,3, LG Fechina2, AV Shestopalov1,4, SA Plyasunova1

1 Dmitrii Rogachev Federal Scientific Clinical Centre of Pediatric Hematology, Oncology and Immunology, 1 Samory Mashela str., Moscow, Russian Federation, 117997

2 Regional Children’s Hospital No. 1, 32 Serafimy Deryabinoi str., Yekaterinburg, Russian Federation, 620149

3 Institute of Medical Cell Technologies, 22a Karla Marksa str., Yekaterinburg, Russian Federation, 620026

4 N.I. Pirogov Russian National Research Medical University, 1 Samory Mashela str., Moscow, Russian Federation, 117997

For correspondence: Aleksandr Mikhailovich Popov, PhD, 1 Samory Mashela str., Moscow, Russian Federation, 117997; Tel.: +7(495)287-65-70; e-mail: uralcytometry@gmail.com

For citation: Popov AM, Verzhbitskaya TYu, Fechina LG, et al. Acute Leukemias: Immunophenotypic Differences between Blast Cells and Their Nonneoplastic Analogues in Bone Marrow. Clinical oncohematology. 2016;9(3):302-13 (In Russ).

DOI: http://dx.doi.org/10.21320/2500-2139-2016-9-3-302-313


ABSTRACT

Flow cytometry immunophenotyping of bone marrow tumor blasts is one of the principal methods used for acute leukemia (AL) diagnosing. Normal lymphopoietic and myelopoietic progenitors have very similar antigenic profile with leukemic cells, thus, making the AL diagnosing more difficult. Genetic disorders resulting in formation of a tumor clone contribute to development of an immunophenotype that differs from normal cells. Aberrant expression of markers detected in AL blast cells alone forms a so-called leukemia-associated immunophenotype. The leukemia-associated immunophenotype detection by multicolor flow cytometry permits distinguishing between normal and neoplastic cells. This requires simultaneous assessment of many markers on the same cells, which is possible only if multicolor flow cytometry with well-designed and well-established antibodies panels is used. Moreover, correct interpretation of the cell population location on dot plot requires adequate cytometer setup, standardized sample preparation and enough experienced personnel. That is why correct immunophenotyping is often possible only in large laboratories performing reference immunophenotyping within the frames of multicenter trials.

Keywords: acute leukemias, flow cytometry, antigenic expression, immunophenotype.

Received: February 19, 2016

Accepted: March 16, 2016

Read in PDF (RUS)pdficon


REFERENCES

  1. Morike A, Zimmermann M, Reiter A, et al. Long-term results of five consecutive trials in childhood acute lymphoblastic leukemia performed by the ALL-BFM study group from 1981 to 2000. Leukemia. 2010;24(2):265–84. doi: 10.1038/leu.2009.257.
  2. Pui CH, Carroll WL, Meshinchi S, Arceci RJ. Biology, risk stratification, and therapy of pediatric acute leukemias: an update. J Clin Oncol. 2011;29(5):551–65. doi: 10.1200/jco.2010.30.7405.
  3. Pui CH, Mullighan CG, Evans WE, Relling MV. Pediatric acute lymphoblastic leukemia: where are we going and how do we get there? Blood. 2012;120(6):1165–74. doi: 10.1182/blood-2012-05-
  4. Bene M, Castoldi G, Knapp W, et al. Proposals for the immunological classification of acute leukemias. European Group for the Immunological Characterization of Leukemias (EGIL). Leukemia. 1995;9(10):1783–6.
  5. van Lochem EG, Wiegers YM, van den Beemd R, et al. Regeneration pattern of precursor-B-cells in bone marrow of acute lymphoblastic leukemia patients depends on the type of preceding chemotherapy. Leukemia. 2000;14(4):688–95. doi: 10.1038/sj.leu.2401749.
  6. McKenna RW, Washington LT, Aquino DA, et al. Immunophenotypic analysis of hematogones (B-lymphocyte precursors) in 662 consecutive bone marrow specimens by 4-color flow cytometry. Blood. 2001;98(8):2498–507. doi: 10.1182/blood.v98.8.2498.
  7. Campana D, Coustan-Smith E. Advances in the immunological monitoring of childhood acute lymphoblastic leukaemia. Best Pract Res Clin Hematol. 2002;15(1):1–19. doi: 1053/beha.2002.0182.
  8. 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):305–13.
  9. Lucio P, Parreira A, van den Beemd MVM, et al. Flow cytometric analysis of normal B cell differentiation: a frame of reference for the detection of minimal residual disease in precursor-B-ALL. Leukemia. 1999;13(3):419–27. doi: 1038/sj.leu.2401279.
  10. Lucio P, Gaipa G, van Lochem EG, et al. BIOMED-I concerted action report: flow cytometric immunophenotyping of precursor B-ALL with standardized triple-stainings. Leukemia. 2001;15(8):1185–92. doi: 10.1038/sj.leu.2402150.
  11. Dworzak MN, Fritsch G, Fleisher C, et al. Multiparameter phenotype mapping of normal and post-chemotherapy B lymphopoiesis in pediatric bone marrow. Leukemia. 1997;11(8):1266–73. doi: 10.1038/sj.leu.2400732.
  12. Попов А.М., Вержбицкая Т.Ю., Цаур Г.А. и др. Аберрации иммунофенотипа, применимые для мониторинга минимальной остаточной болезни методом проточной цитометрии при CD10-позитивном остром лимфобластном лейкозе из В-линейных предшественников. Иммунология. 2010;31(6):299–304.
    [Popov AM, Verzhbitskaya TYu, Tsaur GA, et al. Immunophenotype aberrations used for monitoring of the minimal residual disease using flow cytometry in CD10-positive acute lymphoblastic leukemia from B-linear precursors. 2010;31(6):299–304. (In Russ)]
  13. Мовчан Л.В. Лейкоз-ассоциированный иммунофенотип опухолевых клеток у детей с острым лимфобластным лейкозом из предшественников В-лимфоцитов. Онкогематология. 2012;1:22–8.
    [Movchan LV. Leukemia-associated immunophenotype of tumor cells in childhood B-precursors acute lymphoblastic leukemia. Onkogematologiya. 2012;1:22–8. (In Russ)]
  14. Попов А.М., Вержбицкая Т.Ю., Цаур Г.А. и др. Алгоритм применения проточной цитометрии для мониторинга минимальной остаточной болезни при CD10-негативном остром лимфобластном лейкозе из B-линейных предшественников. Вопросы диагностики в педиатрии. 2012;4(5):31–5.
    [Popov AM, Verzhbitskaya TYu, Tsaur GA, et al. Methodology of flow cytometry application for minimal residual disease monitoring in childhood CD10-negative B-cell precursor acute lymphoblastic leukemia. Voprosy diagnostiki v pediatrii. 2012;4(5):31–5. (In Russ)]
  15. Ciudad J, Orfao A, Vidriales B, et al. Immunophenotypic analysis of CD19+ precursors in normal human adult bone marrow: implications for minimal residual disease detection. Haematologica. 1998;83(12):1069–75.
  16. Veltroni M, De Zen L, Sanzari MC, et al. Expression of CD58 in normal, regenerating and leukemic bone marrow B cells: implications for the detection of minimal residual disease in acute lymphocytic leukemia. Haematologica. 2003;88(11):1245–52.
  17. van Lochem EG, van der Velden VH, Wind HK, et al. Immunophenotypic differentiation patterns of normal hematopoiesis in human bone marrow: reference patterns for age-related changes and disease-induced shifts. Cytometry B Clin Cytom. 2004;60B(1):1–13. doi: 10.1002/cyto.b.20008.
  18. Lee RV, Braylan RC, Rimsza LM. CD58 expression decreases as nonmalignant B cells mature in bone marrow and is frequently overexpressed in adult and pediatric precursor B-cell acute lymphoblastic leukemia. Am J Clin Pathol. 2005;123(1):119–24. doi: 1309/x5vv6fkjq6mublpx.
  19. Robillard N, Cave H, Mechinaud F, et al. Four-color flow cytometry bypasses limitations of IG/TCR polymerase chain reaction for minimal residual disease detection in certain subsets of children with acute lymphoblastic leukemia. Haematologica. 2005;90(11):1516–23.
  20. Seegmiller AC, Kroft SH, Karandikar NJ, McKenna RW. Characterization of immunophenotypic aberrancies in 200 cases of B acute lymphoblastic leukemia. Am J Clin Pathol. 2009;132(6):940–9. doi: 10.1309/AJCP8G5RMTWUEMUU.
  21. Sedek L, Bulsa J, Sonsala A, et al. The immunophenotypes of blast cells in B-cell precursor acute lymphoblastic leukemia: how different are they from their normal counterparts. Cytometry B Clin Cytom. 2014;86(5):329–39. doi: 10.1002/cyto.b.21176.
  22. Hulspas R, O’Gorman MRG, Wood BL, et al. Consideration for the control of background fluorescence in clinical flow cytometry. Cytometry B Clin Cytom. 2009;76В(6):355–64. doi: 10.1002/cyto.b.20485.
  23. Hrusak O, Porwit-MacDonald A. Antigen expression patterns reflecting genotype of acute leukemias. Leukemia. 2002;16(7):1233–58. doi: 10.1038/sj.leu.2402504.
  24. Попов А.М., Цаур Г.А., Вержбицкая Т.Ю. и др. Иммунофенотипическая характеристика острого лимфобластного лейкоза у детей первого года жизни. Онкогематология. 2012;7(2):14–24. doi: 17650/1818-8346-2012-7-2-14-24.
    [Popov AM, Tsaur GA, Verzhbitskaya TY, et al. Immunophenotypic investigation of infant acute lymphoblastic leukemia. Oncohematology. 2012;7(2):14–24. doi: 10.17650/1818-8346-2012-7-2-14-24. (In Russ)]
  25. Fuda FS, Karandikar NJ, Chen W. Significant CD5 expression on normal stage 3 hematogones and mature B-lymphocytes in bone marrow. Am J Clin Pathol. 2009;132(5):733–7. doi: 10.1309/AJCPU5E3NXEKLFIY.
  26. Gaipa G, Basso G, Maglia O, et al. Drug-induced immunophenotypic modulation in childhood ALL: implications for minimal residual disease detection. Leukemia. 2005;19(1):49–56. doi: 10.1038/sj.leu.2403559.
  27. Gaipa G, Basso G, Ratei R, et al. Reply to van der Sluijs-Gelling, et al. Leukemia. 2005;19(12):2351–2. doi: 10.1038/sj.leu.2403912.
  28. van der Sluijs-Gelling AJ, van der Velden VHJ, Roeffen ETJM, et al. Immunophenotypic modulation in childhood precursor-B-ALL can be mimicked in vitro and is related to the induction of cell death. Leukemia. 2005;19(10):1845–7. doi: 10.1038/sj.leu.2403911.
  29. Dworzak MN, Schumich A, Printz D, et al. CD20 up-regulation in pediatric B-cell precursor acute lymphoblastic leukemia during induction treatment: setting the stage for anti-CD20 directed immunotherapy. Blood. 2008;112(10):3982–8. doi: 10.1182/blood-2008-06-
  30. Gaipa G, Basso G, Aliprandi S, et al. Prednisone induces immunophenotypic modulation of CD10 and CD34 in nonapoptotic B-cell precursor acute lymphoblastic leukemia cells. Cytometry B Clin Cytom. 2008;74B(3):150–5. doi: 10.1002/cyto.b.20408.
  31. Попов А.М., Вержбицкая Т.Ю., Цаур Г.А. и др. Изменения иммунофенотипа опухолевых бластов при CD10-позитивном остром лимфобластном лейкозе у детей к 15-му дню индукционной терапии по протоколу ALL-MB-2008. Иммунология. 2010;31(2):60–4.
    [Popov AM, Verzhbitskaya TYu, Tsaur GA, et al. Changes of tumor blast immunophenotype in CD10-positive acute lymphoblastic leukemia in children by the 15th day of induction therapy according to the ALL-MB-2008 protocol. Immunologiya. 2010;31(2):60–4. (In Russ)]
  32. Мовчан Л.В., Шман Т.В., Белевцев М.В. и др. Изменение иммунофенотипа лейкемических клеток на этапах индукционной терапии острого лимфобластного лейкоза из предшественников В-лимфоцитов у детей. Вопросы гематологии/онкологии и иммунопатологии в педиатрии. 2011;10(1):21–6. [Movchan LV, Shman TV, Belevtsev MV, et al. Immunophenotypic modulation of the leukemic cells during induction therapy in children with B-cell precursor acute lymphoblastic leukemia. Voprosy gematologii/onkologii i immunopatologii v pediatrii. 2011;10(1):21–6. (In Russ)]
  33. Dworzak MN, Gaipa G, Schumich A, et al. Modulation of antigen expression in B-cell precursor acute lymphoblastic leukemia during induction therapy is partly transient: evidence for a drug-induced regulatory phenomenon. Results of the AIEOP-BFM-ALL-FLOW-MRD-Study Group. Cytometry B Clin Cytom. 2010;78В(3):147–53. doi: 10.1002/cyto.b.20516.
  34. Borowitz MJ, Pullen DJ, Winick N, et al. Comparison of diagnostic and relapse flow cytometry phenotypes in childhood acute lymphoblastic leukemia: implications for residual disease detection: a report from the Children’s Oncology Group. Cytometry B Clin Cytom. 2005;68В(1):18–24. doi: 1002/cyto.b.20071.
  35. Liu YR, Chang Y, Fu JY, et al. Comparison of the immunophenotype of patients with B lineage acute lymphoblastic leukemia at diagnosis and relapse. Zhonghua Xue Ye Xue Za Zhi. 2006;27(5):335–8.
  36. Dworzak MN, Froschl G, Printz D, et al. Prognostic significance and modalities of flow cytometric minimal residual disease detection in childhood acute lymphoblastic leukemia. Blood. 2002;99(6):1952–8. doi: 10.1182/blood.v99.6.1952.
  37. Coustan-Smith E, Ribeiro RC, Stow P, et al. A simplified flow cytometric assay identifies children with acute lymphoblastic leukemia who have a superior clinical outcome. Blood. 2006;108(1):97–102. doi: 1182/blood-2006-01-0066.
  38. Попов А.М., Вержбицкая Т.Ю., Цаур Г.А. и др. Ограниченная возможность применения упрощенного подхода для определения минимальной остаточной болезни методом проточной цитометрии у детей с острым лимфобластным лейкозом из B-линейных предшественников. Клиническая лабораторная диагностика. 2011;3:25–9.
    [Popov AM, Verzhbitskaya TYu, Tsaur GA, et al. Limited potential for use of simplified approach for determining minimal residual disease by means of flow cytometry in children with acute lymphoblastic leukemia from B-linear precursors. Klinicheskaya laboratornaya diagnostika. 2011;3:25–9. (In Russ)]
  39. Porwit-MacDonald A, Bjorklund E, Lucio P, et al. BIOMED-1 concerted action report: flow cytometric characterization of CD7+ cell subsets in normal bone marrow as a basis for the diagnosis and follow-up of T cell acute lymphoblastic leukemia (T-ALL). Leukemia. 2000;14(5):816–25. doi: 1038/sj.leu.2401741.
  40. Dworzak MN, Fritsch G, Buchinger P, et al. Flow cytometric assessment of human MIC2 expression in bone marrow, thymus, and peripheral blood. Blood. 1994;83(2):415–25.
  41. Dworzak MN, Fritsch G, Fleischer C, et al. CD99 (MIC2) expression in paediatric B-lineage leukaemia/lymphoma reflects maturation-associated patterns of normal B-lymphopoiesis. Br J Haematol. 1999;105(3):690–5. doi:1046/j.1365-2141.1999.01426.x.
  42. Dworzak MN, Froschl G, Printz D, et al. CD99 expression in T-lineage ALL: implications for flow cytometric detection of minimal residual disease. Leukemia. 2004;18(4):703–8. doi:1038/sj.leu.2403303.
  43. Roshal M, Fromm JR, Winter S, et al. Immaturity associated antigens are lost during induction for T cell lymphoblastic leukemia: implications for minimal residual disease detection. Cytometry B Clin Cytom. 2010;78В(3):139–46. doi: 10.1002/cyto.b.20511.
  44. Lund-Johansen F, Terstappen LW. Differential surface expression of cell adhesion molecules during granulocyte maturation. J Leuk Biol. 1993;54(1):47–55.
  45. Terstappen LW, Huang S, Picker LJ. Flow cytometric assessment of human T-cell differentiation in thymus and bone marrow. B 1992;79(3):666–77.
  46. Aalbers AM, van den Heuvel-Eibrink MM, Baumann I, et al. Bone marrow immunophenotyping by flow cytometry in refractory cytopenia of childhood. Haematologica. 2015;100(3):315–23. doi: 10.3324/haematol.2014.107706.
  47. Feng B, Verstovsek S, Jorgensen JL, Lin P. Aberrant myeloid maturation identified by flow cytometry in primary myelofibrosis. Am J Clin Pathol. 2010;133(2):314–20. doi: 10.1309/AJCPNC99DHXIOOTD.
  48. Loken MR, Chu S-Ch, Fritschle W, et al. Normalization of bone marrow aspirates for hemodilution in flow cytometric analyses. Cytometry B Clin Cytom. 2009;76В(1):27–36. doi: 10.1002/cyto.b.20429.
  49. Kussick SJ, Wood BL. Using 4-color flow cytometry to identify abnormal myeloid populations. Arch Pathol Lab Med. 2003;127(9):1140–7.
  50. Leandro MJ, Cooper N, Cambridge G, et al. Bone marrow B-lineage cells in patients with rheumatoid arthritis following rituximab therapy. Rheumatology (Oxford). 2007;46(1):29–36. doi: 10.1093/rheumatology/kel148.
  51. Rehnberg M, Amu S, Tarkowski A, et al. Short- and long-term effects of anti-CD20 treatment on B cell ontogeny in bone marrow of patients with rheumatoid arthritis. Arthritis Res Ther. 2009;11(4):R123. doi: 10.1186/ar2789.
  52. Nakou M, Katsikas G, Sidiropoulos P, et al. Rituximab therapy reduces activated B cells in both the peripheral blood and bone marrow of patients with rheumatoid arthritis: depletion of memory B cells correlates with clinical response. Arthritis Res Ther. 2009;11(4):R131. doi: 10.1186/ar2798.
  53. Borowitz MJ. Minimal residual disease detection in childhood ALL. Haematopoiesis Immunology. 2010;7(1):24–35.
  54. Вержбицкая Т.Ю., Попов А.М., Томилов А.Ф. и др. Определение опухолевых клеток в спинномозговой жидкости у детей с острыми лейкозами методом проточной цитометрии. Вопросы диагностики в педиатрии. 2012;5:31–5.
    [Verzhbitskaya TYu, Popov AM, Tomilov AF, et al. Detection of tumor cells in cerebrospinal fluid in children with acute leukemias using flow cytometry. Voprosy diagnostiki v pediatrii. 2012;5:31–5. (In Russ)]


Chronic Lymphocytic Leukemia: Prognostic Significance of Minimal Residual Disease and Potential of Modern Methods of Its Diagnosis and Therapy (Literature Review)

AYu Kuvshinov, SV Voloshin, IS Martynkevich, EV Kleina, MA Mikhaleva, KM Abdulkadyrov

Russian Scientific Research Institute of Hematology and Transfusiology, 16 2-ya Sovetskaya str., Saint Petersburg, Russian Federation, 191024

For correspondence: Sergei Vladimirovich Voloshin, PhD, 16 2-ya Sovetskaya str., Saint Petersburg, Russian Federation, 191024; Tel.: +7(812)274-37-70; e-mail: kuvshinovmd@gmail.com

For citation: Kuvshinov AYu, Voloshin SV, Martynkevich IS, et al. Chronic Lymphocytic Leukemia: Prognostic Significance of Minimal Residual Disease and Potential of Modern Methods of Its Diagnosis and Therapy (Literature Review). Clinical oncohematology. 2016;9(2):191–8 (In Russ).

DOI: 10.21320/2500-2139-2016-9-2-191-198


ABSTRACT

Achieving a complete remission (CR) in patients with chronic lymphocytic leukemia (CLL) has become a feasible goal directly correlating with a prolonged survival. However, a certain number of tumor cells may be present in the patient’s body even when CR has been achieved, and this phenomenon is called a minimal residual disease (MRD). A lot of data confirming the necessity of MRD diagnosing and monitoring has emerged recently, since the MRD has a significant impact on the prognosis of CLL. Achieving MRD-negative remission is an independent predictor of long-term progression-free survival and overall survival. The occurrence of new diagnostic techniques has allowed to define the MRD and to develop standards for its assessment. This paper presents an overview of literature data about MRD, methods of its evaluation, prognostic significance, as well as the methods of eradication.


Keywords: chronic lymphocytic leukemia, minimal residual disease, flow cytometry.

Received: January 5, 2016

Accepted: January 10, 2016

Read in PDF (RUS)pdficon


REFERENCES

  1. Hallek M, Cheson BD, Catovsky D, et al. Guidelines for the diagnosis and treatment of chronic lymphocytic leukemia: a report from the International Workshop on Chronic Lymphocytic Leukemia updating the National Cancer Institute-Working Group 1996 guidelines. Blood. 2008;111(12):5446–56. doi: 10.1182/blood-2007-06-093906.
  2. Cave H, van der Werff ten Bosch J, Suciu S, 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(9):591–8. doi: 10.1056/nejm199808273390904.
  3. Andersen NS, Pedersen LB, Laurell A, et al. Preemptive treatment with rituximab of molecular relapse after autologous stem cell transplantation in mantle cell lymphoma. J Clin Oncol. 2009;27(26):4365–70. doi: 10.1200/JCO.2008.21.3116.
  4. Grimwade D, Lo Coco F. Acute promyelocytic leukemia: a model for the role of molecular diagnosis and residual disease monitoring in directing treatment approach in acute myeloid leukemia. Leukemia. 2002;16(10):1959–73. doi: 10.1038/sj.leu.2402721.
  5. Vora A, Goulden N, Mitchell C, et al. Augmented post-remission therapy for a minimal residual disease-defined high-risk subgroup of children and young people with clinical standard-risk and intermediate-risk acute lymphoblastic leukaemia (UKALL 2003): a randomised controlled trial. Lancet Oncol. 2014;15(8):809–18. doi: 10.1016/S1470-2045(14)70243-8.
  6. Moreton P, Kennedy B, Lucas G, et al. Eradication of minimal residual disease in B-cell chronic lymphocytic leukemia after alemtuzumab therapy is associated with prolonged survival. J Clin Oncol. 2005;23(13):2971–9. doi: 10.1200/jco.2005.04.021.
  7. Ritgen M, Bottcher S, Stilgenbauer S, et al. Quantitative MRD monitoring identifies distinct GVL response patterns after allogeneic stem cell transplantation for chronic lymphocytic leukemia: results from the GCLLSG CLL3X trial. Leukemia. 2008;22(7):1377–86. doi: 10.1038/leu.2008.96.
  8. Del Poeta G, Del Principe MI, Buccisano F, et al. Consolidation and maintenance immunotherapy with rituximab improve clinical outcome in patients with B-cell chronic lymphocytic leukemia. Cancer. 2008;112(1):119–28. doi: 10.1002/cncr.23144.
  9. Rawstron AC, Kennedy B, Moreton P, et al. Early prediction of outcome and response to alemtuzumab therapy in chronic lymphocytic leukemia. Blood. 2004;103(6):2027–31. doi: 10.1182/blood-2002-10-3270.
  10. Bosch F, Ferrer A, Villamor N, et al. Fludarabine, cyclophosphamide, and mitoxantrone as initial therapy of chronic lymphocytic leukemia: high response rate and disease eradication. Clin Cancer Res. 2008;14(1):155–61. doi: 10.1158/1078-0432.CCR-07-1371.
  11. Kay NE, Geyer SM, Call TG, et al. Combination chemoimmunotherapy with pentostatin, cyclophosphamide, and rituximab shows significant clinical activity with low accompanying toxicity in previously untreated B chronic lymphocytic leukemia. Blood. 2007;109(2):405–11. doi: 10.1182/blood-2006-07-033274.
  12. Ritgen M, Lange A, Stilgenbauer S, et al. Unmutated immunoglobulin variable heavy-chain gene status remains an adverse prognostic factor after autologous stem cell transplantation for chronic lymphocytic leukemia. Blood. 2003;101(5):2049–53. doi: 10.1182/blood-2002-06-1744.
  13. Hillmen P, Skotnicki AB, Robak T, et al. Alemtuzumab compared with chlorambucil as first-line therapy for chronic lymphocytic leukemia. J Clin Oncol. 2007;25(35):5616–23. doi: 10.1200/jco.2007.12.9098.
  14. Robertson LE, Huh YO, Butler JJ, et al. Response assessment in chronic lymphocytic leukemia after fludarabine plus prednisone: clinical, pathologic, immunophenotypic, and molecular analysis. Blood. 1992;80:29–36.
  15. O’Brien SM, Kantarjian HM, Cortes J, et al. Results of the fludarabine and cyclophosphamide combination regimen in chronic lymphocytic leukemia. J Clin Oncol. 2001;19:1414–20.
  16. Tam CS, O’Brien S, Wierda W, et al. Long-term results of the fludarabine, cyclophosphamide, and rituximab regimen as initial therapy of chronic lymphocytic leukemia. Blood. 2008;112(4):975–80. doi: 10.1182/blood-2008-02-140582.
  17. Robak T, Blonski JZ, Gora-Tybor J, et al. Cladribine alone and in combination with cyclophosphamide or cyclophosphamide plus mitoxantrone in the treatment of progressive chronic lymphocytic leukemia: report of a prospective, multicenter, randomized trial of the Polish Adult Leukemia Group (PALG CLL2). Blood. 2006;108(2):473–9. doi: 10.1182/blood-2005-12-4828.
  18. Moreno C, Villamor N, Colomer D, et al. Clinical significance of minimal residual disease, as assessed by different techniques, after stem cell transplantation for chronic lymphocytic leukemia. Blood. 2006;107(11):4563–9. doi: 10.1182/blood-2005-09-3634.
  19. Milligan DW, Fernandes S, Dasgupta R, et al. Results of the MRC pilot study show autografting for younger patients with chronic lymphocytic leukemia is safe and achieves a high percentage of molecular responses. Blood. 2005;105(1):397–404. doi: 10.1182/blood-2004-01-0298.
  20. Bottcher S, Ritgen M, Pott C, et al. Comparative analysis of minimal residual disease detection using four-color flow cytometry, consensus IgH-PCR, and quantitative IgH PCR in CLL after allogeneic and autologous stem cell transplantation. Leukemia. 2004;18(10):1637–45. doi: 10.1038/sj.leu.2403478.
  21. Rawstron AC, Kennedy B, Evans PA, et al. Quantitation of minimal disease levels in chronic lymphocytic leukemia using a sensitive flow cytometric assay improves the prediction of outcome and can be used to optimize therapy. Blood. 2001;98(1):29–35. doi: 10.1182/blood.v98.1.29.
  22. Maloum K, Sutton L, Baudet S, et al. Novel flow-cytometric analysis based on BCD5+ subpopulations for the evaluation of minimal residual disease in chronic lymphocytic leukaemia. Br J Haematol. 2002;119(4):970–5. doi: 10.1046/j.1365-2141.2002.03956.x.
  23. Никитин Е.А. Дифференцированная терапия хронического лимфолейкоза: Дис. ¼ д-ра мед. наук. М., 2014. 203 с.
    [Nikitin EA. Differentsirovannaya terapiya khronicheskogo limfoleikoza. (Differentiated therapy of chronic lymphocytic leukemia.) [dissertation] Moscow; 2014. 203 p. (In Russ)]
  24. Ripolles L, Ortega M, Ortuno F, et al. Genetic abnormalities and clinical outcome in chronic lymphocytic leukemia. Cancer Genet Cytogenet. 2006;171(1):57–64. doi: 10.1016/j.cancergencyto.2006.07.006.
  25. Van Dongen JJ, Langerak AW, Bruggemann M, et al. Design and standardization of PCR primers and protocols for detection of clonal immunoglobulin and T-cell receptor gene recombinations in suspect lymphoproliferations: report of the BIOMED-2 Concerted Action BMH4-CT98-3936. Leukemia. 2003;17(12):2257–317. doi: 10.1038/sj.leu.2403202.
  26. Rawstron AC, Villamor N, Ritgen M, et al. International standardized approach for flow cytometric residual disease monitoring in chronic lymphocytic leukaemia. Leukemia. 2007;21(5):956–64. doi: 10.1038/sj.leu.2404584.
  27. Keating MJ, O’Brien S, Albitar M, et al. Early results of a chemoimmunotherapy regimen of fludarabine, cyclophosphamide, and rituximab as initial therapy for chronic lymphocytic leukemia. J Clin Oncol. 2005;23(18):4079–88. doi: 10.1200/jco.2005.12.051.
  28. Byrd JC, Peterson BL, Morrison VA, et al. Randomized phase 2 study of fludarabine with concurrent versus sequential treatment with rituximab in symptomatic, untreated patients with B-cell chronic lymphocytic leukemia: Results from Cancer and Leukemia Group B 9712 (CALGB 9712). Blood. 2003;101(1):6–14. doi: 10.1182/blood-2002-04-1258.
  29. Eichhorst BF, Busch R, Hopfinger G, et al. Fludarabine plus cyclophosphamide versus fludarabine alone in first-line therapy of younger patients with chronic lymphocytic leukemia. Blood. 2006;107(3):885–91. doi: 10.1182/blood-2005-06-2395.
  30. Cheson BD, Bennett JM, Grever M, et al. National Cancer Institute-sponsored Working Group guidelines for chronic lymphocytic leukemia: revised guidelines for diagnosis and treatment. Blood. 1996;87:4990–7.
  31. Dreger P, Ritgen M, Bottcher S, et al. The prognostic impact of minimal residual disease assessment after stem cell transplantation for chronic lymphocytic leukemia: Is achievement of molecular remission worthwhile? Leukemia. 2005;19(7):1135–8. doi: 10.1038/sj.leu.2403800.
  32. Wendtner CM, Ritgen M, Schweighofer CD, et al. Consolidation with alemtuzumab in patients with chronic lymphocytic leukemia (CLL) in first remission–experience on safety and efficacy within a randomized multicenter phase III trial of the German CLL Study Group (GCLLSG). Leukemia. 2004;18(6):1093–101. doi: 10.1038/sj.leu.2403354.
  33. Montillo M, Tedeschi A, Miqueleiz S, et al. Alemtuzumab as consolidation after a response to fludarabine is effective in purging residual disease in patients with chronic lymphocytic leukemia. J Clin Oncol. 2006;24(15):2337–42. doi: 10.1200/jco.2005.04.6037.
  34. Rawstron AC, de Tute R, Jack AS, et al. Flow cytometric protein expression profiling as a systematic approach for developing disease-specific assays: identification of a chronic lymphocytic leukaemia-specific assay for use in rituximab-containing regimens. Leukemia. 2006;20(12):2102–10. doi: 10.1038/sj.leu.2404416.
  35. Hallek M, Fingerle-Rowson G, Fink A, et al. Immunochemotherapy with fludarabine (F), cyclophosphamide (C), and rituximab (R) (FCR) versus fludarabine and cyclophosphamide (FC) improves response rates and progression-free survival (PFS) of previously untreated patients (pts) with advanced chronic lymphocytic leukemia (CLL). Blood. 2008;112: Abstract 325.
  36. Wierda W, O’Brien S, Wen S, et al. Chemoimmunotherapy with fludarabine, cyclophosphamide, and rituximab for relapsed and refractory chronic lymphocytic leukemia. J Clin Oncol. 2005;23(18):4070–8. doi: 10.1200/jco.2005.12.516.
  37. Bottcher S, Stilgenbauer S, Busch R, et al: Standardized MRD flow and ASO IGH RQ-PCR for MRD quantification in CLL patients after rituximab-containing immunochemotherapy: A comparative analysis. Leukemia. 2009;23(11):2007–17. doi: 10.1038/leu.2009.140.
  38. Ringelstein-Harlev S, Fineman R. Minimal Residual Disease Surveillance in Chronic Lymphocytic Leukemia by Fluorescence-Activated Cell Sorting. Rambam Maimonides Med J. 2014;5(4): e0027. doi: 10.5041/RMMJ.10161.
  39. Bottcher S, Ritgen M, Fischer K, et al. Minimal residual disease quantification is an independent predictor of progression-free and overall survival in chronic lymphocytic leukemia: a multivariate analysis from the randomized GCLLSG CLL8 trial. J Clin Oncol. 2012;30(9):980–8. doi: 10.1200/JCO.2011.36.9348.
  40. Kovacs G, Bottcher S, Bahlo J, et al. Value of minimal residual disease (MRD) negative status at response evaluation in chronic lymphocytic leukemia (CLL): combined analysis of two phase III studies of the German CLL Study Group (GCLLSG). ASH Annual Meeting Abstracts. 2014: Abstract 23.
  41. Eichhorst B, Fink AM, Busch R, et al. Frontline chemoimmunotherapy with fludarabine (F), cyclophosphamide (C), and rituximab (R) (FCR) shows superior efficacy in comparison to bendamustine (B) and rituximab (BR) in previously untreated and physically fit patients (pts) with advanced chronic lymphocytic leukemia (CLL): Final analysis of an international, randomized study of the German CLL Study Group (GCLLSG) (CLL10 study). Proc ASH 2014: Abstract 19.
  42. Garifullin A, Kuvshinov A, Voloshin S, et al. The frequency of occurrence of minimal residual disease (MRD) into different prognostic groups of patients with chronic lymphocytic leukemia (CLL). Intern Hematol Club. [Internet] 2015 Nov 6–7 [cited 2016 April 18] Available from: http://www.comtecmed.com/IHC/2015/poster_list.aspx.
  43. Schweighofer CD, Ritgen M, Eichhorst BF, et al. Consolidation with alemtuzumab improves progression-free survival in patients with chronic lymphocytic leukaemia (CLL) in first remission: long-term follow-up of a randomized phase III trial of the German CLL Study Group (GCLLSG). Br J Haematol. 2009;144(1):95–8. doi: 10.1111/j.1365-2141.2008.07394.x.
  44. Wiernik PH, Adiga GU. Single-agent rituximab in treatment-refractory or poor prognosis patients with chronic lymphocytic leukemia. Curr Med Res Opin. 2011;27(10):1987–93. doi: 10.1185/03007995.2011.615307.
  45. Abrisqueta P, Villamor N, Terol MJ, et al. Rituximab maintenance after first-line therapy with rituximab, fludarabine, cyclophosphamide, and mitoxantrone (R-FCM) for chronic lymphocytic leukemia. Blood. 2013;122(24):3951–9. doi: 10.1182/blood-2013-05-502773.
  46. Shanafelt TD, Ramsay AG, Zent CS, et al. Long-term repair of T-cell synapse activity in a phase II trial of chemoimmunotherapy followed by lenalidomide consolidation in previously untreated chronic lymphocytic leukemia (CLL). Blood. 2013;121(20):4137–41. doi: 10.1182/blood-2012-12-470005.
  47. Dreger P, Dohner H, Ritgen M, et al. Allogeneic stem cell transplantation provides durable disease control in poor-risk chronic lymphocytic leukemia: long-term clinical and MRD results of the German CLL Study Group CLL3X trial. Blood. 2010;116(14)2438–47. doi: 10.1182/blood-2010-03-275420.
  48. Byrd JC, Furman RR, Coutre SE, et al. Targeting BTK with ibrutinib in relapsed chronic lymphocytic leukemia. N Engl J Med. 2013;369(1):32–42. doi: 10.1056/NEJMoa1215637.
  49. O’Brien S, Furman RR, Coutre SE, et al. Ibrutinib as initial therapy for elderly patients with chronic lymphocytic leukaemia or small lymphocytic lymphoma: an open-label, multicentre, phase 1b/2 trial. Lancet Oncol. 2014;15(1):48–58. doi: 10.1016/S1470-2045(13)70513-8.
  50. Furman RR, Sharman JP, Coutre SE, et al. Idelalisib and rituximab in relapsed chronic lymphocytic leukemia. N Engl J Med. 2014;370(11):997–1007. doi: 10.1056/NEJMoa1315226.
  51. O’Brien S, Lamanna N, Kipps TJ, et al. Update on a Phase 2 Study of Idelalisib in Combination with Rituximab in Treatment-Naive Patients ³65 Years with Chronic Lymphocytic Leukemia (CLL) or Small Lymphocytic Lymphoma (SLL). Blood. 2014;124: Abstract 1994.