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ICCS e-newsletter CSI Spring 2012 Weina Chen, MD, PhD Medical Director, Hematopathology Ameripath/Quest Diagnostics Dallas, Texas.

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Presentation on theme: "ICCS e-newsletter CSI Spring 2012 Weina Chen, MD, PhD Medical Director, Hematopathology Ameripath/Quest Diagnostics Dallas, Texas."— Presentation transcript:

1 ICCS e-newsletter CSI Spring 2012 Weina Chen, MD, PhD Medical Director, Hematopathology Ameripath/Quest Diagnostics Dallas, Texas

2 Case History The patient is a 70-year old female presented with mild leukocytosis. She has no prior history of any significant diseases and is asymptomatic.

3 Complete blood count WBC 10.70 NE 50% LY 39.4% MO 9.3% EO 0.7% BASO 0.6% RBC 4.41 HGB 12.7 HCT 37.7% MCV 85.5 MCHC 33.70 RDW 12.6 PLT 211.0

4 Work-up and evaluation Bone marrow (BM) aspirate and biopsy were procured. Flow cytometric analysis was performed on marrow aspirate and results from selected 4-color tubes are provided for review.

5 Flow cytometric analysis Acquisition Beckman Coulter Epics XL (FCS2.0, System II) Analyzed by Paint-A-Gate software (adapted to Coulter) Tubes (FITC/PE/ECD/PC5) –Tube 1: Kappa/lambda/45/19+20 –Tube 2: 5/19/45/10 –Tube 3 : 8/4/45/38 –Tube 4: 15/117/45/34 –Tube 5: 20/10/19/38 –Tube 6: FMC-7/23/5/19 –Tube 7: Kappa/Lambda/5/19

6 Tube 1: Kappa/lambda/45/19+20


8 Tube 2: 5/19/45/10

9 Tube 3 : 8/4/45/38

10 Tube 4: 15/117/45/34

11 Tube 5: 20/10/19/38

12 Tube 6: FMC-7/23/5/19

13 Tube 7: Kappa/Lambda/5/19 [In addition, CD13 -, CD33 - (data not shown); Tdt not tested]

14 Key flow plots in this case CD19(+)/CD20(+) B cells overall exhibiting a pattern of sequential maturation

15 Morphologic evaluation Marrow infiltrated by abundant small to medium-sized lymphoid cells with mature morphologic features although nuclear irregularity/convolution and small cytoplasmic vacuoles observed in a few scattered lymphoid cells.

16 CD20 CD34 CD79a Tdt CD10 Immunohistochemical evaluation A prominent CD79a(+) B-lymphoid hyperplasia of mostly CD10(+) B lymphocytes with increased Tdt(+) cells, some in clusters exceeding 3 or 4 cells


18 Questions… There is an expansion of B cells overall exhibiting a spectrum of maturation. Are these normal maturing B-cell precursors (hematogones) or B-lymphoblasts?

19 A few words on hematogones… Hematogones always express consistent, reproducible, complex spectrum of sequential antigen expression and lack aberrant antigen expression. This defines hematogones into three stages of maturation –Stage 1 hematogones express CD34, high levels of CD10 and CD38, a moderate level of CD22, and absence of CD20. –Intermediate stage 2 hematogones downregulate CD34 completely and CD10 partially, while increasing expression of CD22 and CD20. –Stage 3 hematogones upregulate CD20 expression reaching the intensity of mature B cells, and CD10 and CD38 are slightly down-regulated with increasing expression of polytypic surface immunoglobulin light chains. –Subsequently, these cells mature into CD20(+), CD10(-) mature B cells. –CD5 is expressed on normal, polytypic B cells in a continuum, predominantly at later stages of maturation, specifically on stage 3 hematogones and mature B cells.

20 Comparison to a case with hematogone hyperplasia A case with hematogone (HG) hyperplasia case (bottom plots): Blue, mature B cells; Green, stage 2+3 HG; yellow, stage I HG

21 Comparison to a case with hematogone hyperplasia A case with hematogone (HG) hyperplasia case (bottom plots): Blue, mature B cells; Green, stage 2+3 HG; yellow, stage I HG

22 Questions… These B cells exhibit a spectrum of maturation reminiscent of hematogones and unusual for neoplastic lymphoblasts. Are these hematogones???

23 Answer… No These are B-lymphoblasts. The key finding in this case (on BM sample) – Cytogenetics: 46, XX, t(9;22)(q34;q11.2)[17]/46, XX [3] – Positive FISH for t(9;22)/BCR-ABL1 in 79% of interphase cells

24 Answer… Differential diagnosis – An early chronic myelogenous leukemia (CML) with background hematogone hyperplasia (but the usual morphologic features of CML not apparent) – Lymphoid blast crisis of CML (but no history of CML) – An early B-lymphoblastic leukemia with t(9;22)(q34;q11.2);BCR-ABL1

25 Favored Diagnosis B-lymphoblastic leukemia with t(9;22)(q34;q11.2);BCR-ABL1 Based on the high percent of t(9;22) positive cells (~70%), the entire B-cell population or the majority of B cells including polytypic B cells seems neoplastic.

26 A few words on B-lymphoblastic leukemia with t(9;22)(q34;q11.2);BCR-ABL1 The most frequently observed chromosomal abnormality in adult B- ALL (25% vs. 3-5% in children) Involving the ABL1 oncogene on chromosome 9 and the guanosine triphosphate–binding protein BCR on chromosome 22 The resultant fusion protein having abnormal tyrosine kinase activity, leading to disturbances in proliferation, survival, and adhesion In about 70% of cases of BCR-ABL1 + B-ALL, the expressed protein being 190 kDa, rather than the 210 kDa typically seen in CML Associated with a poor prognosis in both children and adults

27 Unusual features in this case Unusual presentation: close to normal CBC with differential at presentation Unusual morphology: mature morphologic features with mild cytological atypia Unusual immunophenotype: maturation spectrum reminiscent of hematogones (with only subtle deviation) Unusual, indolent clinical course – Follow-up BM in 5 months (with only imatinib mesylate tx) Close to normal CBC, asymptomatic Persistent, but decreased B-lymphoblasts (similar phenotype) RT-PCR: positive BCR-ABL1, p190, further supporting B- ALL

28 What are the clues to avoid misdiagnosis? No apparent causes for hematogone hyperplasia Common causes for hematogone hyperplasia: Reactive conditions: AIDS, immune dysregulation, copper deficiency), BM involved by metastatic tumors Regenerative conditions: post-chemotherapy and stem- cell transplant Relatively high number of hematogones in children Subtle immunophenotypic deviation from hematogones Less distinct “ladder” of CD45 on subsets of B cells Tdt positive cells, some in clusters exceeding 3 or 4 cells The need to add new markers to distinguish hematogones from lymphoblasts CD81, CD123

29 Take home messages The immunophenotype of B-lymphoblasts is variable. While the majority of cases having distinct immunophenotypic aberration deviated from hematogones, rare cases with immunophenotypic feature reminiscent of hematogones do exist. Careful immunophenotypic analysis, clinical correlation for causes of hematogone hyperplasia, ancillary studies (cytogenetics, FISH/molecular studies) are the key elements to reach a correct diagnosis.

30 References 1. Weir EG, Cowan K, LeBeau P, Borowitz MJ. A limited antibody panel can distinguish B-precursor acute lymphoblastic leukemia from normal B precursors with four color flow cytometry: implications for residual disease detection. Leukemia 1999;13:558-67. 2. McKenna RW, Washington LT, Aquino DB, Picker LJ, Kroft SH. Immunophenotypic analysis of hematogones (B-lymphocyte precursors) in 662 consecutive bone marrow specimens by 4-color flow cytometry. Blood 2001;98:2498-507. 3. McKenna RW, Asplund SL, Kroft SH. Immunophenotypic analysis of hematogones (B-lymphocyte precursors) and neoplastic lymphoblasts by 4-color flow cytometry. Leuk Lymphoma 2004;45:277-85. 4. Chen W, Karandikar NJ, McKenna RW, Kroft SH. Stability of leukemia-associated immunophenotypes in precursor B-lymphoblastic leukemia/lymphoma: a single institution experience. Am J Clin Pathol 2007;127:39-46. 5. 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;132:940-9. 6. Loken MR, Shah VO, Dattilio KL, Civin CI. Flow cytometric analysis of human bone marrow. II. Normal B lymphocyte development. Blood 1987;70:1316-24. 7. Ryan DH, Chapple CW, Kossover SA, Sandberg AA, Cohen HJ. Phenotypic similarities and differences between CALLA-positive acute lymphoblastic leukemia cells and normal marrow CALLA- positive B cell precursors. Blood 1987;70:814-21. 8. Campana D, Coustan-Smith E. Detection of minimal residual disease in acute leukemia by flow cytometry. Cytometry 1999;38:139-52. 9. 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;132:733-7.

31 10. Hurwitz CA, Gore SD, Stone KD, Civin CI. Flow cytometric detection of rare normal human marrow cells with immunophenotypes characteristic of acute lymphoblastic leukemia cells. Leukemia 1992;6:233-9. 11. Hurwitz CA, Loken MR, Graham ML, et al. Asynchronous antigen expression in B lineage acute lymphoblastic leukemia. Blood 1988;72:299-307. 12. Kurec AS, Belair P, Stefanu C, Barrett DM, Dubowy RL, Davey FR. Significance of aberrant immunophenotypes in childhood acute lymphoid leukemia. Cancer 1991;67:3081-6. 13. Muzzafar T, Medeiros LJ, Wang SA, Brahmandam A, Thomas DA, Jorgensen JL. Aberrant underexpression of CD81 in precursor B-cell acute lymphoblastic leukemia: utility in detection of minimal residual disease by flow cytometry. Am J Clin Pathol 2009;132:692-8. 14. Hassanein NM, Alcancia F, Perkinson KR, Buckley PJ, Lagoo AS. Distinct expression patterns of CD123 and CD34 on normal bone marrow B-cell precursors ("hematogones") and B lymphoblastic leukemia blasts. Am J Clin Pathol 2009;132:573-80. 15. Muehleck SD, McKenna RW, Gale PF, Brunning RD. Terminal deoxynucleotidyl transferase (TdT)- positive cells in bone marrow in the absence of hematologic malignancy. Am J Clin Pathol 1983;79:277-84. 16. Rimsza LM, Larson RS, Winter SS, et al. Benign hematogone-rich lymphoid proliferations can be distinguished from B-lineage acute lymphoblastic leukemia by integration of morphology, immunophenotype, adhesion molecule expression, and architectural features. Am J Clin Pathol 2000;114:66-75. 17. Sutton L, Vusirikala M, Chen W. Hematogone hyperplasia in copper deficiency. Am J Clin Pathol 2009;132:191-9; quiz 307.

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