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ICCS e-newsletter CSI Spring 2012

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

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ICCS e-newsletter CSI Spring 2012

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  1. ICCS e-newsletter CSISpring 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 • A population of (CD19/20)+ B cells divided into 3 subsets based on intensity of CD45 • Blue (strong CD45) • Violet (intermediate CD45) • Cyan (dim CD45)

  7. Tube 1: Kappa/lambda/45/19+20 • CD(19/20)+ B cells showing variable, polytypic light chain expression • Blue (strong CD45): polytypic surface Ig expression • Violet (intermediate CD45): a subset polytypic sIg • Cyan (dim CD45): no surface Ig

  8. Tube 2: 5/19/45/10 • Differential expression of CD5 and CD10 on 3 subsets of B cells • Blue (subset CD5+, CD10-) • Violet (CD5-, CD10+) • Cyan (CD5-, CD10bright+)

  9. Tube 3 : 8/4/45/38 • Differential expression of CD38 on 3 subsets of B cells • Blue (CD38variable/partial +) • Violet (CD38uniform strong +) • Cyan (CD38+)

  10. Tube 4: 15/117/45/34 • Differential expression of CD34 on 3 subsets of B cells • Blue (CD34-) • Violet (CD34-) • Cyan (CD38+) • Of note, myeloblasts in red express CD117 and CD34

  11. Tube 5: 20/10/19/38 • Differential expression of CD10, 20 and CD38 on 3 subsets of CD19(+) B cells • Blue (CD10-, CD20+, CD38variably/partial+) • Violet (CD10+, CD20variably+, CD38uniform +) • Cyan (CD10bright+, CD20-, CD38+)

  12. Tube 6: FMC-7/23/5/19 • Differential expression of CD5, CD23 and FMC7 on 3 subsets of CD19(+) B cells • Blue (CD5subset+, CD23partial +, FMC7+) • Violet (CD5subset+, CD23-, FMC7-) • Cyan (CD5-, CD23-, FMC7-)

  13. Tube 7: Kappa/Lambda/5/19 • Differential expression of surface Ig and CD5 on subsets of CD19(+) B cells • Blue (CD5+, sIgpolytypic) • Violet (CD5predominantly -, sIg-) [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. Immunohistochemical evaluation CD20 CD79a CD34 Tdt CD10 A prominent CD79a(+) B-lymphoid hyperplasia of mostly CD10(+) B lymphocytes with increased Tdt(+) cells, some in clusters exceeding 3 or 4 cells

  17. 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?

  18. 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.

  19. 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

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

  22. 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

  23. 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

  24. 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.

  25. 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 childrenand adults

  26. 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

  27. 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

  28. 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.

  29. 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.

  30. 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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