Sreedharanunn

Morphologic Confounders and CD19 Negativity in a Case of Hairy Cell Leukemia

Pulkit Rastogi, Sreejesh Sreedharanunni, Uday Yanamandra, Man Updesh Singh Sachdeva and Neelam Varma

Postgraduate Institute of Medical Education and Research (PGIMER), Chandigarh - 160012, India. 

Corresponding author: Dr. Sreejesh Sreedharanunni, MD, DM. Assistant Professor, Dept of Hematology, 5th Floor, Research Block A, Postgraduate Institute of Medical Education and Research, Chandigarh – 160012, India. Tel: +91-9478053220, Fax: +91-172-2747124. sreejesh.s@pgimer.edu.in 

Published: May 1, 2017
Received: January 23, 2017
Accepted: April 1, 2017
Mediterr J Hematol Infect Dis 2017, 9(1): e2017033 DOI 10.4084/MJHID.2017.033
This article is available on PDF format at: 

This is an Open Access article distributed under the terms of the Creative Commons Attribution License
(
https://creativecommons.org/licenses/by-nc/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Objectives: We report a case of hairy cell leukemia (HCL) initially misdiagnosed as plasma cell dyscrasia due to various clinical, morphological and immunophenotypic confounders.
Methods and results: In a patient diagnosed of marrow plasmacytosis and serum monoclonal protein elsewhere and referred to our hospital, morphological evaluation of bone marrow aspirate smears and trephine biopsy, immunophenotyping, and molecular testing (BRAFV600E mutation) were done. Clinically, the patient was asymptomatic; bone marrow revealed plasmacytosis, mastocytosis, and lymphocytosis with a few "hairy" cells. Immunophenotyping showed features of HCL with aberrant CD10 expression and a large subclone of CD19neg cells. A diagnosis of HCL with reactive plasmacytosis and mast cell hyperplasia was made and confirmed by immunophenotyping and molecular studies.
Conclusion: Hematopathologists must be aware of various confounding factors and should judiciously use flow cytometric and molecular studies for attaining a proper diagnosis of HCL. We also report a very rare immunophenotypic aberrancy (CD 19 negativity) in HCL. 



Introduction

Hairy cell leukemia (HCL) is an uncommon yet unique hematolymphoid neoplasm exhibiting a characteristic cytomorphology, immunophenotype, and well defined molecular features. It accounts for 2% of all lymphoid leukemias.[1] Typically, it presents with splenomegaly, pancytopenia, and monocytopenia; and shows a characteristic immunophenotype. The cells are universally CD19positive with co-expression of CD25, CD11c, CD103 and CD123. Patients of HCL can present with a spectrum of different clinical and pathological characteristics often puzzling a clinician or a pathologist. However, the clinical and pathological findings often complement each other to clinch the diagnosis of HCL. A correct diagnosis is mandatory as specific therapy in the form of purine analogues can provide long-term remissions in such patients.[2] We present a case of HCL with atypical clinical and laboratory features confounding the primary hematological abnormality.

Case Reports

Clinical history: A 69-year-old male with no known co-morbidities presented with complaints of breathlessness on exertion for 15 days. It was not associated with fever, cough, or purulent expectoration. He had moderate pallor, tachycardia, and tachypnoea. There was no lymphadenopathy or hepatosplenomegaly. Respiratory system evaluation revealed features suggestive of consolidation confirmed by chest roentgenogram. He improved with parenteral antibiotics. Meanwhile, he was detected to have pancytopenia [Hemoglobin (Hb) – 91g/L, total leukocyte count (TLC) – 2.3x109/L, and platelet count – 122 x109/L] with the nadir absolute neutrophil count (ANC) of 252/µL. He was subjected to bone marrow evaluation which revealed plasmacytosis (plasma cells – 10%).  A serum protein electrophoresis (SPEP) performed subsequently, showed M-spike (0.3g/dL), though no immunofixation studies were done. He was referred to our center for further evaluation of suspected plasma cell dyscrasia.
On the assessment at our center, he was asymptomatic. He did not complain of bone pains. Physical examination was not contributory. His pancytopenia recovered (Hb-112 g/L, TLC-3.2 x10
9/L, ANC-1.4 x109/L, platelets-522 x109/L). A repeat bone marrow examination was performed to evaluate suspected plasma cell dyscrasia. Bone marrow aspirate (BMA) revealed 5% plasma cells; however showed 36% larger lymphoid cells with clumped chromatin and a moderate amount of pale basophilic cytoplasm. A few cells had grooved/reniform nucleus or cytoplasmic projections (Figure 1A). An increase in mast cells was also noted. Trephine biopsy showed an interstitial infiltrate, typical of hairy cell leukemia (Figure 1B, C) along with an increase in mast cells confirmed by mast cell tryptase immunohistochemistry (Figure 1D). No significant clusters of mast cells were highlighted. Repeat SPEP and immunofixation study, done at our center, revealed polyclonal hypergammaglobulinemia.
Multiparametric flow cytometry (Figure 2) was performed on the BMA using four/six color antibody panels by lyse-wash-stain method (antibodies from BD Biosciences, San Jose, CA). One tube containing unstained leukocytes was used as negative control. A minimum of one lakh events was acquired on dual laser BD FACS Canto II and analyzed using BD FACS Diva software. Bright CD19positive low side scatter events (5.3% of viable gated leucocytes) were gated which were positive for CD10, CD20, CD22, CD79b, surface Igκ, CD25, CD11c, CD103, and CD123. Serendipitously, we found a large subclone of cells (15% of viable gated leucocytes) expressing exactly the same immunophenotype markers except for CD19, indicating its loss of expression from hairy cells. The fluorochrome related technical issues were ruled out as the cells showed similar profile using both anti-CD19PECy7 and anti-CD19APC-H7 (clone SJ25C1, BD Biosciences). The CD19 negative cells had an immune profile exactly similar to CD19+ve cells and revealed expression of hairy cell markers along with CD20, CD22, CD79b, CD45, and CD10. The plasma cells (CD38
pos/CD138pos/CD19pos/CD81pos/CD56neg and no light chain restriction – Figure 2) and mast cells (CD117pos/CD33pos/CD2neg/CD25neg) showed normal immunophenotype indicating reactive plasmacytosis and mast cell hyperplasia. A diagnosis of hairy cell leukemia with atypical features (CD19 negative subclone, CD10 positivity, reactive plasmacytosis and mast cell hyperplasia) was made which was subsequently confirmed by amplification-refractory mutation system polymerase-chain-reaction (ARMS-PCR) for BRAF V600E mutation (Figure 1E).
The patient remained asymptomatic, and his laboratory parameters remained normal; not warranting purine analogue therapy. He has been keeping under close medical observation.

Figure 1 Figure 1. (A) Larger lymphoid cells/hairy cells in bone marrow aspirate smear (May-Grunwald Giemsa stain, x1000); (B) Trephine biopsy showing interstitial infiltrate of hairy cell having a typical “fried egg” appearance (Hematoxylin and Eosin stain, x600); (C) Immunohistochemistry for DBA.44 highlights hairy cells (Hematoxylin counterstain, x600); (D) Immunohistochemistry for mast cell tryptase highlights mast cells (Hematoxylin counterstain, x600). (E) ARMS-PCR and agarose gel electrophoresis showing positivity for BRAF V600E mutation (lane 3); 100bp ladder (lane 1), positive control (lane 2), and negative control (lane 4). 

Figure 2 Figure 2. Multiparametric flow cytometry shows two distinct clones (CD19pos and CD19neg) of cells both of which are positive for CD45, CD22, CD10, CD25, CD103, CD11c, CD123 and surface Igκ in similar intensities. The plasma cells do not show clonal restriction.

Discussion

HCL is a unique B-cell non-Hodgkin lymphoma (NHL) characterized by splenomegaly, cytopenias affecting two or more lineages and morphologically by typical hairy cells. Though the majority of cases have this typical presentation, there are scenarios in which the clinical, morphological or immunophenotypic features are atypical, leading to diagnostic confusion. The case presented here exemplifies this intriguing situation where the patient on evaluation for a lower respiratory tract infection was found to have cytopenias, had no palpable splenomegaly and the bone marrow showed only a few "hairy cells" along with confounders in the form of plasmacytosis and mastocytosis. All these together with a minor quantity of  serum “M” protein led to initial misdiagnosis.
Splenomegaly is an important feature seen in up to 90% of patients with HCL. However, its absence should not exclude a diagnosis of HCL.[3] And more importantly, a changing trend has been observed in the symptomatology of HCL over the past 30 years. Number of cases are being diagnosed at an early stage with a less marked splenomegaly.[4]
A co-existence of plasma cell myeloma with HCL as well as the development of myeloma in patients with HCL has been reported in the literature.[5] At times, plasma cell myeloma/leukemia may mimic HCL also.[6-7] Clonal plasma cells were excluded by flow cytometry and SPEP studies. The initial report of small monoclonal band in SPEP from outside our institute might represent a transient monoclonal gammopathy, as has been reported previously with several infections.[8-9] However, a wrong interpretation could not be conclusively resolved in the absence of immunofixation studies. The association of mast cell hyperplasia with HCL has been well characterized by Macon et al.[10] This has been attributed to the angiogenesis and further progression of the disease, confirmed by a latter study.[11] There has also been a case report of systemic mastocytosis associated with a clonal hematopoietic non-mastcell lineage disease (SM-AHNMD) where the coexisting neoplasms were of both lymphoid and myeloid origin.[12] Our case shows a striking mast cell hyperplasia, however, a systemic mastocytosis has been ruled out based on immunophenotype studies.    
Immunophenotype aberrancies have been well described in HCL, like negativity for CD103 or CD25; and positivity for CD10 or CD23.[13] In our case, the cells showed positivity for CD10, and there was a subclone with absence of CD19 expression. While CD10 expression is relatively common (5-26% of cases) and explained by alternate origin of leukemic cells from germinal center,[13] the absence of CD19 expression in HCL has not been previously reported in the literature. CD19 plays an important role in B-cell growth and differentiation and its expression increases as a B-cell matures. This characteristic is often the basis of using it in flow cytometry as a gating marker for the diagnosis and for minimal residual disease (MRD) testing in various B-cell malignancies. In fact, of all the B-NHLs, HCL cases show the maximum level of expression of CD19.[14] The abnormal immunophenotypic pattern should be borne in mind while performing the MRD analysis during follow-up. An alternate marker (CD20) should also be considered for gating leukemic cells in these patients.[15]

Conclusions

We report a case of HCL with unique clinical, morphological and immunophenotypic features. A hematopathologist must be aware of these confounding factors and must deal such cases with a high index of suspicion and a supportive armamentarium of flow cytometry and molecular studies.    

Acknowledgement

The authors are thankful to Mrs. Jasbir Kaur Hira and Mrs Praveen Bose for the technical help in performing molecular and immunophenotypic studies respectively. 

References   

  1. Foucar K, Falini B, Catovsky D, Stein H. Hairy cell leukaemia. In: Swerdlow SH, Campo E, Harris NL, et al. eds. WHO classification of tumours of haematopoietic and lymphoid tissues. Lyon, France: IARC press; 2008:188-190.   
  2. Else M, Dearden CE, Catovsky D. Long-term follow-up after purine analogue therapy in hairy cell leukaemia. Best Pract Res Clin Haematol 2015;28:217-29. https://doi.org/10.1016/j.beha.2015.09.004 PMid:26614900     
  3. Johnston JB, Grever MR. Hairy Cell Leukemia. In: Greer JP, Arber DA, Glader B et al. eds. Wintrobe's Clinical Hematology. Philadelphia; Lippincott Williams & Wilkins; 2014:4395-4443.    
  4. Frassoldati A, Lamparelli T, Federico M, Annino L, Capnist G, Pagnucco G, Dini E, Resegotti L, Damasio EE, Silingardi V. Hairy cell leukemia: a clinical review based on 725 cases of the Italian Cooperative Group (ICGHCL). Italian Cooperative Group for Hairy Cell Leukemia. Leuk Lymphoma 1994;13:307-16. https://doi.org/10.3109/10428199409056295 PMid:7519510      
  5. Saif MW, Greenberg BR. Multiple myeloma and hairy cell leukemia: a rare association or coincidence? Leuk Lymphoma 2001;42:1043-8. https://doi.org/10.3109/10428190109097724 PMid:11697621     
  6. Hanbali A, Alrajeh A, Rasheed W. Plasma cell leukemia mimicking hairy cell leukemia. Hematol Oncol Stem Cell Ther 2015;8:91-2. https://doi.org/10.1016/j.hemonc.2015.05.001 PMid:26013472      
  7. Lesesve JF, Broseus J. Confusing Hairy Cells in a Case of IgG Kappa Plasma Cell Leukemia. Clin Lab 2016;62:749-50. https://doi.org/10.7754/Clin.Lab.2015.150834 PMid:27215099      
  8. Stoimenis D, Spyridonidou C, Papaioannou N. Transient Monoclonal Gammopathy Induced by Disseminated Staphylococcus aureus Infection. Case Rep Med 2012;2012:607104.    
  9. Seve P, Turner R, Stankovic K, Perard L, Broussolle C. Transient monoclonal gammopathy in a patient with Bartonella quintana endocarditis. Am J Hematol 2006;81:115-7. https://doi.org/10.1002/ajh.20499 PMid:16432867      
  10. Macon WR, Kinney MC, Glick AD, Collins RD. Marrow mast cell hyperplasia in hairy cell leukemia. Mod Pathol 1993;6:695-8. PMid:8302811      
  11. Ribatti D, Crivellato E, Molica S. Mast cells and angiogenesis in haematological malignancies. Leuk Res 2009;33:876-9. https://doi.org/10.1016/j.leukres.2009.02.028 PMid:19324412      
  12. Gülen T, Sander B, Nilsson G, Palmblad J, Sotlar K, Horny HP, Hägglund H. Systemic mastocytosis: progressive evolution of an occult disease into fatal mast cell leukemia: unique findings on an unusual hematological neoplasm. Med Oncol 2012;29:3540-6. https://doi.org/10.1007/s12032-012-0261-5 PMid:22661384      
  13. Chen YH, Tallman MS, Goolsby C, Peterson L. Immunophenotypic variations in hairy cell leukemia. Am J Clin Pathol 2006;125:251-9. https://doi.org/10.1309/PMQXVY619Q8Y43AR PMid:16393677      
  14. Ginaldi L, De Martinis M, Matutes E, Farahat N, Morilla R, Catovsky D. Levels of expression of CD19 and CD20 in chronic B cell leukaemias. J Clin Pathol 1998;51:364-9. https://doi.org/10.1136/jcp.51.5.364 PMid:9708202 PMCid:PMC500695    
  15. Sausville JE, Salloum RG, Sorbara L, Kingma DW, Raffeld M, Kreitman RJ, Imus PD, Venzon D, Stetler-Stevenson M. Minimal residual disease detection in hairy cell leukemia. Comparison of flow cytometric immunophenotyping with clonal analysis using consensus primer polymerase chain reaction for the heavy chain gene. Am J Clin Pathol 2003;119:213-7. https://doi.org/10.1309/G6299513NGLCUB1K PMid:12579991        



       

[TOP]

Abstract views:
319

Views:
PDF
61
HTML
179

Article Metrics

Metrics Loading ...

Metrics powered by PLOS ALM


Copyright (c) 2017 Mediterranean Journal of Hematology and Infectious Diseases

Creative Commons License
This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.

 

The Mediterranean Journal of Hematology and Infectious Diseases [eISSN 2035-3006] is owned by the U.C.S.C. and it is published by PAGEPress®, Pavia, Italy. All credits and honors to PKP for their OJS.
 
 
© PAGEPress 2008-2017     -     PAGEPress is a registered trademark property of PAGEPress srl, Italy.     -     VAT: IT02125780185