Cytometry B Clin Cytom. 2003 Jan;51(1):41-4.

Guess what: Chronic 13q14.3+/CD5-/CD23+ lymphocytic leukemia in blood and t(11;14)(q13;q32)+/CD5+/CD23- mantle cell lymphoma in lymph nodes!
Lima M, Pinto L, Dos Anjos Teixeira M, Canelhas A, Mota A, Cabeda JM, Silva C, Queirós ML, Fonseca S, Santos AH, Brochado P, Justiça B.

Service of Clinical Hematology, Hospital Geral de Santo António, Porto, Portugal. mmc.lima@clix.pt

Abstract
We report a case of a patient with two B-cell lymphoproliferative disorders: CD5(-)/CD23(+) B-cell chronic lymphocytic leukemia and CD5(+)/CD23(-) mantle cell lymphoma. These disorders were diagnosed simultaneously based on flow cytometry, immunohistochemistry, fluorescence in situ hybridization, and polymerase chain reaction-based molecular studies. The B-cell lymphocytic leukemia clone predominated in the blood and bone marrow, whereas the mantle cell clone predominated in lymph nodes.

Copyright 2002 Wiley-Liss, Inc.
PMID: 12500296 [PubMed - indexed for MEDLINE

BROCHADO, P.

CABEDA, J.M.

CANELHAS, A.

DOS ANJOS TEIXEIRA, M.

FONSECA, S.

JUSTICA, B.

LIMA, M.

MOTA, A.

PINTO, L.

QUEIROS, M.L.

SANTOS, A.H.

SILVA, C.

Cytometry Part B Clinical Cytometry

Repositório Científico do Centro Hospitalar do Porto

Case Report:Guess What:Chronic 13q14.3/CD5/CD23 Lymphocytic Leukemiain Blood and t(11;14)(q13;q32)/CD5/CD23 MantleCell Lymphoma in Lymph Nodes!Margarida Lima,1* Luı´sa Pinto,2 Maria Dos Anjos Teixeira,1 A´urea Canelhas,3 Alexandra Mota,1Jose´ Manuel Cabeda,4 Cida´lia Silva,1 Maria Luı´s Queiro´s,1 So´nia Fonseca,1 Ana Helena Santos,1Paulo Brochado,3 and Benvindo Justic¸a11Service of Clinical Hematology, Hospital Geral de Santo Anto´nio, Porto, Portugal2Service of Medicine, Hospital S. Gonc¸alo de Amarante, Amarante, Portugal3Service of Pathology, Hospital Geral de Santo Anto´nio, Porto, Portugal4Unit of Molecular Biology, Hospital Geral de Santo Anto´nio, Porto, PortugalWe report a case of a patient with two B-cell lymphoproliferative disorders: CD5/CD23 B-cell chroniclymphocytic leukemia and CD5/CD23 mantle cell lymphoma. These disorders were diagnosed simulta-neously based on flow cytometry, immunohistochemistry, fluorescence in situ hybridization, and polymerasechain reaction–based molecular studies. The B-cell lymphocytic leukemia clone predominated in the bloodand bone marrow, whereas the mantle cell clone predominated in lymph nodes. Cytometry Part B (Clin.Cytometry) 51B:41–44, 2003. © 2002 Wiley-Liss, Inc.Key terms: leukemia; lymphoma; flow cytometry; fluorescence in situ hybridization; B cells; biclonal; mantlecell; chronic lymphocytic leukemia; t(11;14); 13qOver recent years, the recognition that lymphoprolif-erative disorders (LPDs) comprise distinct clinicopatho-logic entities defined by a combination of morphologic,immunophenotypic, and genetic features and the avail-ability of specific therapies for certain of these tumorsmake accurate diagnosis imperative. In most cases, de-tailed immunophenotypic analysis of the leukemia/lym-phoma cells contributes to a specific classification. In fact,the pattern of reactivity for CD20, CD23, CD79b, FMC7,surface immunoglobulins (Igs), and adhesion moleculesproved to be of great help in distinguish between smalllymphocytic lymphoma/chronic lymphocytic leukemia(CLL) and mantle cell lymphoma (MCL) (1–5). Moreover,techniques based on polymerase chain reaction (PCR) andfluorescence in situ hybridization (FISH) to detect specificmolecular and chromosomal abnormalities (6,7) and theuse of antibodies against novel fusion proteins (8,9)greatly improved the ability to detect specific geneticabnormalities. For instance, the chromosomal transloca-tion (11;14)(q13;q32) that fuses the IGH and CCND1genes and leads to cyclin D1 overexpression has beenidentified as the genetic hallmark of MCL (10). Althoughan equivalent genetic marker has not been found forB-CLL, numerical or structural chromosomal abnormalitiesare detected by conventional cytogenetics or FISH analysisin 50% of B-CLL cases, the most common changes beingthe trisomy 12 and the structural abnormalities of 13q,11q, 6q, 14q, and 17p (11,12). Karyotypic analysis andFISH- and PCR-based molecular studies may be helpful inclarifying the diagnosis especially when the B-cell LPD isdifficult to classify based on clinical, morphologic, andphenotypic grounds such as in establishing the differentialdiagnosis between an MCL and a morphologically atypicalB-CLL (13–15) and in identifying B-CLL forms with specificchromosomal abnormalities with prognostic significance(11,16)We report a case of a patient in whom a CD5/CD23B-CLL and CD5/CD23 MCL were diagnosed simulta-neously based on flow cytometry and FISH- and PCR-basedmolecular studies. Aside from the rarity of a true CD5/CD23 B-CLL, to the best of our knowledge, this is the*Correspondence to: Margarida Lima, Unidade de Citometria, Servic¸ode Hematologia, Hospital Geral de Santo Anto´nio, Rua D Manuel II, s/n,4099-001 Porto, PortugalE-mail: mmc.lima@clix.ptReceived 8 May 2002; Accepted 29 August 2002Published online in Wiley InterScience (www.interscience.wiley.com).DOI: 10.1002/cyto.b.10005Cytometry Part B (Clinical Cytometry) 51B:41–44 (2003)© 2002 Wiley-Liss, Inc.first report documenting the simultaneous diagnosis of aB-CLL and an MCL.CASE REPORTAn 86-year-old man was evaluated at the HematologyDepartment of the Hospital Geral de Santo Anto´nio (Porto,Portugal) for persistent, absolute lymphocytosis and ad-renomegalies. White blood cell count was 16.4 109/liter(neutrophils, 22%; lymphocytes, 74%), hemoglobin was9.1 g/dl, and platelet count was 201  109/liter. Thephysical examination revealed multiple cervical, axillary,and inguinal adrenomegalies and hepatomegaly.The peripheral blood (PB) smear showed a markedpredominance of small lymphocytes with clumped chro-matin and scarce cytoplasm and a small population(15%) of large lymphoid cells with less condensed chro-matin, frequently with a visible nucleoli and a more abun-dant, sometimes basophilic, cytoplasm. Gumprecht’sshadows also were present. Flow cytometry–based immu-nophenotyping of PB lymphocytes revealed two popula-tions of phenotypically abnormal B cells (Fig. 1A). Thefirst one, comprising 90% of B cells, consisted of CD19,CD22low, CD23high, CD25low, FMC7, CD5, CD10,CD11a, CD11c, CD20, CD38, CD79b, CD103, andsurface and cytoplasmic Ig. The second population, com-prising only 10% of B cells, consisted of CD19, CD20high,CD22high, CD25low, CD38low, CD79b, FMC7, CD5,CD10, CD11a, CD11c, CD23, CD103, and surfaceIghigh (IgM, ). A few (1%) residual polyclonal B cellswere also present. FISH analysis showed the (11;14) trans-location in only 9% of PB lymphocytes (Fig. 2A) and the13q14.3 deletion in another 21% (Fig. 2B); trisomy 12 andthe (14;18) translocation were not detected (data notshown).On bone marrow smears, lymphocytes comprised 60%of the total cells counted, and immunophenotyping stud-ies revealed that most (59%) were B cells consisting ofCD19/CD5/CD23/surface Ig (73%) and CD19/CD5/CD23/ chains (27%), mature B lymphocytes,and a few residual CD19/CD20/CD38high/CD10/sur-face Ig B-cell precursors (1%). The bone marrow tre-phine biopsy showed a mixed nodular and interstitialinfiltration pattern consisting mainly of CD20/CD5/CD23 small lymphocytes.Flow cytometry of lymph node (LN) lymphocytesshowed that most (90%) had a CD5/CD23/ immuno-phenotype, whereas CD5/CD23high/Ig B cells com-prised only 10% of LN lymphocytes (Fig. 1B). FISH analysisshowed the t(11;14)(q13;q32) in 94% of LN cells (Fig. 2C).The LN histology was consistent with MCL, and immuno-histochemical studies confirmed that most B-lymphomacells expressed CD20 and CD5 and were negative forCD23; a few CD23 B lymphocytes also were observed.PCR-based IgH rearrangement molecular studies con-firmed that the B-cell clone found in the LN was differentfrom the one that predominated in blood (Fig. 3).DISCUSSIONAn increasing number of cases of biclonal B-cell LPDhave been reported in the past few years, likely a conse-quence of a more accurate laboratorian approach to diag-nosis (17–20); in most of these cases, two different cellpopulations were detected by flow cytometry, and, unlessthey express different Ig light chains, additional studiesare usually necessary to distinguish a true biclonal diseasefrom phenotypic heterogeneity due to intraclonal diver-sity. Only a few cases in which two distinct B-cell popu-FIG. 1. Dot plots showing the main immunophenotyping characteristics of peripheral blood (A) and lymph node (B) B cells. CD19/CD5 Blymphocytes are represented as blue dots and CD19/CD5 B lymphocytes are represented as red dots. Gray dots represent CD19 white blood cells.Flow cytometric studies were performed by using a method of staining and washing lyses (FACS lysing solution, Becton Dickinson, San Jose, CA) andfour-color staining that combined allophycocyanin-conjugated anti-CD19 and phycoerythrin–cyanine 5 conjugated anti-CD5 with a large panel of otherphycoerythrin and fluorescein isothiocyanate conjugated monoclonal antibodies directed against other markers combined as follows: FMC7/CD23,CD20/CD22, CD10/CD38, CD103/CD25, CD21/CD79b, CD11a/HLA-DR, / chains. Other specific combinations were used whenever necessary tobetter discriminate the B-cell populations. Data acquisition was carried out in a FACScalibur flow cytometer equipped with a 488-nm argon laser anda 635-nm red-diode laser (Becton Dickinson). For data analysis, Paint-a-Gate PRO software (Becton Dickinson) was used. FSC, forward scatter; SSC, sidescatter.42 LIMA ET AL.lations have been identified fulfill the criteria for diagnosisof two different entities (21).The rare biclonal B-cell LPD described here requiresfurther discussion concerning two points: the criteria fordiagnosis of a CD5 B-CLL and its association with a CD5MCL.Although expression of the CD5 antigen by neoplasticcells is considered an important criterion for diagnosis ofB-CLL, published series have frequently included a num-ber of “CD5 B-CLL” cases. A careful review of cases thathad been classified as “CD5 B-CLL” showed that mosthad other unusual immunophenotypic features, includingbright-surface Ig expression, bright CD20 expression, andabsence of CD23 expression, and displayed an adhesionmolecule profile resembling Non Hodgkin Lymphoma(NHL), suggesting that the majority of them representedNHL in the leukemic phase rather than CD5 B-CLL(22,23). Nonetheless, it is usually accepted that CD5B-CLL, although rare, does exist. We are convinced thatthe present case is one of the very rare cases of CD5B-CLL because it met all the immunophenotypic criteriafor the diagnosis of B-CLL except the expression of CD5and the arbor 13q14.3 deletion, a structural abnormalitycommonly observed in B-CLL; moreover, the lymphocytemorphology was that usually observed in typical CD5B-CLL and the possibility of a CD10 follicular cell lym-phoma was ruled out by immunophenotypic and FISHstudies.Concerning the characterization of the CD5 B-cellpopulation that accounted for only a minority of PB lym-phocytes and predominated in LN, its characteristicCD5, CD23, FMC7, CD20high, CD22high, surfaceIghigh immunophenotype strongly pointed to the diagno-sis of an MCL and made improbable the diagnosis of aB-CLL, a CD5 B-cell LPD that typically displays a CD23,FMC7, CD20/low, CD22/low, and surface Ig/lowFIG. 2. Fluorescence in situ hybridization (FISH) images showing thet(11;14) translocation (A) and the 3q14.3 deletion (B) that were found in9% and 24% of blood cells, respectively. C shows the t(11;14) translo-cation in most (94%) lymph node cells. The t(11;14) translocation isdetected by the presence of a yellow FISH spot in the cells at the leftparts of A and C, in addition to the normal red and green FISH spotsobserved in other cells from the same sample (right parts of A and C). The3q14.3 deletion is demonstrated by the presence of one red FISH spotin the cell at the left part of B, in opposition to the two normal red FISHspots in other cells from the same sample (right part of B). Fluorescencehybridization studies were performed with the following DNA probes:IGH-specific BAC (covering the JH and first constant regions) andCCND1-specific probes to detect the IGH-CCND1 fusion gene associatedwith the t(11;14)(11q13;14q32) translocation (LSI IGH/CCND1); aD13S319 locus-specific probe to detect del(13q14.3) structural abnor-malities (LSI D13S319).FIG. 3. Molecular identification of immunoglobulin H rearrangementclonality in samples of peripheral blood (lane 3) and lymph node (lane 4).Lanes 1 and 2 show molecular weight markers (500-bp ladder in lane 1and 100-bp ladder in lane 2). The lymph node was found to harbor aB-cell clone with a molecular weight different from that predominating inthe peripheral blood. Molecular studies were performed with the B-Lymphoma Molecular Diagnostic Kit from MD-Master Diagnostica(Granada, Spain) as recommended by the manufacturer. Peripheralblood DNA was extracted by using a semi-automated procedure with theNuclisens Extractor (Organon Technika Corporation, Durham, NorthCarolina); lymph node DNA was extracted with the Extracell Kit (Ampli-medica spa-Bioline, Torino, Italy).43B-CLL AND MCLphenotype (4,24–26). FISH studies confirmed our suspi-cion by revealing the presence of the typical chromo-somal abnormality t(11;14)(q13;q32).To the best of our knowledge, this is the first case inwhich a rare form of 13q14.3/CD5 B-CLL was associ-ated with a typical t(11;14)(q13;q32)/CD5 MCL, witheach LPD predominating in different compartments(blood/bone marrow and lymph nodes).LITERATURE CITED1. 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;51:364–369.2. Cabezudo E, Carrara P, Morilla R, Matutes E. Quantitative analysis ofCD79b, CD5 and CD19 in mature B-cell lymphoproliferative disor-ders. 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Guess what: Chronic 13q14.3+/CD5‐ /CD23+ lymphocytic leukemia in blood and t(11;14)(q13;q32)+/CD5+/CD23‐ mantle cell lymphoma in lymph nodes!

http://repositorio.chporto.pt/bitstream/10400.16/573/1/pdf%201.16.pdf

Guess what: Chronic 13q14.3+/CD5‐ /CD23+ lymphocytic leukemia in blood and t(11;14)(q13;q32)+/CD5+/CD23‐ mantle cell lymphoma in lymph nodes!

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