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T-Cell-Rich B-Cell Lymphoma Presenting in the Spleen: A Clinicopathologic Analysis of 3 Cases

Posted on: Wednesday, 25 February 2004, 06:00 CST

We review the clinical, pathologic, and molecular genetic features of 3 splenic T-cellrich B-cell lymphomas and discuss their differential diagnosis. all patients presented with symptomatic splenomegaly and underwent diagnostic/therapeutic splenectomy. Microscopically, the spleen in all cases showed a micronodular proliferation of lymphoid cells. A proportion of the nodules demonstrated central hyalinization or sclerosis. There was also an exuberant extramedullary hematopoiesis. On immunohistochemical stain, the nodules consisted predominantly of small T cells with scattered large atypical B cells. The clonal nature of the atypical B cells was confirmed by polymerase chain reaction assays for immunoglobulin heavy-chain gene rearrangement. In the H&E sections, the differential diagnoses included Hodgkin's lymphoma, follicular lymphoma, peripheral T-cell lymphoma, and nonneoplastic granulomatous process. The presence of exuberant exlramedullary hematopoiesis also raised the possibility of a chronic myeloproliferative disorder. The combined morphologic, immunohistochemical, and molecular genetic data are essential for a correct diagnosis of splenic T-cell-rich B-cell lymphoma. Int J Surg Pathol 12(1)31, 2004

Key words: T-cell-rich B-cell lymphoma, spleen, splenomegaly, extramedullary hematopoiesis.

T-cell-rich B-cell lymphoma (TCRBCL) was originally described by Ramsay et al. [1] in 1988 as a B-cell malignancy with abundant reactive T lymphocytes. It is a rare entity representing about 1-2% of all non-Hodgkin's lymphomas. Histologically, this disease is characterized by the presence of a minor population of large neoplastic B cells in a sea of reactive T cells. A variant form of TCRBCL with numerous histiocytes has also been described by Delabie et al. [2], Because of its unusual morphologic appearance, immunohistochemistry as well as molecular studies are essential for the diagnosis of TCR-BCL. Clinically, TCRBCL and its histiocyte- rich variant frequently present as advanced disease with poor treatment outcome, perhaps with the exception of the primary cutaneous counterpart [3]. In the most recent World Health Organization classification of hematolymphoid malignancies, TCRBCL is considered as a morphologic variant of diffuse large B-cell lymphoma [4].

TCRBCL occurs most frequently as a primary nodal disease. Extranodal presentation has also been reported to occur in skin, lung, and other anatomic sites. Because of its rarity and unusual histomorphology, a diagnosis of TCRBCL arising from an extranodal site can sometimes be challenging. In this report, we describe the clinical, pathologic, and molecular genetic features of 3 splenic TCRECL cases to illustrate the diagnostic difficulty both clinically and pathologically.

case Reports

The pathology databases from Johns Hopkins (1995-2000) and Emory University Hospitals (1999-2002) were searched for spleen with malignant lymphoma. Three cases (1 from Hopkins and 2 from Emory) met the diagnostic criteria of TCRBCL (90% or more of lhc infiltrating lymphocytes were T cells). These were retrieved and the clinical data are summarized in Table 1.

case 1

The patient was an 80-year-old woman who initially presented with hemolytic anemia and leukopenia without fever or night sweats. On physical examination, the patient was found to have significant splenomegaly that was further confirmed by ultrasound. A bone marrow biopsy showed scattered small lymphoid aggregates consistent with a benign process. The patient underwent splenectomy, and a diagnosis of TCRBCL was made in December 2001. Extensive staging was negative for involvement of other anatomic sites. She was managed conservatively without chemotherapy and was alive and well 9 months after initial diagnosis.

case 2

A 73-year-old white woman presented to an outside hospital with anemia and marked splenomegaly. A bone marrow biopsy was reported as hypercellular with many noncaseating granulomas. Two months later, the patient underwent splenectomy because of worsening anemia and abdominal discomfort sec ondary to progressive splenomegaly. The diagnosis of the splenectomy specimen again was reported as multiple epithelioid granulomas suggestive of sarcoidosis. The patient was treated briefly with corticosteroids. The hone marrow biopsy and splenectomy slides were sent to Emory for second opinion, and upon reevaluation, a revised diagnosis of TCR-BCL involving both bone marrow and spleen was rendered. The patient was alive with disease 3 months following splenectomy. No further clinical follow-up information was available.

Table 1. Clinical Data

case3

The patient was a 46-year-old woman who initially presented in june 1998 with fever, soaking night sweats, and fatigue. Laboratory works revealed anemia with abnormal liver function test results. Computerized tomography scans showed marked enlargement of the spleen with no significant visceral adenopathy. Splenectomy was performed in August 1998, and a diagnosis of TCRBCL was made. A liver biopsy also showed involvement by TCRBCL. The patient received 6 cycles of cyclophosphamide, doxorubicin, vincristine, and prednisolone (CHOP) chemotherapy and total body irradiation, followed by allogeneic bone marrow transplantation in January 1999. Her posttransplantation course was complicated by graft-vs-host disease, liver failure, and thmmbotic thrombocytopenic purpura. She expired 1 month after bone marrow transplant.

Materials and Methods

lmmunohistochemistry

Immunohistochemical stains were performed on 5-um sections of formalin-fixed, paraffin-embedded tissues with use of the standard avidin-biotin-complex technique [5]. The antibodies used included CD3, CDlO, CD20 (L26), CD15 (LeuMl), CD30 (Ki1), CD57 (Leu-7), Bcl- 2, CD45 (LCA), and kappa and lambda (DAKO Corporation, Carpinteria, CA).

How Cytometric lmmunophenotyping (FCI)

FCI was performed in cases 1 and 3 using the standard lymphoma protocol as described previously [6]. Briefly, single-cell suspension was prepared from fresh splenic tissue and stained with various combinations of 4 different fluorochrome -conj u gated antibodies. Approximately 10,000 cells were acquired with use of a FACS Calibur flow cytometer and analyzed by use of Paint-a-Gate or CellQuest computer software programs.

Polymerase Chain Reaction (PCR) Assay

Immunoglobulin heavy-chain and T-cell receptor gamma gene rearrangement studies were carried out using PCR assays as described |7-9J. Briefly, genomic DNAs were extracted from formalin-fixed, paraffin-embedded tissue blocks of the splenectomy specimens. Two different consensus primers corresponding to the variable framework region (PR) 3 sequence of the immunoglobulin heavy-chain gene were synthesized (5'-ACACGGCCTGTGATATTACTGT-3' and 5'- CTGTCGACACGGCCGTGTATTACTG-3'). Each of these upstream primers was paired with a common downstream consensus primer of the joining region of the immunoglobulin heavy-chain gene (5'- ACCTGAGGAGACGGTGACC-3') in a PCR assay performed in duplicate for each set of primers. The PCR products were resolved on 8% polyacrylamide gel, stained with ethidium bromide, and visualized under ultraviolet light. The first FR3 primer is further upstream of the second FR3 primer. Therefore, the expected PCR product amplified with the first set of primers is slightly larger than the one generated from the second set. A different set of primers was used for T-cell receptor gamma gene rearrangement assay (forward primer: 5'-TACATCCACTGGTACCTACACCA-3' and reverse primer: 5'- CCCGTCGACTACCTTGGAAATGTTGTATTCTTC-3'), and the PCR products were analyzed by heteroduplex analysis as described. Genomic DNA extracted from normal tonsil tissue was used as negative control. Genomic DNA isolated from SUP-B15 and jurkat cell lines at a dilution of 1:1OO with tonsil DNA was used as positive controls for immunoglobulin heavy-chain and T-cell receptor gamma genes, respectively.

Results

All 3 patients were female, and their age ranged from 46 to 80 years with a mean age of about 66 years. all patients presented with clinical and radiographie evidence of splenomegaly. None of the patients had a prior history of malignant lymphoma, and all patients underwent diagnostic/therapeutic splenonectomy for the presumed clinical diagnosis of functional hypersplenism causing anemia.

The resected spleens were enlarged in all 3 cases, and their weight ranged from 780 to 1,110 grams with an average of about 915 grams. No large tumor masses were noted grossly. Histologie sections of the spleen in all cases demonstrated a micronodular proliferation of lymphoid cells in the white pulp (Fig. IA). The nodules were composed predominantly of small lymphocytes with inconspicuous other types of inflammatory cells such as eosinophils or plasma cells. Within this background were rare, sparsely distributed, large atypical lymphoid cells (Pigs. IB and C) and histiocytes. These atypical cells did not have the characteristic morphologic features of classic Reed-Sternberg cells or lymphohistiocytic-type cells. They were more closely related to large centroblasts or immunoblasts morphologically. A significant proportion of the tumor nodules showed central sclerosis or hyalinization and frequently abutted or surrounded small arterioles (Fig. IB). There was no necrosis. The redpulp was not involved by lymphoma but displayed marked cxtramcdullary hematopoiesis (Fig. IB).

Flow cytometric immunophcnotyping tailed to demonstrate a clonal population of B cells in the 2 cases studied. The lymphocytes were composed almost exclusively of T cells in both cases and they were phenotypically unremarkable (data not shown). On immunohistochemical stains, the small lymphocytes of the tumor nodules were positive for the pan-T cell marker CD3 while the large atypical lymphoid cells were positive for the B-cell marker CD20 (Figs. 2A-D). Focal positivity lor CD30 was noted on the large atypical cells in 1 case, but no CD 15 expression was observed in any of the cases examined. There was no increase in CD57-positive T cells. CD45 staining was difficult to interpret owing to the surrounding T cells bul appeared to be positive in all cases. To further confirm the clonal nature of the large atypical B cells and to rule out a clonal T-cell process, PCR assays were performed. Monoclonality of the atypical B cells was demonstrated in all cases, and a representative one is shown in Fig. 3. The T cells were polyclonal in all 3 cases (data not shown).

One patient received chemotherapy and subsequently allogeneic bone marrow transplantation. One patient received corticosteroid treatment for the presumed diagnosis of sarcoidosis at an outside hospital, and no further treatment information was available after the diagnosis of TCRBCL was made 3 months later. The last patient was managed conservalivcly with spleneclomy only. The patient treated with CHOP and allogeneic bone marrow transplantation died 6 months after initial diagnosis, and the other 2 patients were alive and well following initial diagnosis (3 to 9 months, respectively).

Fig. 1. Representative photomicrographs of the spleen showing a micronodular proliferation of lymphoid cells. (A). A subset of the nodules shows central hyalinization or sclerosis and the red pulp shows exuberant extramedullary hematopoiesis of all 3 lineages. (B). The nodules are composed mostly of small lymphocytes with scattered large atypical lymphoid cells as indicated by the arrow (C).

Discussion

In this report, we described the clinical, pathologic, and genotypic features of splenic TCRBCL in 3 patients. Il appears that splenic TCRBCL is a rare occurrence since only 3 cases were identified from 2 large tertiary healthcare institutions, a finding similar to what has been reported by Kraus et al. [10]. During the 10-year period at their large tertiary care institutions, they encountered only 2 cases of TCR-BCL in their splenectomy specimens. No TCRBCL cases were described in 2 other large study series [11,12]. Clinically, all patients in our report presented with anemia and symptomatic splenomegaly, and underwent diagnostic/ therapeutic splenectomy for the presumed clinical diagnosis of functional hypersplenism.

Fig. 2. The small lymphocytes are CD3-posilive T cells (A and B) while the large atypical lymphoid cells are positive for CD20 (C and D).

To the best of our knowledge, only rare cases of splenic TCRBCL have been reported in the literature [2,13-15]. Delabie et al. [2] and Achten et al. [15] reported a total of 4 cases of TCRBCL in the spleen. Most of these cases probably represented secondary splenic involvement by a nodal lymphoma based on the clinical stage information provided. Histologically, all cases displayed a multifocal nodular pattern of involvement in the white pulp, very similar to the ones described here. Betman and colleagues [14] reported a single case of TCRBCL in Uic spleen, but the pattern of involvement was dif1'usc rather than nodular and little clinical information was provided. In the 5 cases reported by these authors, only 1 case showed focal extramedullary hematopoiesis. This is clearly distinct from what was observed in our cases in which all demonstrated significant extramedullary hematopoiesis in the red pulp. Neither histomorphologic description nor clinical information was given in the cases reported by Krause et al. [10] and Chittal et al. [13].

Fig. 3. A representative photomicrograph showing the donally rearranged immunoglobulin heavy-chain gene in case 1 by polymerase chain reaction. Lane M, [Phi]174 molecular weight marker; Lane N, negative tonsil DNA control in duplicate; Lane P, positive SUP-B 1 5 cell line DNA control; Lanes A and B, patient 1 DNA with first (A) and second (B) primer set in duplicates (see Materials and Methods for details).

In the hematoxylin-and-cosin-stained sections, a nodular proliferation of lymphoid cells with or without histiocytes in the spleen raises many diagnostic possibilities, and ancillary studies such as immunohistochcmistry arc essential for a correct diagnosis of TCRBCL. In classical Hodgkin's lymphoma, Reed-Sternbcrg cells or their variants are easily identified. These cells are difficult to find in the TCRBCL cases described here. In addition, the background inflammatory cells such as eosinophils and plasma cells seen in classical Hodgkin's lymphoma are absent in TCRBCL. On immunohistochemical stains, the neoplastic cells in TCRBCL are positive for CD45 and CD20, a phenotype clearly different from typical classical Hodgkin's lymphoma cells. Distinguishing TCRBCL from lymphocyte-predominant Hodgkin's lymphoma may be a diagnostic challenge since the neoplastic cells share similar immunophenotype. The background cells in the latter category, however, arc composed of a mixture of small T and B lymphocytes while small T lymphocytes with a cytotoxic phenotype predominate in the background infiltrates in the former entity [16]. Furthermore, in lymphocyte-predominant Hodgkin's lymphoma, CD57-positive T cells account for a significant portion of the infiltrating T cells and frequently form collarettes around the tumor cells [17]. Immunophenotypic analysis essentially rules out follicular lymphoma and reactive processes. It also eliminates the diagnostic possibility of most peripheral T-cell lymphomas because of the pattern of involvement.

Even with the help of immunohistochemical stains, TCRBCL in the spleen, particularly those with increased histiocytes, can still be mistaken for a reactive granulomatous process as exemplified by case 2 of this report or Hodgkin's lymphoma as reported by Chittal et al. [13]. It should also be distinguished from the so-called "follicular T-ccll lymphoma" recently described by de Levai et al. [18]. In these situations, it is imperative to perform molecular studies to look for evidence of clonal rearrangement of the immunoglobulin heavy-chain or T-cell receptor genes. As expected, the lymphocytes in a reactive granulomatous process should be polyclonal. The neoplastic cells of "follicular T-cell lymphoma" should demonstrate clonal rearrangement of the T-cell receptor gene. In all 3 cases described here, the atypical B cells show clonal rearrangement of the immunoglobulin heavy-chain gene while the background T cells display a polyclonal pattern of their rearranged T-cell receptor gamma genes, confirming the morphologic and immunohislochemical diagnosis of TCRBCL.

The red pulp of the spleen in these cases was spared from involvement by TCRBCL. However, the exuberant extramedullary hematopoiesis present could potentially be misinterpreted as a manifestation of chronic myeloproliferative disorder, especially when the clinical information is not available. The underlying molecular mechanism for this extensive extramedullary hematopoiesis is not clear. Previous studies have demonstrated that certain cylokines, particularly IL-4, are expressed at increased level in the neoplastic B cells of TCRBCL, and local cytokine release may result in increased recruitment of the background cytotoxic T lymphocytes seen in TCRBCL [19]. Since many cytokines have been implicated in the process of extramedullary hematopoiesis [20,21], it is possible that the same cylokines released by the TCRBCL cells and/or additional cytokines released by the activated cytotoxic T lymphocytes are responsible for the extensive extramedullary hematopoiesis seen in these cases.

TCRBCL presenting in the spleen as secondary involvement by a systemic lymphoma should be treated as diffuse large B-cell lymphoma. However, if the lesion is localized in the spleen with a negative staging (case 1 in this report), a simple splenectomy in conjunction with anti-CD20 might be sufficient to alleviate symptoms and/or to reduce tumor burden, particularly in elderly patients. Since the number of reported cases is limited and there is no published experience with treatment of primary splenic TCRBCL, further study is needed.

In summary, TCRBCL initially presenting in the spleen is a rare occurrence. It should be distinguished from a variety of other disease entities by using an integrated approach including morphology, immunohistochemistry, and molecular methods. Appropriate staging should be performed before the initiation of systemic chemotherapy and/or bone marrow transplantation.

References

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Shiyong Li, MD, PhD, Karen P. Mann, MD, PhD, and Jeannine T. Holden, MD

From the Department of Pathology and Laboratory Medicine, Emory University School of Medicine, Atlanta, GA.

Reprint requests: Shiyong Li, MD, PhD, Department of Pathology and Laboratory Medicine, Emory University Hospital, F143D, 1364 Clifton Road NE, Atlanta, GA 30322.

Copyright Westminster Publications, Inc. Jan 2004

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