Introduction

Anaplastic lymphoma kinase (ALK)-positive diffuse large B cell lymphoma (ALK+/DLBCL) is a rare and distinct subtype of DLBCL, originally described in 1997 by Delsol et al. [1]. To date, approximately 50 cases of ALK+ LBCL have been described in the English literature, and those cases share similar morphologic, immunophenotypic, and molecular genetic characteristics.

ALK+/DLBCL occurs more frequently in male adults and spans all age groups. It shows a very aggressive behavior, high relapse rate, and lack of response to standard regimens [2, 3]. Although it primarily involves lymph nodes, extranodal presentations such as the tongue, nasopharynx, and stomach have been rarely reported [3–7].

Histologically, ALK+/DLBCL exhibits an immunoblastic and/or plasmablastic morphology, often an intrasinusoidal growth pattern, and is derived from B cells based on the expression of monotypic light chain. It shows a unique immunophenotypic profile characterized by strong staining for ALK protein and expression of plasmacytic markers but lack B, T-lineage markers and CD30 [2]. The plasmablastic appearance and CD20 negativity of ALK−DLBCL make this entity a diagnostic challenge with a broad histological differential diagnosis.

Molecular cytogenetics and/or reverse transcriptase polymerase chain reaction has shown that the most commonly observed cytogenetic abnormality of ALK-positive DLBCL is t(2;17)(p23;q23) involving clathrin (CLTC) on 17q23 and ALK on 2p23 [5, 8, 9]. However, cases of ALK-positive DLBCL with underlying t(2;5)(p23;q35) involving ALK on 2p23 and nucleophosmin on 5q35, as seen in the majority of T/null anaplastic large cell lymphoma, have also been described [4]. Other rare cytogenetic abnormalities have been reported including novel ALK fusion partners [10, 11].

In the present study, we describe a case of ALK+/DLBCL diagnosed in the bone marrow trephine, showing the typical morphological and immunohistochemical features of the entity and associated with ALK-1 gene rearrangement detected by fluorescence in situ hybridization (FISH) analysis.

Clinical history

The patient, a 24-year-old male, presented with anemia and B symptoms. Physical examination did not reveal peripheral lymphadenopathy. Laboratory investigation showed normochromic normocytic anemia. The patient was HIV negative. A CT scan of the thorax and abdomen showed multiple liver nodules without additional lymphadenopathy, and a diagnosis of malignancy was suspected. A staging bone marrow biopsy was performed, and the sample was fixed in formalin. Liver biopsy was not performed. Fresh tissue was not procured for conventional cytogenetic analysis.

Materials and methods

Sections were routinely processed in paraffin for light microscopy and immunohistochemistry. FISH analysis was performed using the ALK split probe from DAKO (Denmark) according to the manufacturer’s instructions. Fluorescence images were acquired using a Zeiss Axio Imager Z1 microscope (Carl Zeiss, Jena, Germany) equipped with a Plan Neofluar ×100/1.3 oil objective lens and the appropriate filter settings. Images were recorded using a monochrome progressive scan CV-M4/CL camera (Applied Imaging Corp., CA, USA). The complete setup was controlled by CytoVision software, 3.93.1 release (Applied Imaging Corp.).

Results

Histology showed diffuse infiltration of the marrow trephine by sheets of monomorphic large atypical lymphoid cells with oval to irregular nuclear contours, prominent nucleoli, and abundant eosinophilic or amphophilic cytoplasm (Fig. 1a). Some of the atypical lymphoid cells showed eccentrically located nuclei with perinuclear hof, reminiscent of plasmablasts. In addition, frequent mitoses were easily appreciated.

Fig. 1
figure 1

Large atypical lymphoid cells with irregular nuclei, prominent nucleoli, and abundant eosinophilic or amphophilic cytoplasm (H&E stain, original magnification ×40) (a). The neoplastic cells are CD45 positive (b) and CD138 positive (c) (immunohistochemistry, original magnification ×40). Immunohistochemical stains for kappa (d) and lambda (e) immunoglobulin light chains showing the lymphoma cells to be kappa light chain-restricted (immunohistochemistry, original magnification ×40)

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Immunohistochemically, the atypical lymphoid cells were strongly and diffusely positive for CD45, CD138, and IgA; weakly positive for MUM1; and partially positive for EMA. They showed positive kappa immunoglobulin light chain expression but negative lambda light chain expression (Fig. 1b–d). They exhibited positive ALK immunoreactivity with restricted cytoplasmic granular staining pattern (Fig. 2a) and a high proliferation fraction (approximately 60 %) by Ki-67 staining. The B cell markers PAX-5, CD20, and CD79a were negative. The lymphoid cells were CD30, CD10, Bcl-2, CD3, CD4, and TIA-1 negative. In situ hybridization for Epstein–Barr virus (EBER) was negative as well as immunohistochemistry for human herpes virus-8.

Fig. 2
figure 2

The neoplastic cells display cytoplasmic granular staining for ALK-1 (a) (immunohistochemistry, original magnification ×40). FISH for ALK split probe shows lymphoid cells with a fused signal, as well as isolated red and green signals (arrows) indicating a break in the ALK locus (b)

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On the basis of morphology and immunophenotype, a diagnosis of ALK+/DLBCL was assigned. Interphase FISH analysis showed that the atypical lymphoid cells displayed isolated red or green signals, indicating the presence of a breakpoint in the ALK locus (Fig. 2b). The patient received six cycles of CHOP. Follow-up imaging studies show no evidence of lymphoma. He is currently in complete continuous remission, 16 months from initial diagnosis.

Discussion

Anaplastic lymphoma kinase-positive large B cell lymphoma is a rare variant of diffuse large B cell lymphoma that occurs more frequently in male adults. It involves nodal and extranodal sites, and it is associated with a poor outcome [3–6, 12]. In our case, the patient presented with liver nodules; however, a liver biopsy was not performed and the diagnosis was made on a staging bone marrow trephine. Liver, as the primary site of ALK+/DLBCL, has been reported in only four cases in the English literature [13], whereas initial diagnosis in bone marrow trephine has not been reported previously. A summary of the key data from all cases with extranodal presentation published in the English literature is provided in Table 1.

Table 1 Cases of ALK+ diffuse large B cell lymphoma with extranodal presentation
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Histologically, ALK+/DLBCL shows immunoblastic/plasmablastic cytology and a sinusoidal growth pattern may be seen. Immunohistochemically, in addition to ALK-1 expression, it presents positivity for CD138, VS38c, MUM1, and monotypic cytoplasmic light chain; EMA is usually seen as well. B cell- and T cell-related antigens are negative. CD4, CD57, CD43, and CD45 are variably positive. CD30 and EBV are not expressed [2, 6–8, 12]. Rare tumors express focally CD20 or cytokeratin on scattered cells [6]. A CD30-positive case has been reported [14] as well as a case showing CD10 positivity [7]. Our case showed the typical “plasmacytic” immunohistochemical profile.

ALK+/DLBCL should be distinguished from T-anaplastic large-cell lymphoma (T-ALCL), plasmablastic lymphoma, plasmablastic myeloma, anaplastic variant of diffuse large B cell lymphoma, and carcinoma. The distinction is based on the overall morphologic and immunohistochemical features [2]. T-ALCL is typically CD30+ of T cell phenotype and would be negative for plasma cell markers and immunoglobulin light chain. Plasmablastic lymphomas often occur in the oral region of human immunodeficiency virus-infected individuals and are EBER positive and ALK negative. The anaplastic variant of DLBCL is usually strongly CD20+ and ALK−. Plasmablastic myeloma has not been reported to be ALK positive and would be associated with other features such as lytic bone lesions and a monoclonal protein in serum and/or urine.

ALK+/DLBCL is characterized by ALK rearrangement. In well-characterized cases, three genes have been reported as a fusion partner of ALK: clathrin (CLTC-ALK), nucleophosmin (NPM-ALK), and SEC31A (SEC31A-ALK) [4, 5, 8–10]. Recently, a novel fusion partner of ALK has been reported SEC31A and sequestosome1/SQSTM1 [11]. The ALK immunostaining pattern in ALK+/DLBCL appears to correlate with the type of the underlying rearrangement. Cases with CLTC-ALK/t(2;17) rearrangement show a distinctly cytoplasmic and granular ALK staining pattern, whereas those cases with an NPM-ALK/t(2;5) rearrangement show both cytoplasmic and nuclear staining [2, 6, 8]. The staining pattern for the novel partner sequestosome1/SQSTM1 has been reported as diffusely cytoplasmic [11]. However, this correlation may be imperfect, as Onciu et al. reported one case of ALK+/DLBCL with NPM-ALK fusion that showed cytoplasmic ALK staining only [4]. Because of the heterogeneity of ALK rearrangements in ALK+/DLBCL, in our case, we selected to use a split probe that confirms the ALK rearrangement; however, it does not allow the definition of the specific chromosome partner. Given the cytoplasmic granular staining pattern with ALK immunohistochemistry, it is most likely that the partner in our case is clathrin.

ALK+/DLBCL is a very aggressive neoplasm with high relapse rate and lack of response to standard regimens [2, 3]. The plasmablastic differentiation which is found in these neoplasms is associated with the acquisition of the transcriptional profile of plasma cells in concert with extinction of the B cell differentiation program by proliferating immunoblasts. The deeper insights into the molecular pathogenesis of large B cell lymphomas with plasmablastic differentiation could identify new and effective alternative treatments [15].

In conclusion, we report a rare case of ALK+/DLBCL diagnosed in bone marrow trephine of a young patient who presented with liver masses. The histopathologic diagnosis and differential diagnosis on bone marrow biopsy require detailed immunohistochemical analysis with inclusion of ALK antibody in the immunohistochemical panel for those cases showing anaplastic/plasmablastic morphology and fail to express the usual cellular lineage markers, particularly in the bone marrow biopsy. FISH analysis supports the diagnosis and confirms the ALK gene involvement in the pathogenesis of the disease.