Diagnostic challenges posed by the wide morphological and immunophenotypic spectrum of plasma cell neoplasms

Abstract

Plasma cell neoplasms (PCNs) are a group of malignant neoplasms originating from the monoclonal expansion of immunoglobulin-secreting, terminally differentiated B cells, i.e., plasma cells (PCs). PCNs include monoclonal gammopathy of undetermined significance (MGUS), plasmacytoma (solitary plasmacytoma of the bone and extramedullary plasmacytoma), multiple myeloma (MM)/plasma cell myeloma, and rare PCNs associated with paraneoplastic syndromes. MM is a multifocal, bone marrow-based clonal PCN associated with end-organ damage and a monoclonal paraprotein (M protein) in the serum and/or urine. Morphologically, MMs are typically composed of mature to atypical PCs with a typical immunophenotype (CD38 + bright, CD138 + , CD19-, and monotypic light-chain expression), which are usually diagnostically straightforward. However, rare cases with morphological variants or immunophenotypic aberrancies may pose diagnostic challenges. Small, mature-appearing PC variants of MM at times may be difficult to differentiate from mature B-cell lymphomas with plasmacytic differentiation, such as lymphoplasmacytic lymphoma and marginal zone lymphoma, particularly in core needle biopsies. CD20 + and cyclin D1 + MM may mimic mantle cell lymphoma. Plasmablastic and pleomorphic/anaplastic variants are prone to present as extramedullary lesions mimicking plasmablastic lymphoma (PBL) or various types of lymphomas with pleomorphic/anaplastic morphology. Moreover, neoplastic PCs may exhibit aberrant expression of CD3, CD4, CD117, and cyclin D1, leading to potential diagnostic pitfalls. Diffuse CD30 expression with anaplastic morphology may be mistaken for anaplastic large-cell lymphoma. In rare instances, the neoplastic cells of PCNs are positive for Epstein-Barr virus (EBV), more frequently in cases with plasmablastic morphology and extramedullary involvement, which may raise suspicion for PBL. In this review, we emphasize the rare morphological and immunophenotypic features and variants of PCNs and utilize a few examples to highlight the diagnostic challenges.

Introduction

Plasma cells (PCs) are terminally differentiated B-cells that secrete immunoglobulins. Normal PC exhibits a single eccentrically placed, round nucleus with a "cartwheel" (or clock-face) appearance. The ample amount of cytoplasm is amphophilic with a perinuclear clear area (or perinuclear hof). Occasionally, benign/reactive PCs might be bi-nucleated. Plasma cell neoplasms (PCNs) are a group of malignancies arising from the monoclonal proliferation of PCs. These cells almost always secrete a monoclonal immunoglobulin known as M protein (paraprotein). The main exception is the rare non-secretory MM subtype. PCNs include monoclonal gammopathy of undetermined significance (MGUS), plasmacytoma, (either solitary plasmacytoma of the bone or extramedullary plasmacytoma), and multiple myeloma (MM)/plasma cell myeloma (PCM). When plasma cells (PCs) exhibit significant nuclear atypia, such as irregular nuclear contours, prominent nucleoli, loss of classic clock-face chromatin pattern, multinucleation, and nuclear inclusions (Dutcher body), a neoplastic process should be considered.

According to the 5th edition of the WHO Classification of Haematolymphoid Neoplasms (WHO-HAEM5), PCM is a bone marrow-based, multifocal neoplastic proliferation of PCs. MM is defined by the presence of PCM together with either evidence of disease-related organ damage or, in the absence of organ damage, laboratory or imaging findings that indicate a high risk of developing end-organ damage within 2 years [1]. Disease-related organ damage in MM/PCM includes one or more of the following: hypercalcemia, renal insufficiency, anemia, and lytic bone lesions (CRAB). In the 2022 International Consensus Classification of Mature Lymphoid Neoplasms, clinicians on the Clinical Advisory Committee (CAC) strongly favored the term “MM” over “PCM.” [2] Moreover, MM is more commonly used than PCM in the literature. Therefore, we use “MM” rather than “PCM” throughout this review.

In WHO-HAEM5, the morphology of neoplastic PCs ranges from mature-appearing PCs to immature, plasmablastic, and pleomorphic (including multinucleated and multilobated) variants [1]. Immunophenotypically, neoplastic PCs in MM exhibit terminal B-cell differentiation characterized by bright CD38 expression, CD138 and IRF4/MUM1 expression, monotypic light-chain expression, and absent expression of B-cell markers CD19, CD20 and PAX5. In this review, we focus on rare morphological and phenotypic variants and features of MM, using selected examples to highlight the diagnostic challenges.

Morphological challenges

Small-cell variant

The small-cell or lymphoplasmacytic variant of MM has been described in the Revised 4th edition of the WHO Classification of Hematolymphoid Tumours, but not in the WHO-HAEM5 [3]. In the study of 351 MM cases, Heerema-McKenney et al. identified 12 (3.4%) cases of the small-cell variant [4]. Immunophenotypically, the neoplastic PCs usually show strong expression of CD20 and cyclin D1, mimicking mantle cell lymphoma (MCL) because of the common presence of the t(11q13;14q32)(IGH::CCND1) translocation [5–10]. However, this variant is distinguished from B-cell lymphoma by the lack of CD45 and the presence of CD138, and the clinical presentation of MM [4]. Fig. 1 depicts a case of MM with the small-cell variant presenting with a compression fracture of the lumbar vertebra.

Fig. 1.

Small-cell variant of MM presenting with a lumbar spine fracture in a 41-year-old male. A Sheets of tumor cells admixed with blood in a transpedicular bone biopsy (H&E, × 40). B The tumor cells are small, some with a plasmacytoid appearance (H&E; × 1000). The neoplastic cells express CD20 (C), CD138 (D), and cyclin D1 (E, × 400) and exhibit CCND1 rearrangement by dual-color FISH probe (F)

The major diagnostic challenge for the small-cell variant is lymphoplasmacytic lymphoma (LPL), a clonal B-cell neoplasm comprising small lymphocytes with plasmacytoid and/or plasma cell differentiation, a frequent monoclonal IgM paraprotein, and the presence of MYD88 L265P mutation (95%) [11]. The presence of lytic bone lesions, the absence of MYD88 L265P mutation, and a high serum free light-chain ratio would favor the diagnosis of MM. Awareness of this rare small-cell variant of MM would help avoid misdiagnosis and ensure appropriate management. The diagnosis of the small-cell variant of myeloma can be established by correlating the clinical, morphological, and immunophenotypic features as listed in Table 1.

Table 1.

Differential diagnoses of MM of the small-cell variant

	MM, small-cell variant MM	MCL, small-cell variant MCL	Lymphoplasmacytic lymphoma (LPL)
Serum M protein	Present	Absent	Present, usually IgM
CRAB	Present	Absent	Absent
CD5	Negative	Usually positive	Usually negative
CD19	Negative	Positive	Positive
CD45	Usually negative	Positive	Positive
CD117	Positive in one third	Negative	Negative
CD138	Positive	Negative	Only in the plasmacytic component
MUM1	Positive	Sometimes positive	Only in the plasmacytic component
Cyclin D1	Usually positive	Positive	Negative
SOX11	Negative	Positive	Negative
t(11;14)(q13;q32)	Usually present	Present	Absent

CRAB hypercalcemia, renal insufficiency, anemia, and bone lesions

In addition to LPL and MCL mentioned above, variable extents of plasmacytic differentiation have been reported in many types of small B-cell lymphomas, such as marginal zone lymphoma, small lymphocytic lymphoma/chronic lymphocytic leukemia, and follicular lymphoma. In particular, marginal zone lymphoma can exhibit extensive plasmacytic differentiation. This can make it difficult to differentiate it from LPL or PCN. Awareness of plasma cell differentiation in various types of small B-cell lymphomas is important for considering differential diagnoses, particularly in cases of limited needle core biopsy specimens. Correlation of clinical findings, immunophenotypic data, and molecular study results, particularly the MYD88 mutation assay, is needed to reach the correct diagnosis.

Several findings may suggest the possibility of B-cell lymphoma with extensive clonal plasmacytic differentiation. Plasma cells associated with B-cell lymphoma are more frequently CD45 +, CD19 +, CD56-, and surface immunoglobulin light-chain positive [12, 13]. The identification of background clonal B-cells by flow cytometry or ultrasensitive kappa/lambda mRNA by in situ hybridization, as well as atypical B-cells with aberrant CD43 or BCL-2 expression, also suggests the possibility of small B-cell lymphoma with plasmacytic differentiation. Additionally, patients with LPL and MZL often show IgM type M-protein, which is rare in PCN. Furthermore, LPL and MZL usually lack the abnormal FISH findings seen in PCN.

Plasmablastic variant

Based on Wright-stained marrow aspirate smears, Greipp P.R. et al. first defined the plasmablastic morphology of MM as follows: a large nucleus with fine reticular chromatin, a large nucleolus, and no or very little hof region in the less abundant cytoplasm (less than one-half of the nuclear area) [14]. Nuclear immaturity and pleomorphism of PCs are reliable indicators of malignancy [1].

In histological sections, plasmablasts are characterized by large, round, and eccentrically located vesicular nuclei, usually with a prominent nucleolus, and abundant basophilic or amphophilic cytoplasm [15].

Plasmablastic myeloma (PBM) and plasmablastic lymphoma (PBL) exhibit nearly identical cytomorphology and immunophenotypic profile (Table 2), making them impossible to differentiate based purely on pathological features [16]. In general, EBV infection is far more commonly associated with PBL than PBM, with some exceptions. Figure 2 depicts a case of PBM with extensive marrow involvement by clonal plasmablasts in an HIV-infected patient. In our previous studies, we found that, in immunocompetent patients, the plasmablastic variant of both plasmacytoma and MM is more frequently associated with EBV than the non-blastic variants [17]. Furthermore, PBMs in Taiwan frequently presented as extramedullary and extranodal masses mimicking PBL [18]. In these cases, the presence of myeloma-defining features (Table 2), including extensive marrow involvement (≥ 60% plasmablasts), a serum free light-chain ratio of ≥ 100, CRAB, and serum M protein, favors the diagnosis of PBM according to the WHO-HAEM5 [1]. Additionally, FISH for targeted genetic alterations such as del(13q14), gain of chromosome 1q, loss of chromosome 1p, and IGH::FGFR3 may be helpful in differentiating PBM from PBL [18].

Table 2.

Differential diagnostic features of plasmablastic myeloma (PBM) from plasmablastic lymphoma (PBL)

	Plasmablastic myeloma (PBM)	Plasmablastic lymphoma (PBL)
Bone marrow	Clonal bone marrow plasma cell percentage ≥ 60%*	Negative or partially involved
CRAB	Present	Absent
Ratio of serum free light chain	≥ 100	Within normal ranges
M protein	Present	Absent
EBV infection	Rare	Frequent

CRAB hypercalcemia, renal insufficiency, anemia, and lytic bone lesion, M protein, monoclonal paraprotein

^*Plasmablastic myeloma may present initially as extramedullary disease, either with or without marrow involvement, yet exhibiting other myeloma-defining features such as CRAB to be qualified as MM

Fig. 2.

Plasmablastic myeloma occurring in a 41-year-old male infected with HIV. A and B, marrow infiltration by sheets of plasmablasts (A, × 200 and B, × 1000; H&E) expressing CD138 (C, × 400) and MYC (D, × 400). Marrow aspirate (E) and peripheral blood (F) smears show plasmablasts with prominent nucleoli and amphophilic cytoplasm with microvesicles (×1000; Liu stain)

Pleomorphic (anaplastic) variant

Pleomorphic (or anaplastic) myeloma is a very rare morphologic variant of MM, defined as malignant PCs with significant cytological atypia. Only isolated case reports and small case series have been reported [19–22]. In histopathology, anaplastic PCs exhibit anaplastic or pleomorphic nuclei, including multinucleated and multilobated forms. This variant is an aggressive form of MM that may occur at initial presentation or metachronously at relapse from transformation or dedifferentiation of well-differentiated myelomatous PCs [19, 21]. Pleomorphic or anaplastic myeloma usually presents in extramedullary sites and shows a poor response to conventional and novel therapies [19, 21, 23]. Pleomorphic or anaplastic PCs usually coexist with plasmablastic PCs in the same patient. Figure 3 shows a case of MM comprising predominantly plasmablasts in the marrow, whereas the nodal lesion exhibits anaplastic morphology [24]. In the literature, some authors have considered plasmablastic and anaplastic MM to be the same morphological variant [20, 22]. In the authors’ opinion, because plasmablastic and pleomorphic or variant are morphologically distinct and have different differential diagnostic considerations, they should be regarded as separate morphological variants rather than being lumped together into a single variant.

Fig. 3.

Pleomorphic/anaplastic myeloma initially presenting as inguinal lymphadenopathy in a 71-year-old male. A Scanning view shows a diffuse nodal infiltration (H&E, × 12.5). B The neoplastic cells are anaplastic (H&E, × 1000) and express CD56 (C, × 400) and CD138 (D, × 400). The neoplastic PCs are plasmablastic/anaplastic in the marrow aspirate (E; Liu stain) and peripheral blood (F; Liu stain)

Extramedullary presentation of MM

Extramedullary disease (EMD) is defined by the presence of clonal PCs outside the marrow. EMD occurs in 7%–18% of patients with MM and is more frequent in cases with plasmablastic and pleomorphic morphology [18, 25]. EMD is usually a manifestation of advanced disease and has a higher prevalence among patients with relapsed MM [18, 26, 27]. According to a systematic review by Bladé J et al., EMD is defined by soft-tissue plasmacytoma that arises from hematogenous spread and has no contact with bony structures (to exclude direct invasion from the underlying marrow tumor). The sites of EMD vary according to the phase of MM [28]. At diagnosis, EMD is typically found in skin and soft tissues. At relapse, the frequently involved sites include the liver, kidney, lymph nodes, the central nervous system, the breast, the pleura, and the pericardium. Another form of EMD is plasma cell leukemia (PCL), which could occur as either a primary or a secondary disease [29].

Cutaneous involvement by MM with blastic morphology and an aberrant phenotype may pose a significant diagnostic challenge, particularly when differentiating it from PBL. In our recent study of 25 cases of PBM, defined by ≥ 30% plasmablasts in tissue sections, 15 (60%) patients had EMD, which occurred as the initial presentation in nine cases [18]. In our small cohort of patients, the most common extramedullary sites of MM were soft tissue and/or skin (10/15, 67%), followed by the pleura, the lungs, and lymph nodes.

Myelomatous effusion refers to malignant pleural, pericardial, or ascitic effusion caused by neoplastic PCs. Myelomatous effusion usually develops during disease progression in patients with MM, yet it may occur at initial presentation as an ominous sign [30]. The key diagnostic clue lies in the typical cytological features of PCs, such as round nuclei and basophilic or amphophilic cytoplasm with a perinuclear hof. Although neoplastic PCs exhibit a wide cytomorphologic range, these features persist even in multinucleated pleomorphic PCs as depicted in Fig. 4.

Fig. 4.

Myelomatous pleural effusion occurring 13 months after diagnosis of multiple myeloma in a 60-year-old female. A Cellular effusion comprising numerous neoplastic PCs (×200). B Two mesothelial cells among the neoplastic PCs (×1000). C and D Close-up views showing multinucleated neoplastic PCs, still with a perinuclear hof (Liu stain in all panels)

When the neoplastic PCs in myelomatous effusion exhibit plasmablastic morphology, PBM should be differentiated from effusion-associated PBL, primary effusion lymphoma (PEL), and fluid overload-associated large B-cell lymphoma. PEL can be readily distinguished from PBM by its universal association with Kaposi’s sarcoma-associated herpesvirus/human herpesvirus 8 (KSHV/HHV8), a feature not seen in PBM. FO-LBCL often shows expression of the B-cell markers (CD20 and PAX5), which are absent in PBM. Finally, the presence of myeloma-defining features such as M protein and CRAB supports the diagnosis of malignant effusion of MM with plasmablastic morphology [31].

Plasma cell leukemia (PCL) was initially defined by Prof. Robert Kyle in 1974 as requiring both more than 20% circulating PCs and an absolute PC count greater than 2 × 10⁹/L in peripheral blood (PB) [32]. Because of the poor prognosis of PCL, the International Myeloma Working Group redefined the diagnostic criteria in 2021 as the presence of 5% or more circulating PCs in PB in patients otherwise diagnosed with symptomatic MM [33]. This definition is adopted by the WHO-HAEM5, in which primary PCL is considered a clinical variant of MM [1]. In a recent study using multiparameter flow cytometry in newly diagnosed myeloma patients, Jelinek T. et al. found that patients with 2%–20% circulating neoplastic PCs had outcomes comparable to those with primary PCL. Moreover, the patients with 2%–5% circulating neoplastic PCs had outcomes similar to those with 5%–20% circulating neoplastic PCs [34]. Their study suggests that ≥ 2% circulating neoplastic PCs is a biomarker of occult primary PCL and supports the assessment of circulating PCs by flow cytometry during the diagnostic workup of MM.

PCL is considered primary when it presents de novo in patients without evidence of previous MM and the primary form accounts for 60%–70% of PCL cases [35]. In primary PCL, the constellation of adverse biologic prognostic factors in patients with advanced aggressive myeloma is already present at diagnosis. Furthermore, primary PCL has a more aggressive clinical presentation than MM [35]. The prognosis of primary PCL is poor, with a median progression-free survival of only four months, even in the era of proteasome inhibitors and immunomodulatory agents as reported in a cohort of 53 Turkish patients [36]. Fig. 5 depicts an example of primary PCL.

Fig. 5.

Primary plasma cell leukemia in a 59-year-old male. A-D Various morphological features of leukemic PCs in the peripheral blood. E and F, Subsequent marrow study after the diagnosis of PCL confirms MM (all Liu stains)

In the remaining 30%–40% of patients, secondary PCL develops during disease progression or relapse, occurring in around 1% of MM cases during the disease course [33]. In a multicenter retrospective study of 101 patients with secondary PCL, defined by ≥ 20% circulating PCs and an absolute PC count greater than 2 × 10⁹/L in PB, Jurczyszyn A. found that the median time to the development of secondary PCL from initial MM diagnosis was 31months [37]. The prognosis was poor. Of the 72 patients who received treatment, a platelet count ≤ 100 × 10⁹/L at the time of secondary PCL diagnosis was the only independent predictor of a poorer overall survival. They suggested that patients with secondary PCL may benefit from salvage autologous stem cell transplantation and proteasome inhibitor-based therapy [37]. As survival among patients with MM increases, the frequency of secondary PCL is expected to increase. There are some differences in the phenotypic and molecular genetic findings between primary and secondary PCL. For details, readers may refer to the review by Jurczyszyn A. et al [29].

Immunophenotypic challenges

PCs are terminally differentiated B-cells that secrete immunoglobulins. Benign PCs typically express CD19, CD38, and CD138 and exhibit polytypic light-chain expression. In contrast, neoplastic (malignant) PCs are monotypic for light chain expression, with frequent loss of CD19 expression, and may aberrantly express CD56, CD117, or other less common antigens. The following section provides a more detailed discussion of these antigens.

CD45

CD45 is present during the early developmental stages of PCs. The majority of PCNs are negative for CD45, with CD45 expression being reported in up to 14% of MM [38, 39]. Using flow cytometric immunophenotyping, Kumar et al. revealed that the proportion of neoplastic PCs expressing CD45 was higher among those with early disease (MGUS or smoldering MM) compared to those with advanced disease (new or relapsed MM) [39]. Accordingly, CD45 expression in a hematolymphoid neoplasm should not be used to exclude PCNs [24].

CD19 and CD56

As mentioned previously, reactive/benign PCs express CD19. In contrast, neoplastic PCs usually lose CD19 expression (> 95% of cases), making CD19 a relatively reliable marker for differentiating reactive from neoplastic PCs in clinical practice [1, 40].

CD56, an NK cell marker, is expressed in 60%–80% of MM cases [1, 41]. Most MM cells are CD19-/CD56 +, with a smaller proportion showing a CD19 −/CD56 − or CD19 +/CD56 + phenotype [40]. Using flow cytometry, Rawstron AC et al. reported that 65% of patients with MM had a CD19 −/CD56 + phenotype, 30% with CD19 −/CD56(low) phenotype, and 5% had a CD19 +/CD56 + phenotype. These two antigens discriminated myeloma from normal plasma cells, which were all CD19 +/CD56(-/low) [42]. Although extremely rare, neoplastic PCs might be positive for CD19 and negative for CD56 [43].In contrast to typical PCNs, however, PCs in LPL are almost always CD19-positive [44]. PCs with aberrant antigen expression, such as CD56 and CD117, and light-chain restriction should be considered neoplastic [1]. Interestingly, CD56 is usually negative in the leukemic PCs in both primary and secondary PCL [41].

CD117

CD117 (KIT) is a type III receptor tyrosine kinase involved in cell signal transduction in several cell types, including mast cells, some hematopoietic stem cells, germ cells, melanocytes, and Cajal cells of the gastrointestinal tract [45]. Accordingly, CD117 is expressed in neoplasms arising from these cells, including acute myeloid leukemia, melanocytic neoplasms, and various sarcomas, including gastrointestinal stromal tumors [45].

CD117 is negative in benign PCs. PCs with aberrant antigen expression, such as CD117, and light-chain restriction should be considered neoplastic [1]. In myelomatous PCs, the reported expression rates ranged from 3% to one third of cases. [45–49] Marks E et al. reported that CD117 was expressed in PBM (6 of 8 cases) but not in any of the eight cases of PBL, and they suggested that CD117 expression could serve as a useful marker for differentiating PBM from PBL [50]. However, in our recent study of 25 cases of PBM, none of these cases expressed CD117 [18]. In brief, aberrant CD117 expression in PCs is an indicator of plasmacytic neoplasms, and the usefulness of CD117 for differentiating PBM from PBL warrants additional studies. Additionally, CD117 expression together with the absence of CD45 expression may result in the misdiagnosis of extramedullary PBM as granulocytic or myeloid sarcoma.

In addition, the co-expression of CD56 and CD117 has been reported to be of prognostic significance. In a retrospective study of 188 patients with MM using flow cytometric immunophenotyping, Keski H. et al. found that double positivity for CD56 and CD117 was associated with a poorer prognosis. Patients with double positivity also had a lower overall response rate, shorter overall survival, and higher mortality compared to patients expressing either marker alone or neither [51].

CD3

Another antigenic aberrancy in PCN is CD3 expression, which is a relatively specific T-cell antigen. Pan Z. et al. reported a series of 17 cases of B-cell neoplasms expressing CD3. These included 12 cases of PBL and five cases of PBM, all with blastic morphology and occurring at extranodal sites [52]. Similarly, we recently described a case of anaplastic myeloma with nodal presentation and strong and diffuse expression of both CD3 and CD45. The case was initially misdiagnosed as nodal peripheral T-cell lymphoma [24].

CD4

CD4 is a T-cell marker and has been reported to be rarely expressed in myelomatous PCs [53–56]. Aberrant acquisition of T-cell antigens including CD3 and CD4 in PCNs has been reported to be associated with plasmablastic and anaplastic morphology, aggressive behavior, and poor prognosis [54–56]. In cases with CD4 expression particularly with co-expression of CD56, various hematologic malignancies, including blastic plasmacytoid dendritic cell neoplasm and various T- and NK/T-cell lymphomas, may enter the differential diagnosis.

In cases of myeloma with aberrant T-cell antigen expression, comprehensive immunophenotypic evaluation is key to making the correct diagnosis. This evaluation should include additional T-cell markers (CD2, CD5, CD7, cytotoxic markers, T-cell receptor (TCR)-βF1, and TCR-delta) and PC markers (CD38, CD138, MUM1, Blimp1, and XBP1), together with clonality studies (IGH and TCR gene rearrangement), awareness of a history of MM, and the rare phenotypic aberrancy, are key to making the correct diagnosis.

MM is genetically heterogeneous, with two main groups defined by cytogenetics. Approximately 40%–50% of patients show recurrent IGH translocations whereas up to 55% lack IGH translocations and are characterized by hyperdiploidy. A small subset of patients does not fall into either category [1, 2]. Table 3 lists gene fusions relatively specific to MM that can be detected by interphase FISH and their clinical significance. Although these FISH findings are highly informative for the diagnosis of MM, some may also occur in B-cell lymphomas. Therefore, accurate diagnosis requires an integrated approach incorporating histomorphology, immunophenotyping, and molecular findings.

Table 3.

Gene fusions relatively specific for Multiple Myeloma

FISH abnormality	Gene#	Clinical significance
t(4;14)(p16;q32)	FGFR3/NSD2 (MMSET)	High-risk myeloma
t(6;14)(p21;q32)	CCND3	Uncommon
t(11;14)(q13;q32)*	CCND1	Most common translocation (~15%–20%); associated with Cyclin D1 expression and sensitivity to venetoclax
t(14;16)(q32;q23)	MAF	High-risk myeloma
t(14;20)(q32;q12)	MAFB	High-risk myeloma
t(8;14)(q24;q32)**	MYC	Often secondary progression event

^*Also a hallmark fusion of mantle cell lymphoma

^**This is also frequently involved in plasmablastic lymphoma

^#All genes are fused with IGH

Cyclin D1

Cyclin D1 is a critical regulator of the cell cycle in lymphocytes, and its function is essential for normal lymphocyte proliferation and differentiation in response to mitogenic stimuli. Aberrant cyclin D1 expression resulting from the chromosomal translocation t(11;14)(q13;q32) with IGH::CCND1 fusion is a hallmark of mantle cell lymphoma. Cyclin D1 expression by immunohistochemistry has been reported in around one-quarter of MM cases [5, 57, 58]. Using FISH for the 14q32 and 11q13 chromosomal loci in a large cohort of patients with MM, Fonseca R. et al. found that 16% (53/336) of cases had abnormal FISH patterns compatible with the t(11;14)(q13;q32) [59].This unique subset of patients with MM is characterized by cyclin D1 upregulation and a lymphoplasmacytic morphology (small-cell variant) as exemplified in Fig. 1. [59].

In routine diagnostic practice, in a marrow specimen comprising small lymphocytes and PCs (i.e. small-cell variant of PCM), cyclin D1 expression might help lead to the diagnosis of MM. However, an exceptional case of cyclin D1-expressing LPL has been reported [60]. Due to the technical difficulty in immunohistochemistry for light-chain stains, particularly in the marrow trephine, cyclin D1 could also be applied in conjunction with CD19 for differentiating benign vs. mature-appearing neoplastic PCs. The former typically exhibit a phenotype of CD19 +/cyclin D1-, whereas the latter are usually CD19-/cyclin D1 +.

Aberrant expression of cyclin D1 in patients with myeloma should prompt pathologists to perform FISH for t(11;14)(q13;q32)/IGH::CCND1, as the presence of this translocation may identify patients who could benefit from venetoclax-based therapy [61].

EMA and cytokeratin

Epithelial membrane antigen (EMA) is considered a marker of epithelial cells, yet it is also expressed by benign or reactive PCs and in around 60% of MM cases [38]. Cytokeratin has been reported to be expressed in one-quarter to one-third of plasmacytoma/MM cases [38, 62]. Expression of EMA and/or cytokeratin in MM with EMD could easily lead to a diagnosis of metastatic carcinoma, a potential diagnostic pitfall for MM with EMD, particularly those with plasmablastic and pleomorphic/anaplastic morphology.

Association with EBV

MM is generally not associated with Epstein-Barr virus (EBV), as demonstrated by in situ hybridization [17, 18, 50, 63]. The rare EBV-positive myeloma cases occur predominantly in extramedullary sites with a plasmablastic and/or pleomorphic/anaplastic morphology mimicking PBL and more frequently in HIV-infected patients. [17, 50, 63, 64] However, the differential diagnosis between PBM and PBL relies on comprehensive clinicopathological analysis, primarily based on myeloma-defining features, while EBV association alone is not a sufficient distinguishing factor.

Differential diagnostic summaries

Reactive plasmacytosis might mimic PCN, particularly when there is extensive plasmacytosis in the marrow [65, 66]. Reactive plasmacytosis might occur in the PB or BM, and most commonly in patients with (1) infection, (2) angioimmunoblastic T-cell lymphoma (AITL), (3) other malignancies, (4) drug-related, (5) autoimmune diseases and (6) others, according to a systematic literature review [65]. Immunophenotypically, reactive PCs express CD19 and CD138, but not CD56, CD117 or cyclin D1. They are polytypic for light-chain expression, which can be determined by flow cytometry, immunohistochemistry or in situ hybridization. A clonality assay for immunoglobulin gene rearrangement might help differentiate reactive from neoplastic PCs, although this is usually unnecessary because morphology combined with immunophenotyping is often sufficient for diagnosis.

PCNs composed of atypical or immature PCs that otherwise exhibit typical PC features are generally straightforward to diagnose with the aid of flow cytometric immunophenotyping or immunohistochemistry. The major diagnostic challenges in PCNs tend to arise from the two morphological extremes: the small-cell(mature) and the large-cell (plasmablastic and pleomorphic/anaplastic) variants.

Table 1 lists the major differential diagnoses of the small-cell variant of PCN, namely small-cell variant of MCL and LPL. Compared with the small-cell variant of PCN, the neoplastic cells in the small-cell variant of MCL exhibit slightly irregular nuclear contours and usually express CD5 and SOX11, but not CD138. Cyclin D1 expression is not useful in this diagnostic setting. LPL is a marrow-based neoplasm comprising small B cells, plasmacytoid lymphocytes and PCs with an IgM paraprotein [11]. LPL could pose a serious diagnostic challenge when differentiating it from the small-cell variant of PCN. In addition to morphological features, LPL is distinct from the small-cell variant of PCN by immunophenotype (negative for CD19, cyclin D1, and SOX11, and positive for CD45). Genetically, the great majority of LPL cases exhibit the MYD88 L265P mutation, which can also be detected in effusion cytology specimens [67].

For blastic PCNs, the major differential diagnoses include diffuse large B-cell lymphoma (DLBCL) and PBL. DLBCL is distinguished from PBM by the absence of amphophilic cytoplasm and by a B-cell rather than plasmacytic phenotype. The differential between PBL and PBM is particularly challenging because PBM frequently presents as an extramedullary and extranodal mass, mimicking PBL [15, 18]. Please refer to Table 2 for the major differential diagnostic features between PBM and PBL.

For pleomorphic/anaplastic MM, the major differential considerations are anaplastic lymphoma kinase-positive large B-cell lymphoma (ALK + LBCL), primary effusion lymphoma (PEL; including extra-cavitary PEL), and Kaposi’s sarcoma-associated herpesvirus/human herpesvirus 8 (KSHV/HHV8)-positive DLBCL. Table 4 summarizes the major differential diagnostic features. Morphologically, the tumor cells in these three entities might be immunoblastic, plasmablastic, or anaplastic. Phenotypically, ALK + LBCL and PEL express PC markers, whereas KSHV/HHV8-positive DLBCL shows a B-cell phenotype. Moreover, by immunohistochemistry, ALK + LBCL and KSHV/HHV8 + DLBCL are positive for ALK and KSHV/HHV8-latency-associated nuclear antigen (LANA), respectively.

Table 4.

Differential diagnostic considerations of pleomorphic/anaplastic myeloma

	Pleomorphic/anaplastic myeloma	ALK + LBCL	PEL	KSHV/HHV8-positive DLBCL
Clinical features	*Sharing similar features with MM *Either occurring at initial presentation or at relapse from MM *Usually HIV-unrelated *Aggressive behaviour	*No association with immunodeficiency	*Mostly HIV-infected patients *50% patients with KSHV/HHV8 infection-related features (KS and MCD)	*Usually with profound immunodeficiency, most often HIV-positive and HHV8-positive MCD
Sites of involvement	*BM *Extramedullary sites including serous effusion	*Nodal (75%) *Less frequently, extra-nodal	*Pleural, pericardial, and/or peritoneal effusion *No nodal or extranodal mass (except for extracavitary PEL)	*Nodal *Less frequently, extranodal (including spleen)
Morphology	*Pleomorphic *Anaplastic *Multilobulated *Multinucleated	*Immunoblastic, plasmablastic, or anaplastic	*Immunoblastic, plasmablastic, or anaplastic	*Immunoblastic, plasmablastic, or anaplastic
Immunophenotype	*Positive for PC markers	*Positive for PC markers *CD20-negative or variable *ALK-positive	*Positive for PC markers *Negative for B-cell markers	*B-cell phenotype *Usually IgM lambda *Often negative for PC markers
KSHV/HHV8 by LANA	Negative	Negative	Positive	Positive
EBER	Usually negative	Negative	Positive in most HIV-infected patients	Usually Negative

The most commonly used PC markers include CD38, CD138, MUM1, Blimp1 and XBP1

ALK + LBCL anaplastic lymphoma kinase-positive large B-cell lymphoma, DLBCL diffuse large B-cell lymphoma, KS Kaposi’s sarcoma, KSHV/HHV8 KS-associated herpesvirus/human herpesvirus 8, LANA latency-associated nuclear antigen, MCD multicentric Castleman disease, PEL primary effusion lymphoma

Conclusion

The sheer increase in PCs in the marrow is not diagnostic of MM, as reactive plasmacytosis in the marrow may exceed 20% [65]. There is a wide morphological range of PCs in PCNs, and morphology alone is usually not diagnostic. Immunophenotyping is critical in establishing the correct diagnosis. However, phenotypic aberrancies and pitfalls, as discussed above, exist. Detailed morphological evaluation, laboratory assays, appropriate application and interpretation of immunophenotypic markers, molecular studies (clonality and FISH), and clinical correlation, as summarized in Table 5, are mandatory for the correct diagnosis.

Table 5.

Key differential diagnostic features of benign/reactive plasma cells from neoplastic plasma cells in plasma cell neoplasms

	Benign/reactive PCs	Neoplastic PCs/plasma cell neoplasms
Morphology	*Mature *Rarely bi-nucleated	*Mature, immature, plasmablastic, or pleomorphic (anaplastic) *Nuclear convolution, multi-lobulation and multi-nucleation *May have cytoplasmic and intranuclear vacuoles and inclusions (Russel and Dutcher bodies)
Phenotypic	*CD19 + (both surface and cytoplasmic by FCM) *CD19 + (IHC) *Polytypic light chain expression (FCM, IHC, and ISH) *No aberrant antigen express	*Surface CD19-/cytoplasmic CD19 + (by FCM) *CD19- (IHC) *Monotypic light chain expression *Aberrant antigen expression: CD3, CD4, CD56, CD117, cyclin D1
CRAB	Negative	Usually present
M protein	Negative	Usually present
Serum FLC	Normal ranges and kappa/lambda ratio	Involved light chain ≥ 100 mg/L and involved/uninvolved FLC ratio ≥ 100 for MM
FISH	Negative for genetic alterations	Presence of myeloma-associated genetic abnormalities (Table 3)
BCR-GR	Polyclonal	Clonal in the majority of MM cases (70%–90%)

BCR-GR B-cell receptor gene rearrangement, FCM flow cytometric immunophenotyping, FLC free light chain, IHC immunohistochemistry, SH in situ hybridization

Abbreviations

ALK + LBCL

Anaplastic lymphoma kinase-positive large B-cell lymphoma

BM

Bone marrow

EBV

Epstein-Barr virus

DLBCL

Diffuse large B-cell lymphoma

FISH

Fluorescence in situ hybridization

HIV

Human immunodeficiency virus

KSHV/HHV8

Kaposi’s sarcoma-associated herpesvirus/human herpesvirus 8

LPL

Lymphoplasmacytic lymphoma

MCL

Mantle cell lymphoma

MM

Multiple myeloma

MGUS

Monoclonal gammopathy of undetermined significance

PBL

Plasmablastic lymphoma

PBM

Plasmablastic myeloma

PC

Plasma cell

PCL

Plasma cell leukemia

PCM

Plasma cell myeloma

PCN

Plasma cell neoplasm

PEL

Primary effusion lymphoma

TCR

T-cell receptor

Authors’ contributions

SSC conceived the idea and wrote the draft. CCC updated the draft. Both authors approved the final manscript.

Funding

This review received no funding.

Data availability

No datasets were generated or analysed during the current study.

Declarations

Competing interests

The authors declare no competing interests.